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WO2025030165A1 - Pharmaceutical compositions for delivery of herpes simplex virus antigens and related methods - Google Patents

Pharmaceutical compositions for delivery of herpes simplex virus antigens and related methods Download PDF

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Publication number
WO2025030165A1
WO2025030165A1 PCT/US2024/040874 US2024040874W WO2025030165A1 WO 2025030165 A1 WO2025030165 A1 WO 2025030165A1 US 2024040874 W US2024040874 W US 2024040874W WO 2025030165 A1 WO2025030165 A1 WO 2025030165A1
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WIPO (PCT)
Prior art keywords
hsv
polypeptides
antigenic
antigenic fragments
polypeptide
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PCT/US2024/040874
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French (fr)
Inventor
Alptekin GÜLER
Ricardo Sanchez Velazquez
Julia UEBELE
Annette VOGEL
Christina PFAFENROT
Anna Luise Ernst
Stephanie HEIN
Sabrina Hinz
Ugur Sahin
Michael Steven ROONEY
Ekaterina ESAULOVA
Adam ZUIANI
Theresa ADDONA
Asaf PORAN
Scott GOULDING
Alexandra WALLS
John SROUJI
Gavin PALOWITCH
Charles Lefco DULBERGER
Sarah Catharina Dany
Stefan Thomas STRAUSS
Gary H. Cohen
Harvey Friedman
Sita Awasthi
Drew Weissman
Lauren Michelle HOOK
Kevin EGAN
Tina M. Cairns
Doina ATANASIU
Wan Ting SAW
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Biontech SE
University of Pennsylvania Penn
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Biontech SE
University of Pennsylvania Penn
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Publication of WO2025030165A1 publication Critical patent/WO2025030165A1/en
Pending legal-status Critical Current
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K39/12—Viral antigens
    • A61K39/245—Herpetoviridae, e.g. herpes simplex virus
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • A61P31/12—Antivirals
    • A61P31/20—Antivirals for DNA viruses
    • A61P31/22—Antivirals for DNA viruses for herpes viruses
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K2039/51—Medicinal preparations containing antigens or antibodies comprising whole cells, viruses or DNA/RNA
    • A61K2039/53—DNA (RNA) vaccination
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K2039/555—Medicinal preparations containing antigens or antibodies characterised by a specific combination antigen/adjuvant
    • A61K2039/55511—Organic adjuvants
    • A61K2039/55555—Liposomes; Vesicles, e.g. nanoparticles; Spheres, e.g. nanospheres; Polymers
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K2039/57—Medicinal preparations containing antigens or antibodies characterised by the type of response, e.g. Th1, Th2
    • A61K2039/575—Medicinal preparations containing antigens or antibodies characterised by the type of response, e.g. Th1, Th2 humoral response
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K2039/70—Multivalent vaccine
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2710/00—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA dsDNA viruses
    • C12N2710/00011—Details
    • C12N2710/16011—Herpesviridae
    • C12N2710/16611—Simplexvirus, e.g. human herpesvirus 1, 2
    • C12N2710/16634—Use of virus or viral component as vaccine, e.g. live-attenuated or inactivated virus, VLP, viral protein

Definitions

  • Herpes simplex viruses commonly referred to only as herpes, are categorized into two types: herpes simplex virus, type 1 (HSV-1, or oral herpes) and herpes simplex virus, type 2 (HSV-2, or genital herpes).
  • HSV-1 herpes simplex virus
  • HSV-2 herpes simplex virus
  • HSV-1 type 1
  • HSV-2 herpes simplex virus
  • HSV-1 prevalence is understood as being highest in Africa and lowest in the Americas.
  • HSV-2 More women are infected with HSV-2 than men, because sexual transmission of HSV is more efficient from men to women than from women to men. Prevalence of HSV-2 infection was estimated to be highest in Africa, followed by the Americas. Prevalence of HSV-2 was also shown to increase with age, though the highest numbers of people newly-infected have historically been in adolescents. Both HSV-1 and HSV-2 infections are lifelong.
  • technologies e.g., combinations, compositions, methods, etc.
  • the present disclosure provides pharmaceutical compositions (e.g., immunogenic compositions, e.g., vaccines) for delivering herpes simplex virus (HSV) antigen constructs to a subject (e.g., a patient) and related technologies (e.g., methods).
  • pharmaceutical compositions e.g., immunogenic compositions, e.g., vaccines
  • HSV compositions e.g., immunogenic compositions, e.g., vaccines
  • related technologies e.g., methods.
  • HSV glycoprotein C or antigenic portions thereof, HSV glycoprotein D (gD) or antigenic portions thereof, glycoprotein E (gE) or antigenic portions thereof, or combinations thereof can be useful in preventing or treating HSV, e.g., HSV-1, HSV-2, or both, as further disclosed herein.
  • the present disclosure provides, for example, combinations comprising a plurality of polyribonucleotides, wherein the plurality of polyribonucleotides comprises a first set of polyribonucleotides that encode one or more glycoprotein (GP) polypeptides.
  • GP polypeptide comprises an HSV glycoprotein or antigenic portions thereof.
  • polyribonucleotides that encode one or more GP polypeptides provided herein encode one or more of HSV-2 gC, gD, and/or gE or antigenic portions thereof (e.g., in a construct).
  • such polyribonucleotides of a first set of polyribonucleotides can be part of an RNA construct.
  • a polyribonucleotide that encodes a GP polypeptide or RNA construct as described herein can be part of a composition (e.g., a pharmaceutical composition, e.g., an immunogenic composition, e.g., a vaccine).
  • the present disclosure also provides a combination comprising a plurality of polyribonucleotides, wherein the plurality of polyribonucleotides comprises a first set of polyribonucleotides and a second set of polyribonucleotides.
  • a second set of polyribonucleotides encode one or more T-cell string polypeptides.
  • a T-cell string polypeptide comprises one or more HSV T-cell antigens or antigenic portions thereof.
  • such polyribonucleotides of a second set of polyribonucleotides can be part of an RNA construct.
  • a polyribonucleotide that encodes a T-cell string polypeptide or a corresponding RNA construct as described herein can be part of a composition (e.g., a pharmaceutical composition, e.g., an immunogenic composition, e.g., a vaccine).
  • a composition e.g., a pharmaceutical composition, e.g., an immunogenic composition, e.g., a vaccine.
  • FIG.2 has been modified from Ibanez, F.J., et al., “Experimental Dissection of the Lytic Replication Cycles of Herpes Simplex Virus in vitro,” Front Microbiol. 2018; 9: 2406, which is incorporated herein by reference in its entirety.
  • FIGS.3A-3F show HSV-2 gC, gD, or gE expression in HEK293T cells transfected with 0.2 ⁇ g/mL LNP-formulated trivalent nucleoside-modified RNA (modRNA) encoding gC, gD, and gE (drug product, DP).
  • modRNA trivalent nucleoside-modified RNA
  • HSV-2 gC, gD and gE protein were detected by flow cytometry using primary monoclonal mouse antibodies detecting the respective antigen and a secondary fluorescent tagged anti-mouse antibody.
  • Representative data from one experiment showing percentage of gC (FIG.3A), gD (FIG.3B), and gE (FIG.3C) protein-expressing cells and median fluorescence intensities (MFI) (FIG.3D, 3E, and 3F, respectively) of the total HEK293T population are depicted per antigen. Data shown are mean+SD of HEK293T transfections performed in triplicates.
  • FIG.4 shows a schematic overview of a study in guinea pigs investigating a composition candidate against HSV-2.
  • Guinea pigs were immunized IM on day 0 and day 28 with an HSV-2 composition candidate containing total HSV-2 gC/gD/gE RNA at a concentration of 3 ⁇ g, 15 ⁇ g, or PBS control, as outlined in Table 24.
  • the guinea pigs were challenged with a lethal dose of 5 x 10 5 PFU of HSV-2 strain MS (25-fold LD 50 ).
  • FIGS.5A-5C show serum IgG antibody titers observed one month after a 2nd immunization in guinea pigs immunized with an HSV-2 composition candidate described herein.
  • Serum antibody titers were determined by ELISA at day 56, 4 weeks after the second immunization with a composition (“trivalent vaccine”) comprising three polyribonucleotides encoding glycoprotein C (gC), glycoprotein D (gD) and glycoprotein E (gE), respectively.
  • the dose level represents total RNA content of three RNAs encoding for the respective HSV-2 gC, HSV-2 gD and HSV-2 gE antigens in a 1:1:1 ratio.
  • Geometric mean ⁇ 95% CI and individual animal values are shown. P values were calculated by Kruskal-Wallis test.
  • gC2 glycoprotein C from herpes simplex virus-2
  • gD2 glycoprotein D from herpes simplex virus-2
  • gE2 glycoprotein E from herpes simplex virus-2
  • IgG immunoglobulin G
  • RNA ribonucleic acid
  • GMT geometric mean
  • CI confidence interval.
  • FIG.5A administration of the HSV-2 composition candidate induced high IgG antibody titers against each of HSV-2 gC (FIG.5A), HSV-2 gD (FIG.5B) and HSV-2 gE (FIG.5C), with a 15 ⁇ g dose inducing higher titers for gC and gD antigens than a 3 ⁇ g dose.
  • FIGS.6A-6C show vaginal IgG antibody titers in guinea pigs one month after a 2nd immunization with an HSV-2 modRNA composition described herein. Vaginal antibody titers were determined by ELISA at day 56, four weeks after the second immunization with a trivalent composition.
  • the dose level represents total RNA content of three RNAs encoding for the respective HSV-2 gC, HSV-2 gD and HSV-2 gE antigens in a 1:1:1 ratio. Geometric mean ⁇ 95% CI and individual animal values are shown. P values were calculated by Kruskal-Wallis test.
  • gC2 glycoprotein C from herpes simplex virus-2
  • gD2 glycoprotein D from herpes simplex virus-2
  • gE2 glycoprotein E from herpes simplex virus- 2
  • IgG immunoglobulin G
  • RNA ribonucleic acid
  • GMT geometric mean
  • CI confidence interval.
  • FIG.6A shows serum neutralizing antibody titers to HSV-2 in guinea pigs one month after a 2nd immunization with an HSV-2 modified RNA (modRNA) composition described herein.
  • modified RNA modified RNA
  • Neutralizing antibody titers were determined using a serum HSV-2 plaque reduction assay and defined as highest dilution of serum with 5% human complement that reduced the number of HSV-2 plaques by 50%. Samples were collected at day 56, 4 weeks after the second immunization. The dose level represents total RNA content of three RNAs encoding for the respective HSV-2 gC, HSV-2 gD and HSV-2 gE antigens in a 1:1:1 ratio. Geometric mean ⁇ 95% CI and individual animal values are shown. P values were calculated by Mann-Whitney test.
  • FIGS.8A-8C shows weight loss in guinea pigs administered an HSV-2 composition described herein, following HSV-2 viral challenge.
  • RNA ribonucleic acid
  • FIG.9 shows survival of guinea pigs immunized with an HSV-2 composition described herein, up to day 48 after HSV-2 viral challenge. Probability of survival of guinea pigs up to 48 days after lethal intravaginal challenge with HSV-2 at day 60, approximately one month after second immunization with 3 ⁇ g or 15 ⁇ g of trivalent a composition, or PBS.
  • the dose level represents total RNA content of three RNAs encoding for the respective HSV-2 gC, HSV-2 gD and HSV-2 gE antigens in a 1:1:1 ratio.
  • FIGS.10A-10C show individual evaluation of genital disease in guinea pigs administered an RNA composition described herein, up to day 48 after challenge with a lethal intravaginal dose of HSV-2. Results at day 60, approximately one month after the second vaccination with an HSV-2 modified RNA (modRNA) composition are shown.
  • FIG.10A shows the mean number of days with genital disease during this period and
  • FIG.10B shows the mean severity of genital lesions of days with genital disease. Mean ⁇ SEM and individual animal values are shown.
  • FIG.10C shows the mean number of urinary retention days.
  • the dose level represents total RNA content of three RNAs encoding for the respective HSV-2 gC, HSV-2 gD and HSV-2 gE antigens in a 1:1:1 ratio.
  • P values were calculated by Mann-Whitney test. Black circles with red outlines are associated with animals that succumbed after viral challenge in the PBS group.
  • HSV-2 herpes simplex virus-2
  • SEM standard error of the mean
  • PBS phosphate buffered saline
  • gC2 glycoprotein C from herpes simplex virus-2
  • gD2 glycoprotein D from herpes simplex virus-2
  • gE2 glycoprotein E from herpes simplex virus-2
  • RNA ribonucleic acid
  • FIG.11 shows cumulative disease score in guinea pigs administered an RNA composition described herein, up to day 48 after challenge with a lethal intravaginal dose of HSV-2. Results at day 60 are shown, approximately one month after the second vaccination. The mean number of days with genital disease per group is shown over the course of 48 days. The dose level represents total RNA content of three RNAs encoding for the respective gC2, gD2 and gE2 antigens in a 1:1:1 ratio.
  • HSV-2 herpes simplex virus-2
  • PBS phosphate buffered saline
  • gC2 glycoprotein C from herpes simplex virus-2
  • gD2 glycoprotein D from herpes simplex virus-2
  • gE2 glycoprotein E from herpes simplex virus-2
  • RNA ribonucleic acid.
  • FIG.11 administration of 3 ⁇ g of an HSV-2 composition significantly decreased mean cumulative disease days, and administration 15 ⁇ g decreased mean cumulative disease days further still.
  • FIGS.12A-12C show vaginal virus titers in guinea pigs administered an HSV-2 composition described herein, 2 and 4 days after viral challenge.
  • Vaginal HSV-2 titers were determined by plaque assay 2 days (FIG.12A) and 4 days (FIG.12B) after a lethal intravaginal challenge with HSV-2. Results are plotted as means ⁇ SEM and individual animal values. Mean days of genital shedding of HSV-2 DNA were analyzed by PCR and displayed in (FIG.12C). The dose level for the HSV-2 composition represents total RNA content of three RNAs in a 1:1:1 ratio encoding for the respective HSV-2 gC, HSV-2 gD and HSV-2 gE antigens. P values were calculated by Kruskal-Wallis test.
  • FIGS.13A-13B show DNA copy numbers in DRG and spinal cord of guinea pigs administered an HSV-2 composition disclosed herein, on day 48 after viral challenge. DRG and spinal cord HSV-2 DNA copy numbers in guinea pigs on day 48 following viral challenge with a lethal intravaginal dose of HSV-2 were analyzed by qPCR.
  • HSV-2 genome copies in DRG (FIG.13A) and spinal cord (FIG.13B) relative to GAPDH expression at day 48 after viral challenge are shown for immunized animals. Mean ⁇ SEM and individual animal values are shown.
  • the dose level for the HSV-2 composition represents total RNA content of three RNAs in a 1:1:1 ratio encoding for the respective HSV-2 gC, HSV-2 gD and HSV-2 gE antigens. P values were calculated by Mann-Whitney test.
  • FIGS.14A-14I show transfection rates and expression levels in HEK293T cells transfected with RNA encoding HSV-2 gC (gC2), HSV-2 gD (gD2) and HSV-2 gE (gE2) antigens.
  • Cells were transfected with 0.2 ⁇ g/mL modRNA encoding antigens using a commercial transfection reagent. Expression of gC2, gD2 and gE2 protein was detected by flow cytometry using primary monoclonal mouse antibodies detecting the respective antigen and a secondary fluorescent tagged anti-mouse antibody. Representative data from one experiment showing percentage of gC2 (FIG.14A), gD2 (FIG.14D) and gE2 (FIG.14G) protein-expressing cells and median fluorescence intensities (MFI) (FIG.14B, FIG.14E, and FIG.14H respectively) of the total HEK293T population are depicted per antigen.
  • MFI median fluorescence intensities
  • 2138 HSV-2 gE secretory signal and HSV-2 gC antigen (version 4)
  • 2140 HSV-1 gD secretory signal and HSV-2 gC antigen (version 4)
  • 2141 HSV-1 gB secretory signal and HSV-2 gC antigen (version 4)
  • 1876 IL2 secretory signal and HSV-2 gC antigen (version 3).
  • HSV-2 gD secretory signal and HSV-2 gD antigen version 1
  • 1877 HSV-2 gD secretory signal and HSV-2 gD antigen (version 3)
  • 1659 HSV-2 gD secretory signal and HSV-2 gD antigen (version 2)
  • 1660 HSV-2 gD secretory signal and HSV-2 gE antigen (version 2)
  • 2143 HSV-1 gD secretory signal and HSV-2 gE antigen (version 4).
  • FIGS.15A-15F show transfection rates and expression levels in HEK293T cells transfected with RNA encoding HSV-2 gC (gC2), HSV-2 gD (gD2) and HSV-2 gE (gE2) antigens.
  • Cells were transfected with 0.2 ⁇ g/mL modRNA encoding antigens using a commercial transfection reagent.
  • Expression of gC2, gD2 and gE2 protein was detected by flow cytometry using primary monoclonal mouse antibodies detecting the respective antigen and a secondary fluorescent tagged anti-mouse antibody.
  • FIG.15A Representative data from one experiment showing percentage of gC2 (FIG.15A) and gE2 (FIG.15B) protein-expressing cells and median fluorescence intensities of gC2, gD2 and gE2 (MFI) (FIG.15C, FIG.15D, and FIG.15E respectively) of the total HEK293T population are depicted per antigen. Data shown are mean+SD of HEK293T transfections performed in triplicates.
  • FIG.15 depicts conformational changes in HSV glycoprotein B (gB).
  • FIG.17 depicts conservation scores determined for amino acids located at positions along an UL27 consensus sequence.
  • the UL27 open reading frame encodes HSV gB.
  • complete HSV-1 and HSV-2 genomes were downloaded from VIPR database, and HSV-1 strain 17 and HSV-2 strain HG52 were used as reference strains for HSV-1 and HSV-2 respectively.
  • FIGS.18A-18D depict four HSV T-cell string polypeptide constructs, referred to as A) RNA construct 1 (Het 1), B) RNA construct 3 (Het 3), C) RNA construct 5 (Het 5), and D) RNA construct 7 (Het 7).
  • RNA construct 1 (Het 1) includes RL2, RL2, RS1 and UL54 T cell antigenic fragments.
  • RNA construct 3 (Het 3) includes UL29, UL39, UL49, and UL9 T cell antigenic fragments.
  • RNA construct 5 (Het 5) includes UL30, UL40, UL5, and UL52 T cell antigenic fragments.
  • RNA construct 7 (Het 7) includes UL1, UL19, UL21, UL27, UL46, UL47, UL25 and UL48 T cell antigenic fragments.
  • FIG.20 shows flow diagrams of Part A and Part B of Example 34. The flow diagrams show an overview of the protocol for safety and immunigenicity of an HSV RNA-based composition as three IM administrations at Visits 1, 4, and 7 in healthy subjects.
  • FIG.21 shows flow diagram of Part C of Example 34.
  • the flow diagram shows an overview of the protocol for safety and immunogenicity of an HSV RNA-based composition as two IM administrations in subjects with a history of genital herpes.
  • Syringe icons represent composition administration.
  • Swab icons represent 28 day twice daily anogenital swabbing periods with daily symptom diary.
  • HSV Herpes simplex virus
  • IM intramuscular
  • Wk week.
  • FIG.22 shows a dose escalating schema for Part A.
  • FIG.23 depicts four HSV antigen constructs referred to as A) RNA construct 1 (Het 1), B) RNA construct 4 (Het 4), C) RNA construct 6 (Het 6), and D) RNA construct 8 (Het 8).
  • RNA construct 1 (Het 1) includes RL2, RL2, RS1 and UL54 T cell antigenic fragments.
  • RNA construct 4 (Het 4) includes UL9, UL49, UL39, and UL29 T cell antigenic fragments.
  • RNA construct 6 (Het 6) includes UL52, UL5.1, UL5.2, UL40, UL30.1, and T cell antigenic fragments.
  • RNA construct 8 (Het 8) includes UL48, UL25, UL47, UL46, UL27.1, UL27.2, UL21, UL19, and UL1 T cell antigenic fragments. HLA-I peptides were detected by mass spectrometry.
  • FIGS.25A-25B depict a graphic overview of all HLA-I epitopes A) and specific A*02:01 epitopes B) in response to RNA construct 4 (Het 4) (SEQ ID NO: 622).
  • FIGS.26A-26B depict a graphic overview of all HLA-I epitopes A) and specific A*02:01 epitopes B) in response to RNA construct 6 (Het 6) (SEQ ID NO: 624).
  • FIGS.27A-27B depict a graphic overview of all HLA-I epitopes A) and specific A*02:01 epitopes B) in response to RNA construct 8 (Het 8) (SEQ ID NO: 626).
  • FIGS.28A-28B depict antigen specific T cell response in mice.
  • A) data is compared to vehicle control;
  • B) data is compared to DMSO. Mice were immunized with 1 ug of each of RNA construct 1 (Het 1), RNA construct 4 (Het 4), RNA construct 6 (Het 6), and RNA construct 8 (Het 8) (Group 5 in Table 23 Example 37).
  • FIGS.29A-29B depict antigen specific T cell response in mice.
  • A) data is compared to vehicle control
  • B) data is compared to DMSO. Mice were immunized with 1 ug of each of RNA construct 1 (Het 1) and RNA construct 8 (Het 8) (Group 6 in Table 23 Example 37).
  • FIGS.30A-30B depict antigen specific T cell response in mice.
  • A) data is compared to vehicle control;
  • B) data is compared to DMSO. Mice were immunized with 1 ug of each of RNA construct 1 (Het 1), RNA construct 4 (Het 4), and RNA construct 6 (Het 6) (Group 7 in Table 23 Example 37).
  • FIG.31 depicts antigen specific T cell response in mice. Mice received saline and data is compared to DMSO.
  • FIG.32 depicts antigen specific T cell response for each T-cell antigenic fragment. Fragments were classified as having a low curated score if the fragments resulted in ⁇ 100 spots per 1x10 6 cells.
  • FIGS.33A-33B depict T-cell response to the antigenic fragments.
  • FIGS.34A-34B depict T-cell response to the antigenic fragments.
  • FIGS.36A-36B depict A) polyfunctional CD4 T- cell response and B) polyfunctional CD8 T cell response in mice immunized with 1 ug of each of RNA construct 1 (Het 1), RNA construct 4 (Het 4), RNA construct 6 (Het 6), and RNA construct 8 (Het 8) (Group 5; total 4 ug, 1 ug of each RNA construct).
  • FIGS.37A-37D depict A) UL54, B) UL29, C) UL40, and D) UL47-specific polyfunctional CD8 T- cell response.
  • FIG.38 shows an exemplary study design.
  • FIG.39 depicts female A02 mice bodyweight 0-12 days post intravaginally challenge with HSV-2 strain MS (5 ⁇ 10 5 PFU (25 LD 50 )).
  • FIG.40 depicts survival curve for female A02 mice 0-12 days post intravaginally challenge with HSV-2 strain MS (5 ⁇ 10 5 PFU (25 LD 50 )).
  • FIGS.41A-41C depict survival curve of female A02 mice immunized with individual RNA constructs. Mice received A) RNA construct 1 (Het 1) or BNT163; B) RNA construct 4 (Het 4) or RNA construct 6 (Het 6); or C) RNA construct 8 (Het 8. BNT163 was used a control: BNT163 is a trivalent composition comprising RNA encoding a glycoprotein C (gC) antigenic fragment, a glycoprotein D (gD) antigenic fragment, and a glycoprotein E (gE) antigenic fragment.
  • FIGS.42A-42C depict survival curve of female A02 mice immunized with a combination of RNA constructs.
  • FIGS.43A-43B depict survival curves.
  • FIGS.44A-44B depict A) survival curve using criteria including clinical readouts such as hindleg paralysis and B) survival curve using criteria excluding hindleg paralysis as a clinical readout for female A02 mice 0- 12 days post intravaginally challenge with HSV-2 strain MS (1x10 6 PFU HSV-2 (500 LD 50 )) immunized with all four construct: RNA construct 1 (Het 1), RNA construct 4 (Het 4), RNA construct 6 (Het 6), and RNA construct 8 (Het 8) (total 4 ug, 1 ug of each RNA construct).
  • FIGS.45A-45B depict cumulative survival days for A) individual RNA constructs; RNA construct 1 (Het 1), RNA construct 4 (Het 4), RNA construct 6 (Het 6), or RNA construct 8 (Het 8) and B) combination of constructs.
  • BNT163 was used a control: BNT163 is a trivalent composition comprising RNA encoding a glycoprotein C (gC) antigenic fragment, a glycoprotein D (gD) antigenic fragment, and a glycoprotein E (gE) antigenic fragment.
  • FIGS.46A-46B depict cumulative survival days for A) all construct and combination of constructs and B) constructs and combination of constructs showing best and worst cumulative survival.
  • BNT163 was used a control: BNT163 is a trivalent composition comprising RNA encoding a glycoprotein C (gC) antigenic fragment, a glycoprotein D (gD) antigenic fragment, and a glycoprotein E (gE) antigenic fragment.
  • FIGS.47A-47D depict vaginal titers using A+B) plaque assay or C+D) qPCR at A+C) 2 days after challenge or B+D) 4 days after challenge.
  • FIG.48 depicts ten HSV antigen constructs: RNA construct 15 (Het 15); RNA construct 16 (Het 16); RNA construct 17 (Het 17); RNA construct 18 (Het 18); RNA construct 19 (Het 19); RNA construct 20 (Het 20); RNA construct 21 (Het 21); RNA construct 22 (Het 22); RNA construct 23 (Het 23); RNA construct 24 (Het 4).
  • RNA construct 15 and RNA construct 16 are immediate early and late constructs and include RL2.1, UL54, UL47, UL46 and UL21 T cell antigenic fragments.
  • RNA construct 17 and RNA construct 18 are early constructs and include UL29, UL39, UL9, and UL5.1, UL40, and UL30.1 T cell antigenic fragments.
  • RNA construct 19 and RNA construct 20 are immediate early and early constructs and include UL2.1, UL54, UL9, UL39, and UL5.1 T cell antigenic fragments.
  • RNA construct 21 and RNA construct 22 are late and early constructs and include UL47, UL46, UL21, UL5.2, UL40, UL30.1, and UL29 T cell antigenic fragments.
  • RNA construct 23 and RNA construct 24 include UL2.1, U54, UL5.2., UL40, UL47, and UL46 cell antigenic fragments.
  • FIGS.49A-49E depict protein expression of the RNA constructs shown in FIG.80 by pair with proteasome inhibitor.
  • FIGS.51A-51B depict antigen specific T cell response in A02 mice under A) loose saturation ( ⁇ 220 spots per 1x10 6 cells) and B) under stringent saturation ( ⁇ 100 spots per 1x10 6 cells). Mice were immunized with 2 ug of RNA construct 15 (Het 15) and 2 ug of RNA construct 17 (Het 17) (Group 1 in Table 24 Example 39).
  • FIGS.52A-52B depict antigen specific T cell response in A02 mice under A) loose saturation ( ⁇ 220 spots per 1x10 6 cells) and B) under stringent saturation ( ⁇ 100 spots per 1x10 6 cells). Mice were immunized with 2 ug of RNA construct 20 (Het 20) and 2 ug of RNA construct 22 (Het 22) (Group 2 in Table 24 Example 39).
  • FIGS.53A-53B depict antigen specific T cell response in A02 mice under A) loose saturation ( ⁇ 220 spots per 1x10 6 cells) and B) under stringent saturation ( ⁇ 100 spots per 1x10 6 cells).
  • FIG.54 depicts antigen specific T cell response in Balb/c mice immunized with 2 ug of RNA construct 15 (Het 15) and 2 ug of RNA construct 17 (Het 17) (Group 1 in Table 24 Example 39).
  • FIG.55 depicts antigen specific T cell response in Balb/c mice immunized with 2 ug of RNA construct 20 (Het 20) and 2 ug of RNA construct 22 (Het 22) (Group 2 in Table 24 Example 39).
  • FIG.56 depicts antigen specific T cell response in Balb/c mice immunized with 4 ug of RNA construct 23 (Het 23) (Group 3 in Table 24 Example 39).
  • FIGS.57A-57C depict CD4 and CD8 T-cell responses in mice immunized with 2 ug of RNA construct 15 (Het 15) and 2 ug of RNA construct 17 (Het 17) (Group 1 in Table 24 Example 39).
  • FIGS.58A-58B depict CD4 and CD8 T-cell responses in mice immunized with 2 ug of RNA construct 15 (Het 15) and 2 ug of RNA construct 17 (Het 17) (Group 1 in Table 24 Example 39).
  • FIGS.60A-60B depict CD4 and CD8 T-cell response in mice immunized with 4 ug of RNA construct 23 (Het 23) (Group 3 in Table 24 Example 39).
  • FIG.61 Immunized mice (Day 0 and Day 21) were injected with medroxyprogesterone (subcutaneous injection 2mg/mouse at Day 46) and challenged intravaginally with HSV-2 strain MS (5x10 3 PFU HSV- 2 ( ⁇ 10xLD 50 )) at Day 51 and monitored for survival, genital disease scoring, weight and vaginal virus titers at 6-h, days 2, 4 and 7.
  • FIGS.62A-62E depict female mice bodyweight 0-16 days post intravaginally challenge with HSV- 2 strain MS (5x10 3 PFU HSV-2 ( ⁇ 10xLD 50 )). Mice were administered with A) PBS (Group 1 in Table 25 Example 40); B) 4 ug of RNA construct 23 (Het 23) (Group 2 in Table 25 Example 40); C) with 2 ug of RNA construct 15 (Het 15) and 2 ug of RNA construct 17 (Het 17) (Group 3 in Table 25 Example 40); D) a trivalent (BNT163) composition comprising RNA encoding a glycoprotein C (gC) antigenic fragment, a glycoprotein D (gD) antigenic fragment, and a glycoprotein E (gE) antigenic fragment; and E) combined graph.
  • A) PBS Group 1 in Table 25 Example 40
  • B) 4 ug of RNA construct 23 (Het 23) Group 2 in Table 25 Example 40
  • FIG.63 depicts survival for female mice 0-15 days post intravaginally challenge with HSV-2 strain MS (5x10 3 PFU HSV-2 ( ⁇ 10xLD 50 )).
  • FIGS.64A-64B depict effect of RNA construct immunization on vaginal disease following HSV-2 infection.
  • FIG.65 depicts vaginal HSV-2 replication kinetics over 7 days.
  • FIG.66 depicts cumulative survival for female mice 0-15 days post intravaginally challenge with HSV-2 strain MS (5x10 3 PFU HSV-2 ( ⁇ 10xLD 50 )).
  • FIGS.67A-67B depict the effect of RNA construct immunizations on HSV-2 replication in mice vaginal cavity.
  • FIGS.68A-68B depict the effect of RNA construct immunizations on HSV-2 replication in mice vaginal cavity. A) day 6-h post challenge; and B) day 7 post challenge.
  • FIG.69 depicts female mice bodyweight 0-16 days post intravaginally challenge with HSV-2 strain MS (5x10 3 PFU HSV-2 ( ⁇ 10xLD 50 )).
  • BNT163 a trivalent composition comprising RNA encoding a glycoprotein C (gC) antigenic fragment, a glycoprotein D (gD) antigenic fragment, and a glycoprotein E (gE) antigenic fragment, .
  • TCS-23 RNA construct 23 (Het 23);
  • TCS 15+17 RNA constructs 15+17 (Het 15+ 17).
  • FIG.70 depicts survival for female mice 0-15 days post intravaginally challenge with HSV-2 strain MS (5x10 3 PFU HSV-2 ( ⁇ 10xLD 50 )).
  • FIGS.71A-71B depict effect of RNA construct immunization on vaginal disease following HSV-2 infection.
  • FIGS.72A-72D depict the effect of RNA construct immunizations on HSV-2 replication kinetics in mice vaginal cavity.
  • BNT163 a trivalent composition comprising RNA encoding a glycoprotein C (gC) antigenic fragment, a glycoprotein D (gD) antigenic fragment, and a glycoprotein E (gE) antigenic fragment
  • FIGS.73A-73F depict expression levels in HEK293T cells transfected with RNA encoding HSV-2 gC (gC2), gD (gD2) and gE (gE2) antigens.
  • FIGS.74A-74F depict expression levels in HEK293T cells transfected with RNA encoding HSV-2 gC (gC2) antigens.
  • FIGS.75A-75B depict expression levels in HEK293T cells transfected with RNA encoding HSV-2 gD (gD2) antigens.
  • FIGS.76A-76D depict expression levels in HEK293T cells transfected with RNA encoding HSV-2 gE (gE2) antigens.
  • FIGS.77A-77C depict secretion levels of HEK293T cells transfected with RNA encoding HSV-2 encoding HSV-2 gC (gC2), gD (gD2) and gE (gE2) antigens.
  • FIG.78 shows cumulative recurrent genital lesion days per group.
  • FIG.79 shows days with recurrent genital lesions for each animal starting 1 day after the second immunization. The same animals as in FIG.78 are shown here.
  • FIG.80 shows cytokine production by CD4+ T cell in response to compositions comprising HSV-2 glycoproteins.
  • Mice were immunized twice as described herein with 10 ⁇ g of an exemplary immunogenic fragment of HSV-2 gB (gB2) RNA-LNP. Splenocytes from these mice were stimulated with a gB2 overlapping peptide pool. CD4+ cytokine-producing T cells were analyzed by flow cytometry.
  • FIG.81 shows cytokine production by CD8+ T cell in response to compositions comprising HSV-2 glycoproteins.
  • FIG.82 depicts expression levels in HEK293T cells transfected with RNA encoding HSV-2 gB (gB2) antigens.
  • FIGS.83A-83B show recurrent genital lesion days per group.
  • Guinea pigs were immunized twice on days 25 and 65 post-infection with nucleoside modified RNAs encapsulated in a lipid nanoparticle and expressing an exemplary HSV-2 gE (gE2) immunogenic fragment and an exemplary HSV-2 gI (gI2) immunogenic fragment (15ug each), or PBS (control). Animals were scored daily Monday to Friday for recurrent genital lesions from 1 day after the first immunization until the end of the study on day 116. From the time of the second immunization, significant differences appeared comparing guinea pigs immunized with modified RNAs encoding exemplary gE2 immunogenic fragment and an exemplary gI2 immunogenic fragment, with the PBS group.
  • FIG.83A shows the cumulative recurrent genital lesion days per group.
  • FIG.83B shows days with recurrent genital lesions for each animal starting 1 day after the second immunization. P values were calculated by the two-tailed Mann Whitney test and demonstrate highly significant differences comparing gE2/gI2 with PBS (**, P ⁇ 0.01; ns, P value not significant).
  • FIGS.84A-84D show T cell responses to gE2 stimulation in mice immunized with gE2/gI2 bivalent RNA composition in CD4+ T cells (FIG.84A) and in CD8+ T cells (FIG.84B); and T cell responses to gI2 stimulation in mice immunized with E2/gI2 bivalent RNA composition in CD4+ T cells (FIG.84C) and in CD8+ T cells (FIG.84D).
  • FIG.85 shows antibody responses to gE2/gI2 in mice immunized with gE2 RNA composition.
  • FIGS.86A-86C show antibody responses to gE2/gI2 (FIG.86A), gE2 (FIG.86B), and gI2 (FIG. 86C) in mice immunized with gE2/gI2 bivalent RNA composition.
  • FIG.87 shows a scheme of the protocol of the mice model experiments described in Example 51
  • FIGS.88A-88C show survival (FIG.88A), weight loss (FIG.88B), and disease score (FIG.88C) in a mouse model of HSV-2 in mice immunized with gE2/gI2 bivalent RNA
  • FIGS.89A-89C show HSV virus titer 2 days (FIG.89A) and 4 days (FIG.89B) after infection in mice immunized with gE2/gI2 bivalent; as well as HSV-2 DNA copy number in DRG 28 days after infection (FIG. 89C).
  • FIGS.91A-91D show the prophylactic effect of BNT163, gB, gE2/gI2, or a combination of them on survival (FIG.91A), disease severity (FIG.91B), genital lesions (FIG.91C), and urinary retention (FIG.91D) in guinea pigs infected with HSV-2.
  • FIGS.92A-92B show the therapeutic effect of BNT163, gB and gE2/gI2 on recurrent lesions in a first (FIG.92A) and the combined results of two experiments (FIG.92B) experiments.
  • the term “about” may encompass a range of values that within 25%, 20%, 19%, 18%, 17%, 16%, 15%, 14%, 13%, 12%, 11%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, or less of the referred value.
  • Agent may refer to a physical entity or phenomenon. In some embodiments, an agent may be characterized by a particular feature and/or effect.
  • an agent may be a compound, molecule, or entity of any chemical class including, for example, a small molecule, polypeptide, nucleic acid, saccharide, lipid, metal, or a combination or complex thereof.
  • the term “agent” may refer to a compound, molecule, or entity that comprises a polymer. In some embodiments, the term may refer to a compound or entity that comprises one or more polymeric moieties. In some embodiments, the term “agent” may refer to a compound, molecule, or entity that is substantially free of a particular polymer or polymeric moiety. In some embodiments, the term may refer to a compound, molecule, or entity that lacks or is substantially free of any polymer or polymeric moiety.
  • amino acid refers to a compound and/or substance that can be, is, or has been incorporated into a polypeptide chain, e.g., through formation of one or more peptide bonds.
  • an amino acid has the general structure H 2 N–C(H)(R)–COOH.
  • an amino acid is a naturally-occurring amino acid.
  • an amino acid is a non-natural amino acid; in some embodiments, an amino acid is a D-amino acid; in some embodiments, an amino acid is an L-amino acid.
  • Standard amino acid refers to any of the twenty standard L-amino acids commonly found in naturally occurring peptides.
  • Nonstandard amino acid refers to any amino acid, other than the standard amino acids, regardless of whether it is prepared synthetically or obtained from a natural source.
  • an amino acid, including a carboxy- and/or amino-terminal amino acid in a polypeptide can contain a structural modification as compared with the general structure above.
  • an amino acid may be modified by methylation, amidation, acetylation, pegylation, glycosylation, phosphorylation, and/or substitution (e.g., of the amino group, the carboxylic acid group, one or more protons, and/or the hydroxyl group) as compared with the general structure.
  • such modification may, for example, alter the circulating half-life of a polypeptide containing the modified amino acid as compared with one containing an otherwise identical unmodified amino acid.
  • such modification does not significantly alter a relevant activity of a polypeptide containing the modified amino acid, as compared with one containing an otherwise identical unmodified amino acid.
  • the term “amino acid” may be used to refer to a free amino acid; in some embodiments it may be used to refer to an amino acid residue of a polypeptide.
  • Antibody agent refers to an agent that specifically binds to a particular antigen. In some embodiments, the term encompasses a polypeptide or polypeptide complex that includes immunoglobulin structural elements sufficient to confer specific binding.
  • an antibody agent is or comprises a polypeptide whose amino acid sequence includes one or more structural elements recognized by those skilled in the art as a complementarity determining region (CDR); in some embodiments an antibody agent is or comprises a polypeptide whose amino acid sequence includes at least one CDR (e.g., at least one heavy chain CDR and/or at least one light chain CDR) that is substantially identical to one found in a reference antibody. In some embodiments an included CDR is substantially identical to a reference CDR in that it is either identical in sequence or contains between 1-5 amino acid substitutions as compared with the reference CDR.
  • CDR complementarity determining region
  • an included CDR is substantially identical to a reference CDR in that it shows at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with the reference CDR. In some embodiments an included CDR is substantially identical to a reference CDR in that it shows at least 96%, 96%, 97%, 98%, 99%, or 100% sequence identity with the reference CDR. In some embodiments an included CDR is substantially identical to a reference CDR in that at least one amino acid within the included CDR is deleted, added, or substituted as compared with the reference CDR but the included CDR has an amino acid sequence that is otherwise identical with that of the reference CDR.
  • an included CDR is substantially identical to a reference CDR in that 1-5 amino acids within the included CDR are deleted, added, or substituted as compared with the reference CDR but the included CDR has an amino acid sequence that is otherwise identical to the reference CDR. In some embodiments an included CDR is substantially identical to a reference CDR in that at least one amino acid within the included CDR is substituted as compared with the reference CDR but the included CDR has an amino acid sequence that is otherwise identical with that of the reference CDR. In some embodiments an included CDR is substantially identical to a reference CDR in that 1-5 amino acids within the included CDR are deleted, added, or substituted as compared with the reference CDR but the included CDR has an amino acid sequence that is otherwise identical to the reference CDR.
  • an antibody agent is or comprises a polypeptide whose amino acid sequence includes structural elements recognized by those skilled in the art as an immunoglobulin variable domain.
  • an antibody agent is a polypeptide protein having a binding domain which is homologous or largely homologous to an immunoglobulin-binding domain.
  • an antibody agent may be or comprise a polyclonal antibody preparation.
  • an antibody agent may be or comprise a monoclonal antibody preparation.
  • an antibody agent may include one or more constant region sequences that are characteristic of a particular organism, such as a camel, human, mouse, primate, rabbit, rat; in many embodiments, an antibody agent may include one or more constant region sequences that are characteristic of a human.
  • an antibody agent may include one or more sequence elements that would be recognized by one skilled in the art as a humanized sequence, a primatized sequence, a chimeric sequence, etc.
  • an antibody agent may be a canonical antibody (e.g., may comprise two heavy chains and two light chains).
  • an antibody agent may be in a format selected from, but not limited to, intact IgA, IgG, IgE or IgM antibodies; bi- or multi- specific antibodies (e.g., Zybodies®, etc); antibody fragments such as Fab fragments, Fab’ fragments, F(ab’)2 fragments, Fd’ fragments, Fd fragments, and isolated CDRs or sets thereof; single chain Fvs; polypeptide-Fc fusions; single domain antibodies (e.g., shark single domain antibodies such as IgNAR or fragments thereof); cameloid antibodies; masked antibodies (e.g., Probodies®); Small Modular ImmunoPharmaceuticals (“SMIPs TM” ); single chain or Tandem diabodies (TandAb®); VHHs; Anticalins®; Nanobodies® minibodies; BiTE®s; ankyrin repeat proteins or DARPINs®; Avimers®; DARTs; TCR-like antibodies;, Adnectins
  • an antibody may lack a covalent modification (e.g., attachment of a glycan) that it would have if produced naturally.
  • an antibody may contain a covalent modification (e.g., attachment of a glycan, a payload (e.g., a detectable moiety, a therapeutic moiety, a catalytic moiety, etc.), or other pendant group (e.g., poly-ethylene glycol, etc.)).
  • Antigen refers to a molecule that is recognized by the immune system, e.g., in some embodiments the adaptive immune system, such that it elicits an antigen-specific immune response.
  • an antigen-specific immune response may be or comprise generation of antibodies and/or antigen-specific T cells.
  • an antigen is a peptide or polypeptide that comprises at least one epitope against which an immune response can be generated.
  • an antigen is presented by cells of the immune system such as antigen presenting cells like dendritic cells or macrophages.
  • an antigen or a processed product thereof such as a T-cell epitope is bound by a T- or B-cell receptor, or by an immunoglobulin molecule such as an antibody. Accordingly, an antigen or a processed product thereof may react specifically with antibodies or T lymphocytes (T cells).
  • an antigen is a parasitic antigen.
  • an antigen may be delivered by RNA molecules as described herein.
  • a peptide or polypeptide antigen can be 2-100 amino acids, including for example, 5 amino acids, 10 amino acids, 15 amino acids, 20 amino acids, 25 amino acids, 30 amino acids, 35 amino acids, 40 amino acids, 45 amino acids, or 50 amino acids in length.
  • a peptide or polypeptide antigen can be greater than 50 amino acids. In some embodiments, a peptide or polypeptide antigen can be greater than 100 amino acids.
  • an antigen is recognized by an immune effector cell. In some embodiments, an antigen if recognized by an immune effector cell is able to induce in the presence of appropriate co-stimulatory signals, stimulation, priming and/or expansion of the immune effector cell carrying an antigen receptor recognizing the antigen. In the context of the embodiments of the present disclosure, in some embodiments, an antigen can be presented or present on the surface of a cell, e.g., an antigen presenting cell.
  • an antigen is presented by a diseased cell such as a virus-infected cell.
  • an antigen receptor is a TCR which binds to an epitope of an antigen presented in the context of MHC.
  • binding of a TCR when expressed by T cells and/or present on T cells to an antigen presented by cells such as antigen presenting cells results in stimulation, priming and/or expansion of said T cells.
  • binding of a TCR when expressed by T cells and/or present on T cells to an antigen presented on diseased cells results in cytolysis and/or apoptosis of the diseased cells, wherein said T cells preferably release cytotoxic factors, e.g., perforins and granzymes.
  • Two events or entities are “associated” with one another, as that term is used herein, if the presence, level, degree, type and/or form of one is correlated with that of the other.
  • a particular entity e.g., poypeptide, genetic signature, metabolite, microbe, etc.
  • two or more entities are physically “associated” with one another if they interact, directly or indirectly, so that they are and/or remain in physical proximity with one another.
  • two or more entities that are physically associated with one another are covalently linked to one another; in some embodiments, two or more entities that are physically associated with one another are not covalently linked to one another but are non- covalently associated, for example by means of hydrogen bonds, van der Waals interaction, hydrophobic interactions, magnetism, and combinations thereof.
  • binding typically refers to a non-covalent association between or among entities or moieties. In some embodiments, binding data are expressed in terms of “IC50”.
  • IC50 is the concentration of an assessed agent in a binding assay at which 50% inhibition of binding of reference agent known to bind the relevant binding partner is observed.
  • assays are run under conditions in which the assays are run (e.g., limiting binding target and reference concentrations), these values approximate K D values.
  • Assays for determining binding are well known in the art and are described in detail, for example, in PCT publications WO 94/20127 and WO 94/03205, and other publications such Sidney et al., Current Protocols in Immunology 18.3.1 (1998); Sidney, et al., J. Immunol. 154:247 (1995); and Sette, et al., Mol. Immunol.
  • binding can be expressed relative to binding by a reference standard peptide.
  • a reference standard peptide For example, can be based on its IC 50 , relative to the IC 50 of a reference standard peptide.
  • Binding can also be determined using other assay systems including those using: live cells (e.g., Ceppellini et al., Nature 339:392 (1989); Christnick et al., Nature 352:67 (1991); Busch et al., Int. Immunol.2:443 (1990); Hill et al., J. Immunol.147:189 (1991); del Guercio et al., J. Immunol.
  • Cap refers to a structure comprising or essentially consisting of a nucleoside-5 '-triphosphate that is typically joined to a 5'-end of an uncapped RNA (e.g., an uncapped RNA having a 5'- diphosphate).
  • a cap is or comprises a guanine nucleotide.
  • a cap is or comprises a naturally-occurring RNA 5’ cap, including, e.g., but not limited to a 7- methylguanosine cap, which has a structure designated as “m7G.”
  • a cap is or comprises a synthetic cap analog that resembles an RNA cap structure and possesses the ability to stabilize RNA if attached thereto, including, e.g., but not limited to anti-reverse cap analogs (ARCAs) known in the art).
  • ARCAs anti-reverse cap analogs
  • a capped RNA may be obtained by in vitro capping of RNA that has a 5' triphosphate group or RNA that has a 5' diphosphate group with a capping enzyme system (including, e.g., but not limited to vaccinia capping enzyme system or Saccharomyces cerevisiae capping enzyme system).
  • a capped RNA can be obtained by in vitro transcription (IVT) of a single- stranded DNA template in the presence of a dinucleotide or trinucleotide cap analog.
  • Cell-mediated immunity “Cell-mediated immunity,” “cellular immunity,” “cellular immune response,” or similar terms are meant to include a cellular response directed to cells characterized by expression of an antigen, in particular characterized by presentation of an antigen with class I or class II MHC.
  • a cellular response relates to immune effector cells, in particular to T cells or T lymphocytes which act as either “helpers” or “killers.”
  • the helper T cells also termed CD4 + T cells or CD4 T cells
  • the killer cells also termed cytotoxic T cells, cytolytic T cells, CD8 + T cells, CD8 T cells, or CTLs
  • kill diseased cells such as virus-infected cells, preventing the production of more diseased cells.
  • co-administration refers to use of a pharmaceutical composition (e.g., immunogenic composition, e.g., vaccine) described herein and an additional therapeutic agent.
  • a pharmaceutical composition e.g., immunogenic composition, e.g., vaccine
  • an additional therapeutic agent may be performed concurrently or separately (e.g., sequentially in any order).
  • a pharmaceutical composition e.g., immunogenic composition, e.g., vaccine
  • an additional therapeutic agent may be combined in one pharmaceutically- acceptable carrier, or they may be placed in separate carriers and delivered to a target cell or administered to a subject at different times.
  • Codon-optimized refers to alteration of codons in a coding region of a nucleic acid molecule to reflect the typical codon usage of a host organism without preferably altering the amino acid sequence encoded by the nucleic acid molecule.
  • coding regions are codon-optimized for optimal expression in a subject to be treated using the RNA molecules described herein.
  • codon-optimization may be performed such that codons for which frequently occurring tRNAs are available are inserted in place of “rare codons.”
  • codon- optimization may include increasing guanosine/cytosine (G/C) content of a coding region of RNA described herein as compared to the G/C content of the corresponding coding sequence of a wild type RNA, wherein the amino acid sequence encoded by the RNA is preferably not modified compared to the amino acid sequence.
  • G/C guanosine/cytosine
  • Combination therapy refers to those situations in which a subject is simultaneously exposed to two or more therapeutic regimens (e.g., two or more therapeutic agents).
  • the two or more regimens may be administered simultaneously; in some embodiments, such regimens may be administered sequentially (e.g., all “doses” of a first regimen are administered prior to administration of any doses of a second regimen); in some embodiments, such agents are administered in overlapping dosing regimens.
  • “administration” of combination therapy may involve administration of one or more agent(s) or modality(ies) to a subject receiving the other agent(s) or modality(ies) in the combination.
  • Comparable refers to two or more agents, entities, situations, sets of conditions, etc., that may not be identical to one another but that are sufficiently similar to permit comparison there between so that one skilled in the art will appreciate that conclusions may reasonably be drawn based on differences or similarities observed.
  • comparable sets of conditions, circumstances, individuals, or populations are characterized by a plurality of substantially identical features and one or a small number of varied features.
  • corresponding to may be used to designate the position/identity of a structural element in a compound or composition relative to another compound or composition (e.g., to an appropriate reference compound or composition).
  • a monomeric residue in a polymer e.g., an amino acid residue in a polypeptide or a nucleic acid residue in a polynucleotide
  • corresponding to a residue in an appropriate reference polymer.
  • residues in a polypeptide are often designated using a canonical numbering system based on a reference related polypeptide, so that an amino acid “corresponding to” a residue at position 190, for example, need not actually be the 190 th amino acid in a particular amino acid chain but rather corresponds to the residue found at 190 in the reference polypeptide; those of ordinary skill in the art readily appreciate how to identify “corresponding” amino acids.
  • sequence alignment strategies including software programs such as, for example, BLAST, CS-BLAST, CUSASW++, DIAMOND, FASTA, GGSEARCH/GLSEARCH, Genoogle, HMMER, HHpred/HHsearch, IDF, Infernal, KLAST, USEARCH, parasail, PSI-BLAST, PSI-Search, ScalaBLAST, Sequilab, SAM, SSEARCH, SWAPHI, SWAPHI-LS, SWIMM, or SWIPE that can be utilized, for example, to identify “corresponding” residues in polypeptides and/or nucleic acids in accordance with the present disclosure.
  • software programs such as, for example, BLAST, CS-BLAST, CUSASW++, DIAMOND, FASTA, GGSEARCH/GLSEARCH, Genoogle, HMMER, HHpred/HHsearch, IDF, Infernal, KLAST, USEARCH, parasail, PSI-BLAST, PSI-Search, Scala
  • corresponding to may be used to describe an event or entity that shares a relevant similarity with another event or entity (e.g., an appropriate reference event or entity).
  • a gene or protein in one organism may be described as “corresponding to” a gene or protein from another organism in order to indicate, in some embodiments, that it plays an analogous role or performs an analogous function and/or that it shows a particular degree of sequence identity or homology, or shares a particular characteristic sequence element.
  • amino acid sequence “derived from” a designated amino acid sequence (peptide or polypeptide) “derived from” a designated amino acid sequence (peptide or polypeptide), it refers to a structural analogue of a designated amino acid sequence.
  • an amino acid sequence which is derived from a particular amino acid sequence has an amino acid sequence that is identical, essentially identical or homologous to that particular sequence or a fragment thereof.
  • Amino acid sequences derived from a particular amino acid sequence may be variants of that particular sequence or a fragment thereof.
  • the antigens suitable for use herein may be altered such that they vary in sequence from the naturally occurring or native sequences from which they were derived, while retaining the desirable activity of the native sequences.
  • the term “designed” refers to an agent (i) whose structure is or was selected by the hand of man; (ii) that is produced by a process requiring the hand of man; and/or (iii) that is distinct from natural substances and other known agents.
  • Dosing regimen may be used to refer to a set of unit doses (typically more than one) that are administered individually to a subject, typically separated by periods of time.
  • a given therapeutic agent has a recommended dosing regimen, which may involve one or more doses.
  • a dosing regimen comprises a plurality of doses each of which is separated in time from other doses.
  • individual doses are separated from one another by a time period of the same length; in some embodiments, a dosing regimen comprises a plurality of doses and at least two different time periods separating individual doses.
  • all doses within a dosing regimen are of the same unit dose amount. In some embodiments, different doses within a dosing regimen are of different amounts. In some embodiments, a dosing regimen comprises a first dose in a first dose amount, followed by one or more additional doses in a second dose amount different from the first dose amount. In some embodiments, a dosing regimen comprises a first dose in a first dose amount, followed by one or more additional doses in a second dose amount same as the first dose amount. In some embodiments, a dosing regimen is correlated with a desired or beneficial outcome when administered across a relevant population (i.e., is a therapeutic dosing regimen).
  • Encode refers to sequence information of a first molecule that guides production of a second molecule having a defined sequence of nucleotides (e.g., mRNA) or a defined sequence of amino acids.
  • a DNA molecule can encode an RNA molecule (e.g., by a transcription process that includes a DNA-dependent RNA polymerase enzyme).
  • An RNA molecule can encode a polypeptide (e.g., by a translation process).
  • a gene, a cDNA, or an RNA molecule encodes a polypeptide if transcription and translation of RNA corresponding to that gene produces the polypeptide in a cell or other biological system.
  • a coding region of an RNA molecule encoding a target antigen refers to a coding strand, the nucleotide sequence of which is identical to the RNA sequence of such a target antigen.
  • a coding region of an RNA molecule encoding a target antigen refers to a non-coding strand of such a target antigen, which may be used as a template for transcription of a gene or cDNA.
  • Engineered refers to the aspect of having been manipulated by the hand of man.
  • a polynucleotide is considered to be “engineered” when two or more sequences that are not linked together in that order in nature are manipulated by the hand of man to be directly linked to one another in the engineered polynucleotide and/or when a particular residue in a polynucleotide is non-naturally occurring and/or is caused through action of the hand of man to be linked with an entity or moiety with which it is not linked in nature.
  • Epitope refers to a moiety that is specifically recognized by an immunoglobulin (e.g., antibody or receptor) binding component.
  • an epitope may be recognized by a T cell, a B cell, or an antibody.
  • an epitope is comprised of a plurality of chemical atoms or groups on an antigen.
  • such chemical atoms or groups are surface-exposed when the antigen adopts a relevant three-dimensional conformation.
  • such chemical atoms or groups are physically near to each other in space when the antigen adopts such a conformation.
  • an epitope of an antigen may include a continuous or discontinuous portion of the antigen.
  • an epitope is or comprises a T cell epitope.
  • an epitope may have a length of about 5 to about 30 amino acids, or about 10 to about 25 amino acids, or about 5 to about 15 amino acids, or about 5 to 12 amino acids, or about 6 to about 9 amino acids.
  • a gene product can be a transcript.
  • a gene product can be a polypeptide.
  • expression of a nucleic acid sequence involves one or more of the following: (1) production of an RNA template from a DNA sequence (e.g., by transcription); (2) processing of an RNA transcript (e.g., by splicing, editing, etc.); (3) translation of an RNA into a polypeptide or protein; and/or (4) post-translational modification of a polypeptide or protein.
  • Five prime untranslated region refers to a sequence of an RNA molecule between a transcription start site and a start codon of a coding region of an RNA.
  • “5’ UTR” refers to a sequence of an RNA molecule that begins at a transcription start site and ends one nucleotide (nt) before a start codon (usually AUG) of a coding region of an RNA molecule, e.g., in its natural context.
  • fragment as used herein in the context of a nucleic acid sequence (e.g., RNA sequence) or an amino acid sequence may typically be a portion of a reference sequence.
  • a reference sequence is a full-length sequence of e.g., a nucleic acid sequence or an amino acid sequence.
  • a fragment typically, refers to a sequence that is identical to a corresponding stretch within a reference sequence.
  • a fragment comprises a continuous stretch of nucleotides or amino acid residues that corresponds to at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95% of the total length of a reference sequence from which the fragment is derived.
  • fragment with reference to an amino acid sequence (peptide or polypeptide), relates to a part of an amino acid sequence, e.g., a sequence which represents the amino acid sequence shortened at the N-terminus and/or C-terminus.
  • a fragment of an amino acid sequence comprises at least 6, in particular at least 8, at least 12, at least 15, at least 20, at least 30, at least 50, or at least 100 consecutive amino acids from an amino acid sequence.
  • homology refers to the overall relatedness between polynucleotide molecules (e.g., DNA molecules and/or RNA molecules) and/or between polypeptide molecules.
  • polynucleotide molecules e.g., DNA molecules and/or RNA molecules
  • polypeptide molecules are considered to be “homologous” to one another if their sequences are at least 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 99% identical.
  • polynucleotide molecules e.g., DNA molecules and/or RNA molecules
  • polypeptide molecules are considered to be “homologous” to one another if their sequences are at least 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 99% similar (e.g., containing residues with related chemical properties at corresponding positions).
  • certain amino acids are typically classified as similar to one another as “hydrophobic” or “hydrophilic” amino acids, and/or as having “polar” or “non-polar” side chains.
  • Humoral immunity As used herein, the term “humoral immunity” or “humoral immune response” refers to antibody production and the accessory processes that accompany it, including: Th2 activation and cytokine production, germinal center formation and isotype switching, affinity maturation and memory cell generation. It also refers to the effector functions of antibodies, which include pathogen neutralization, classical complement activation, and opsonin promotion of phagocytosis and pathogen elimination.
  • Identity refers to the overall relatedness between polynucleotide molecules (e.g., DNA molecules and/or RNA molecules) and/or between polypeptide molecules.
  • polynucleotide molecules e.g., DNA molecules and/or RNA molecules
  • polypeptide molecules are considered to be “substantially identical” to one another if their sequences are at least 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical.
  • Calculation of the percent identity of two nucleic acid or polypeptide sequences can be performed by aligning the two sequences for optimal comparison purposes (e.g., gaps can be introduced in one or both of a first and a second sequence for optimal alignment and non-identical sequences can be disregarded for comparison purposes).
  • the length of a sequence aligned for comparison purposes is at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or substantially 100% of the length of a reference sequence.
  • the nucleotides at corresponding positions are then compared.
  • the percent identity between the two sequences is a function of the number of identical positions shared by the sequences, taking into account the number of gaps, and the length of each gap, which needs to be introduced for optimal alignment of the two sequences.
  • the comparison of sequences and determination of percent identity between two sequences can be accomplished using a mathematical algorithm. For example, the percent identity between two nucleotide sequences can be determined using the algorithm of Meyers and Miller, 1989, which has been incorporated into the ALIGN program (version 2.0).
  • nucleic acid sequence comparisons made with the ALIGN program use a PAM120 weight residue table, a gap length penalty of 12 and a gap penalty of 4.
  • the percent identity between two nucleotide sequences can, alternatively, be determined using the GAP program in the GCG software package using an NWSgapdna.CMP matrix.
  • an assessed value achieved with a provided pharmaceutical composition may be “increased” relative to that obtained with a comparable reference pharmaceutical composition (e.g., immunogenic composition, e.g., vaccine).
  • an assessed value achieved in a subject may be “increased” relative to that obtained in the same subject under different conditions (e.g., prior to or after an event; or presence or absence of an event such as administration of a pharmaceutical composition (e.g., immunogenic composition, e.g., vaccine) as described herein, or in a different, comparable subject (e.g., in a comparable subject that differs from the subject of interest in prior exposure to a condition, e.g., absence of administration of a pharmaceutical composition (e.g., immunogenic composition, e.g., vaccine) as described herein.).
  • a pharmaceutical composition e.g., immunogenic composition, e.g., vaccine
  • comparative terms refer to statistically relevant differences (e.g., that are of a prevalence and/or magnitude sufficient to achieve statistical relevance). Those skilled in the art will be aware, or will readily be able to determine, in a given context, a degree and/or prevalence of difference that is required or sufficient to achieve such statistical significance.
  • the term “reduced” or equivalent terms refers to a reduction in the level of an assessed value by at least 5%, at least 10%, at least 20%, at least 50%, at least 75% or higher, as compared to a comparable reference.
  • the term “reduced” or equivalent terms refers to a complete or essentially complete inhibition, i.e., a reduction to zero or essentially to zero.
  • the term “increased” or “induced” refers to an increase in the level of an assessed value by at least 10%, at least 20%, at least 30%, at least 40%, at least 50%, at least 80%, at least 100%, at least 200%, at least 500%, or higher, as compared to a comparable reference.
  • Ionizable refers to a compound or group or atom that is charged at a certain pH. In the context of an ionizable amino lipid, such a lipid or a function group or atom thereof bears a positive charge at a certain pH. In some embodiments, an ionizable amino lipid is positively charged at an acidic pH.
  • an ionizable amino lipid is predominately neutral at physiological pH values, e.g., in some embodiments about 7.0-7.4, but becomes positively charged at lower pH values.
  • an ionizable amino lipid may have a pKa within a range of about 5 to about 7.
  • Isolated means altered or removed from the natural state. For example, a nucleic acid or a peptide naturally present in a living animal is not “isolated”, but the same nucleic acid or peptide partially or completely separated from the coexisting materials of its natural state is “isolated”.
  • RNA lipid nanoparticle As used herein, the term “RNA lipid nanoparticle” refers to a nanoparticle comprising at least one lipid and RNA molecule(s).
  • an RNA lipid nanoparticle comprises at least one ionizable amino lipid. In some embodiments, an RNA lipid nanoparticle comprises at least one ionizable amino lipid, at least one helper lipid, and at least one polymer-conjugated lipid (e.g., PEG-conjugated lipid). In various embodiments, RNA lipid nanoparticles as described herein can have an average size (e.g., Z-average) of about 100 nm to 1000 nm, or about 200 nm to 900 nm, or about 200 nm to 800 nm, or about 250 nm to about 700 nm.
  • an average size e.g., Z-average
  • RNA lipid nanoparticles can have a particle size (e.g., Z- average) of about 30 nm to about 200 nm, or about 30 nm to about 150 nm, about 40 nm to about 150 nm, about 50 nm to about 150 nm, about 60 nm to about 130 nm, about 70 nm to about 110 nm, about 70 nm to about 100 nm, about 80 nm to about 100 nm, about 90 nm to about 100 nm, about 70 to about 90 nm, about 80 nm to about 90 nm, or about 70 nm to about 80 nm.
  • a particle size e.g., Z- average
  • an average size of lipid nanoparticles is determined by measuring the particle diameter.
  • RNA lipid nanoparticles may be prepared by mixing lipids with RNA molecules described herein.
  • Lipidoid As used herein, a “lipidoid” refers to a lipid-like molecule. In some embodiments, a lipoid is an amphiphilic molecule with one or more lipid-like physical properties. In the context of the present disclosure, the term lipid is considered to encompass lipidoids.
  • Nanoparticle As used herein, the term “nanoparticle” refers to a particle having an average size suitable for parenteral administration.
  • a nanoparticle has a longest dimension (e.g., a diameter) of less than 1,000 nanometers (nm). In some embodiments, a nanoparticle may be characterized by a longest dimension (e.g., a diameter) of less than 300 nm. In some embodiments, a nanoparticle may be characterized by a longest dimension (e.g., a diameter) of less than 100 nm. In many embodiments, a nanoparticle may be characterized by a longest dimension between about 1 nm and about 100 nm, or between about 1 ⁇ m and about 500 nm, or between about 1 nm and 1,000 nm.
  • a population of nanoparticles is characterized by an average size (e.g., longest dimension) that is below about 1,000 nm, about 500 nm, about 100 nm, about 50 nm, about 40 nm, about 30 nm, about 20 nm, or about 10 nm and often above about 1 nm.
  • a nanoparticle may be substantially spherical so that its longest dimension may be its diameter.
  • a nanoparticle has a diameter of less than 100 nm as defined by the National Institutes of Health. [0137]
  • Naturally occurring The term “naturally occurring” as used herein refers to an entity that can be found in nature.
  • Neutralization refers to an event in which binding agents such as antibodies bind to a biological active site of a virus such as a receptor binding protein, thereby inhibiting the parasitic infection of cells. In some embodiments, the term “neutralization” refers to an event in which binding agents eliminate or significantly reduce ability of infecting cells.
  • Nucleic acid particle can be used to deliver nucleic acid to a target site of interest (e.g., cell, tissue, organ, and the like).
  • a nucleic acid particle may comprise at least one cationic or cationically ionizable lipid or lipid-like material, at least one cationic polymer such as protamine, or a mixture thereof and nucleic acid.
  • a nucleic acid particle is a lipid nanoparticle.
  • a nucleic acid particle is a lipoplex particle.
  • nucleic acid refers to a polymer of at least 10 nucleotides or more.
  • a nucleic acid is or comprises DNA.
  • a nucleic acid is or comprises RNA.
  • a nucleic acid is or comprises peptide nucleic acid (PNA).
  • PNA peptide nucleic acid
  • a nucleic acid is or comprises a single stranded nucleic acid.
  • a nucleic acid is or comprises a double-stranded nucleic acid.
  • a nucleic acid comprises both single and double-stranded portions.
  • a nucleic acid comprises a backbone that comprises one or more phosphodiester linkages. In some embodiments, a nucleic acid comprises a backbone that comprises both phosphodiester and non-phosphodiester linkages. For example, in some embodiments, a nucleic acid may comprise a backbone that comprises one or more phosphorothioate or 5'-N-phosphoramidite linkages and/or one or more peptide bonds, e.g., as in a “peptide nucleic acid”.
  • a nucleic acid comprises one or more, or all, natural residues (e.g., adenine, cytosine, deoxyadenosine, deoxycytidine, deoxyguanosine, deoxythymidine, guanine, thymine, uracil). In some embodiments, a nucleic acid comprises on or more, or all, non-natural residues.
  • natural residues e.g., adenine, cytosine, deoxyadenosine, deoxycytidine, deoxyguanosine, deoxythymidine, guanine, thymine, uracil.
  • a non-natural residue comprises a nucleoside analog (e.g., 2-aminoadenosine, 2- thiothymidine, inosine, pyrrolo-pyrimidine, 3 -methyl adenosine, 5-methylcytidine, C-5 propynyl-cytidine, C-5 propynyl-uridine, 2-aminoadenosine, C5-bromouridine, C5-fluorouridine, C5-iodouridine, C5-propynyl-uridine, C5 - propynyl-cytidine, C5-methylcytidine, 2-aminoadenosine, 7-deazaadenosine, 7-deazaguanosine, 8-oxoadenosine, 8- oxoguanosine, 6-O-methylguanine, 2-thiocytidine, methylated bases, intercalated bases, and combinations thereof).
  • a non-natural residue comprises one or more modified sugars (e.g., 2'-fluororibose, ribose, 2'-deoxyribose, arabinose, and hexose) as compared to those in natural residues.
  • a nucleic acid has a nucleotide sequence that encodes a functional gene product such as an RNA or polypeptide.
  • a nucleic acid has a nucleotide sequence that comprises one or more introns.
  • a nucleic acid may be prepared by isolation from a natural source, enzymatic synthesis (e.g., by polymerization based on a complementary template, e.g., in vivo or in vitro, reproduction in a recombinant cell or system, or chemical synthesis.
  • enzymatic synthesis e.g., by polymerization based on a complementary template, e.g., in vivo or in vitro, reproduction in a recombinant cell or system, or chemical synthesis.
  • a nucleic acid is at least 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 225, 250, 275, 300, 325, 350, 375, 400, 425, 450, 475, 500, 600, 700, 800, 900, 1000, 1500, 2000, 2500, 3000, 3500, 4000, 4500, 5000, 5500, 6000, 6500, 7000, 7500, 8000, 8500, 9000, 9500, 10,000, 10,500, 11,000, 11,500, 12,000, 12,500, 13,000, 13,500, 14,000, 14,500, 15,000, 15,500, 16,000, 16,500, 17,000, 17,500, 18,000, 18,500, 19,000, 19,500, or 20,000 or more residues or nucleotides long.
  • Nucleotide refers to its art-recognized meaning. When a number of nucleotides is used as an indication of size, e.g., of a polynucleotide, a certain number of nucleotides refers to the number of nucleotides on a single strand, e.g., of a polynucleotide.
  • Patient refers to any organism who is suffering or at risk of a disease or disorder or condition. Typical patients include animals (e.g., mammals such as mice, rats, rabbits, non- human primates, and/or humans).
  • a patient is a human. In some embodiments, a patient is suffering from or susceptible to one or more diseases or disorders or conditions. In some embodiments, a patient displays one or more symptoms of a disease or disorder or condition. In some embodiments, a patient has been diagnosed with one or more diseases or disorders or conditions. In some embodiments, a disease or disorder or condition that is amenable to provided technologies is or includes an HSV infection. In some embodiments, a patient is receiving or has received certain therapy to diagnose and/or to treat a disease, disorder, or condition. In some embodiments, a patient is a patient suffering from or susceptible to an HSV infection.
  • PEG-conjugated lipid refers to a molecule comprising a lipid portion and a polyethylene glycol portion.
  • Pharmaceutical composition refers to an active agent, formulated together with one or more pharmaceutically acceptable carriers.
  • active agent is present in unit dose amount appropriate for administration in a therapeutic regimen that shows a statistically significant probability of achieving a predetermined therapeutic effect when administered to a relevant population.
  • pharmaceutical compositions may be specially formulated for parenteral administration, for example, by subcutaneous, intramuscular, or intravenous injection as, for example, a sterile solution or suspension formulation.
  • compositions comprising: pharmaceutically effective amount or “therapeutically effective amount” refers to the amount which achieves a desired reaction or a desired effect alone or together with further doses.
  • a desired reaction in some embodiments relates to inhibition of the course of the disease. In some embodiments, such inhibition may comprise slowing down the progress of a disease and/or interrupting or reversing the progress of the disease.
  • a desired reaction in a treatment of a disease may be or comprise delay or prevention of the onset of a disease or a condition.
  • compositions e.g., immunogenic compositions, e.g., vaccines
  • an effective amount of pharmaceutical compositions will depend, for example, on a disease or condition to be treated, the severity of such a disease or condition, individual parameters of the patient, including, e.g., age, physiological condition, size and weight, the duration of treatment, the type of an accompanying therapy (if present), the specific route of administration and similar factors. Accordingly, doses of pharmaceutical compositions (e.g., immunogenic compositions, e.g., vaccines) described herein may depend on various of such parameters. In the case that a reaction in a patient is insufficient with an initial dose, higher doses (or effectively higher doses achieved by a different, more localized route of administration) may be used.
  • Poly(A) sequence As used herein, the term “poly(A) sequence” or “poly-A tail” refers to an uninterrupted or interrupted sequence of adenylate residues which is typically located at the 3'-end of an RNA molecule. Poly(A) sequences are known to those of skill in the art and may follow the 3’-UTR in the RNAs described herein. An uninterrupted poly(A) sequence is characterized by consecutive adenylate residues. In nature, an uninterrupted poly(A) sequence is typical.
  • RNAs disclosed herein can have a poly(A) sequence attached to the free 3'-end of the RNA by a template-independent RNA polymerase after transcription or a poly(A) sequence encoded by DNA and transcribed by a template-dependent RNA polymerase.
  • Polypeptide refers to a polymeric chain of amino acids.
  • a polypeptide has an amino acid sequence that occurs in nature.
  • a polypeptide has an amino acid sequence that does not occur in nature.
  • a polypeptide has an amino acid sequence that is engineered in that it is designed and/or produced through action of the hand of man.
  • a polypeptide may comprise or consist of natural amino acids, non-natural amino acids, or both. In some embodiments, a polypeptide may comprise or consist of only natural amino acids or only non-natural amino acids. In some embodiments, a polypeptide may comprise D-amino acids, L-amino acids, or both. In some embodiments, a polypeptide may comprise only D-amino acids. In some embodiments, a polypeptide may comprise only L-amino acids.
  • a polypeptide may include one or more pendant groups or other modifications, e.g., modifying or attached to one or more amino acid side chains, at the polypeptide’s N-terminus, at the polypeptide’s C-terminus, or any combination thereof.
  • such pendant groups or modifications comprise acetylation, amidation, lipidation, methylation, pegylation, etc., including combinations thereof.
  • a polypeptide may be cyclic, and/or may comprise a cyclic portion.
  • a polypeptide is not cyclic and/or does not comprise any cyclic portion.
  • a polypeptide is linear.
  • a polypeptide may be or comprise a stapled polypeptide.
  • the term “polypeptide” may be appended to a name of a reference polypeptide, activity, or structure; in such instances it is used herein to refer to polypeptides that share the relevant activity or structure and thus can be considered to be members of the same class or family of polypeptides.
  • the present specification provides and/or those skilled in the art will be aware of exemplary polypeptides within the class whose amino acid sequences and/or functions are known; in some embodiments, such exemplary polypeptides are reference polypeptides for the polypeptide class or family.
  • a member of a polypeptide class or family shows significant sequence homology or identity with, shares a common sequence motif (e.g., a characteristic sequence element) with, and/or shares a common activity (in some embodiments at a comparable level or within a designated range) with a reference polypeptide of the class; in some embodiments with all polypeptides within the class).
  • a common sequence motif e.g., a characteristic sequence element
  • shares a common activity in some embodiments at a comparable level or within a designated range
  • a member polypeptide shows an overall degree of sequence homology or identity with a reference polypeptide that is at least about 30-40%, and is often greater than about 50%, 60%, 70%, 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more and/or includes at least one region (e.g., a conserved region that may in some embodiments be or comprise a characteristic sequence element) that shows very high sequence identity, often greater than 90% or even 95%, 96%, 97%, 98%, or 99%.
  • a conserved region that may in some embodiments be or comprise a characteristic sequence element
  • a conserved region usually encompasses at least 3-4 and often up to 20 or more amino acids; in some embodiments, a conserved region encompasses at least one stretch of at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15 or more contiguous amino acids.
  • a relevant polypeptide may comprise or consist of a fragment of a parent polypeptide.
  • Recombinant in the context of the present disclosure means “made through genetic engineering”. In some embodiments, a “recombinant” entity such as a recombinant nucleic acid in the context of the present disclosure is not naturally occurring.
  • Reference As used herein, the term “reference” describes a standard or control relative to which a comparison is performed. For example, in some embodiments, an agent, animal, individual, population, sample, sequence or value of interest is compared with a reference or control agent, animal, individual, population, sample, sequence or value.
  • RNA Ribonucleic acid
  • RNA Ribonucleic acid
  • polyribonucleotide refers to a polymer of ribonucleotides.
  • an RNA is single stranded.
  • an RNA is double stranded. In some embodiments, an RNA comprises both single and double stranded portions. In some embodiments, an RNA can comprise a backbone structure as described in the definition of “Nucleic acid / Polynucleotide” above.
  • An RNA can be a regulatory RNA (e.g., siRNA, microRNA, etc.), or a messenger RNA (mRNA). In some embodiments where an RNA is a mRNA. In some embodiments where an RNA is a mRNA, a RNA typically comprises at its 3’ end a poly(A) region.
  • an RNA typically comprises at its 5’ end an art-recognized cap structure, e.g., for recognizing and attachment of a RNA to a ribosome to initiate translation.
  • an RNA is a synthetic RNA.
  • Synthetic RNAs include RNAs that are synthesized in vitro (e.g., by enzymatic synthesis methods and/or by chemical synthesis methods).
  • Ribonucleotide encompasses unmodified ribonucleotides and modified ribonucleotides.
  • unmodified ribonucleotides include the purine bases adenine (A) and guanine (G), and the pyrimidine bases cytosine (C) and uracil (U).
  • Modified ribonucleotides may include one or more modifications including, but not limited to, for example, (a) end modifications, e.g., 5' end modifications (e.g., phosphorylation, dephosphorylation, conjugation, inverted linkages, etc.), 3' end modifications (e.g., conjugation, inverted linkages, etc.), (b) base modifications, e.g.
  • ribonucleotide also encompasses ribonucleotide triphosphates including modified and non-modified ribonucleotide triphosphates.
  • risk is expressed as a percentage. In some embodiments, risk is from 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, 60, 70, 80, 90 up to 100%. In some embodiments risk is expressed as a risk relative to a risk associated with a reference sample or group of reference samples. In some embodiments, a reference sample or group of reference samples have a known risk of a disease, disorder, condition and/or event. In some embodiments a reference sample or group of reference samples are from individuals comparable to a particular individual. In some embodiments, relative risk is 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, or more.
  • RNA lipoplex particle refers to a complex comprising liposomes, in particular cationic liposomes, and RNA molecules. Without wishing to bound by a particular theory, electrostatic interactions between positively charged liposomes and negatively charged RNA results in complexation and spontaneous formation of RNA lipoplex particles.
  • positively charged liposomes may comprise a cationic lipid, such as in some embodiments DOTMA, and additional lipids, such as in some embodiments DOPE.
  • an RNA lipoplex particle is a nanoparticle.
  • specific interaction is dependent upon the presence of a particular structural feature of the target entity (e.g., an epitope, a cleft, a binding site). It is to be understood that specificity need not be absolute. In some embodiments, specificity may be evaluated relative to that of a target-binding moiety for one or more other potential target entities (e.g., competitors). In some embodiments, specificity is evaluated relative to that of a reference specific binding moiety. In some embodiments, specificity is evaluated relative to that of a reference non- specific binding moiety.
  • Subject refers to an organism to be administered with a composition described herein, e.g., for experimental, diagnostic, prophylactic, and/or therapeutic purposes. Typical subjects include animals (e.g., mammals such as mice, rats, rabbits, non-human primates, domestic pets, etc.) and humans. In some embodiments, a subject is a human subject. In some embodiments, a subject is suffering from a disease, disorder, or condition (e.g., an HSV infection). In some embodiments, a subject is susceptible to a disease, disorder, or condition (e.g., an HSV infection).
  • a disease, disorder, or condition e.g., an HSV infection
  • a subject displays one or more symptoms or characteristics of a disease, disorder, or condition (e.g., an HSV infection). In some embodiments, a subject displays one or more non-specific symptoms of a disease, disorder, or condition (e.g., an HSV infection). In some embodiments, a subject does not display any symptom or characteristic of a disease, disorder, or condition (e.g., an HSV infection). In some embodiments, a subject is someone with one or more features characteristic of susceptibility to or risk of a disease, disorder, or condition (e.g., an HSV infection). In some embodiments, a subject is a patient. In some embodiments, a subject is an individual to whom diagnosis and/or therapy is and/or has been administered.
  • Suffering from An individual who is “suffering from” a disease, disorder, and/or condition has been diagnosed with and/or displays one or more symptoms of a disease, disorder, and/or condition.
  • Susceptible to An individual who is “susceptible to” a disease, disorder, and/or condition is one who has a higher risk of developing the disease, disorder, and/or condition than does a member of the general public.
  • an individual who is susceptible to a disease, disorder and/or condition may not have been diagnosed with the disease, disorder, and/or condition.
  • an individual who is susceptible to a disease, disorder, and/or condition may exhibit symptoms of the disease, disorder, and/or condition.
  • an individual who is susceptible to a disease, disorder, and/or condition may not exhibit symptoms of the disease, disorder, and/or condition. In some embodiments, an individual who is susceptible to a disease, disorder, and/or condition will develop the disease, disorder, and/or condition. In some embodiments, an individual who is susceptible to a disease, disorder, and/or condition will not develop the disease, disorder, and/or condition. [0159] Synthetic: As used herein, the term “synthetic” refers to an entity that is artificial, or that is made with human intervention, or that results from synthesis rather than naturally occurring.
  • a synthetic nucleic acid or polynucleotide refers to a nucleic acid molecule that is chemically synthesized, e.g., in some embodiments by solid-phase synthesis.
  • the term “synthetic” refers to an entity that is made outside of biological cells.
  • a synthetic nucleic acid or polynucleotide refers to a nucleic acid molecule (e.g., an RNA) that is produced by in vitro transcription using a template.
  • a therapeutic agent or therapy is any substance that can be used to alleviate, ameliorate, relieve, inhibit, prevent, delay onset of, reduce severity of, and/or reduce incidence of one or more symptoms or features of a disease, disorder, and/or condition.
  • a therapeutic agent or therapy is a medical intervention (e.g., surgery, radiation, phototherapy) that can be performed to alleviate, relieve, inhibit, present, delay onset of, reduce severity of, and/or reduce incidence of one or more symptoms or features of a disease, disorder, and/or condition.
  • a medical intervention e.g., surgery, radiation, phototherapy
  • 3' UTR refer to a sequence of an RNA molecule that begins following a stop codon of a coding region of an open reading frame sequence. In some embodiments, the 3' UTR begins immediately after a stop codon of a coding region of an open reading frame sequence, e.g., in its natural context.
  • Threshold level refers to a level that are used as a reference to attain information on and/or classify the results of a measurement, for example, the results of a measurement attained in an assay.
  • a threshold level means a value measured in an assay that defines the dividing line between two subsets of a population (e.g., a batch that satisfy quality control criteria vs. a batch that does not satisfy quality control criteria).
  • a value that is equal to or higher than the threshold level defines one subset of the population, and a value that is lower than the threshold level defines the other subset of the population.
  • a threshold level can be determined based on one or more control samples or across a population of control samples. A threshold level can be determined prior to, concurrently with, or after the measurement of interest is taken. In some embodiments, a threshold level can be a range of values.
  • Treat As used herein, the term “treat,” “treatment,” or “treating” refers to any method used to partially or completely alleviate, ameliorate, relieve, inhibit, prevent, delay onset of, reduce severity of, and/or reduce incidence of one or more symptoms or features of a disease, disorder, and/or condition.
  • Treatment may be administered to a subject who does not exhibit signs of a disease, disorder, and/or condition. In some embodiments, treatment may be administered to a subject who exhibits only early signs of the disease, disorder, and/or condition, for example for the purpose of decreasing the risk of developing pathology associated with the disease, disorder, and/or condition. In some embodiments, treatment may be administered to a subject at a later-stage of disease, disorder, and/or condition.
  • Vaccination refers to the administration of a composition intended to generate an immune response, for example to a disease-associated (e.g., disease-causing) agent.
  • vaccination can be administered before, during, and/or after exposure to a disease-associated agent, and in certain embodiments, before, during, and/or shortly after exposure to the agent.
  • vaccination includes multiple administrations, appropriately spaced in time, of a vaccine composition.
  • vaccination generates an immune response to an infectious agent.
  • Vaccine refers to a composition that induces an immune response upon administration to a subject. In some embodiments, an induced immune response provides protective immunity.
  • Variant As used herein in the context of molecules, e.g., nucleic acids, proteins, or small molecules, the term “variant” refers to a molecule that shows significant structural identity with a reference molecule but differs structurally from the reference molecule, e.g., in the presence or absence or in the level of one or more chemical moieties as compared to the reference entity. In some embodiments, a variant also differs functionally from its reference molecule. In general, whether a particular molecule is properly considered to be a “variant” of a reference molecule is based on its degree of structural identity with the reference molecule. As will be appreciated by those skilled in the art, any biological or chemical reference molecule has certain characteristic structural elements.
  • a variant by definition, is a distinct molecule that shares one or more such characteristic structural elements but differs in at least one aspect from the reference molecule.
  • a variant polypeptide or nucleic acid may differ from a reference polypeptide or nucleic acid as a result of one or more differences in amino acid or nucleotide sequence and/or one or more differences in chemical moieties (e.g., carbohydrates, lipids, phosphate groups) that are covalently components of the polypeptide or nucleic acid (e.g., that are attached to the polypeptide or nucleic acid backbone).
  • moieties e.g., carbohydrates, lipids, phosphate groups
  • a variant polypeptide or nucleic acid shows an overall sequence identity with a reference polypeptide or nucleic acid that is at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, or 99%.
  • a variant polypeptide or nucleic acid does not share at least one characteristic sequence element with a reference polypeptide or nucleic acid.
  • a reference polypeptide or nucleic acid has one or more biological activities.
  • a variant polypeptide or nucleic acid shares one or more of the biological activities of the reference polypeptide or nucleic acid.
  • a variant polypeptide or nucleic acid lacks one or more of the biological activities of the reference polypeptide or nucleic acid. In some embodiments, a variant polypeptide or nucleic acid shows a reduced level of one or more biological activities as compared to the reference polypeptide or nucleic acid. In some embodiments, a polypeptide or nucleic acid of interest is considered to be a “variant” of a reference polypeptide or nucleic acid if it has an amino acid or nucleotide sequence that is identical to that of the reference but for a small number of sequence alterations at particular positions.
  • a variant polypeptide or nucleic acid comprises about 10, about 9, about 8, about 7, about 6, about 5, about 4, about 3, about 2, or about 1 substituted residues as compared to a reference.
  • a variant polypeptide or nucleic acid comprises a very small number (e.g., fewer than about 5, about 4, about 3, about 2, or about 1) number of substituted, inserted, or deleted, functional residues (i.e., residues that participate in a particular biological activity) relative to the reference.
  • a variant polypeptide or nucleic acid comprises not more than about 5, about 4, about 3, about 2, or about 1 addition or deletion, and, in some embodiments, comprises no additions or deletions, as compared to the reference.
  • a variant polypeptide or nucleic acid comprises fewer than about 25, about 20, about 19, about 18, about 17, about 16, about 15, about 14, about 13, about 10, about 9, about 8, about 7, about 6, and commonly fewer than about 5, about 4, about 3, or about 2 additions or deletions as compared to the reference.
  • a reference polypeptide or nucleic acid is one found in nature.
  • Vector refers to a nucleic acid molecule capable of transporting another nucleic acid to which it has been linked.
  • plasmid refers to a circular double stranded DNA loop into which additional DNA segments may be ligated.
  • vectors are capable of autonomous replication in a host cell into which they are introduced (e.g., bacterial vectors having a bacterial origin of replication and episomal mammalian vectors).
  • Other vectors e.g., non-episomal mammalian vectors
  • certain vectors are capable of directing the expression of genes to which they are operatively linked.
  • vectors are referred to herein as “expression vectors.”
  • known techniques may be used, for example, for generation or manipulation of recombinant DNA, for oligonucleotide synthesis, and for tissue culture and transformation (e.g., electroporation, lipofection).
  • Enzymatic reactions and purification techniques may be performed according to manufacturer's specifications or as commonly accomplished in the art or as described herein.
  • the foregoing techniques and procedures may be generally performed according to conventional methods well known in the art and as described in various general and more specific references that are cited and discussed throughout the present specification. See e.g., Sambrook et al., Molecular Cloning: A Laboratory Manual (4th ed., Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y.
  • compositions e.g., immunogenic compositions, e.g., vaccines
  • HSV herpes simplex virus
  • HSV-2 antigen constructs e.g., HSV-2 antigen constructs, or a combination thereof
  • HSV herpes simplex virus
  • the present disclosure provides HSV (e.g., HSV-1, HSV-2, or both) compositions (e.g., immunogenic compositions, e.g., vaccines) and related technologies (e.g., methods).
  • the present disclosure provides for example, polyribonucleotides that encode one or more GP polypeptides that each comprise an HSV glycoprotein or an antigenic portion thereof.
  • the present disclosure also provides, for example, polyribonucleotides that encode one or more T-cell string polypeptides that each comprises one or more HSV T-cell antigens or antigenic portions threof.
  • a polyribonucleotide provided herein can be part of an RNA construct.
  • a polyribonucleotide or RNA construct as described herein can be part of a composition (e.g., a pharmaceutical composition, e.g., an immunogenic composition, e.g., a vaccine).
  • HSV Herpes Simplex Virus
  • HSV-1 and HSV-2 The structure of HSV-1 and HSV-2 mainly include (from inside to outside) a DNA core, capsid, tegument and envelope. Each of HSV-1 and HSV-2 have a double stranded DNA genome of about 153kb, encoding at least 80 genes.
  • the DNA core is enclosed by an icosapentahedral capsid composed of 162 capsomeres, 150 hexons and 12 pentons, made of six different viral proteins.
  • the DNA is surrounded by at least 20 different viral tegument proteins that have structural and regulatory roles. Some of them participating in capsid transport to the nucleus and other organelles, viral DNA entry into the nucleus, activation of early genes transcription, suppression of cellular protein biosynthesis, and RNA degradation.
  • the viral envelope surrounding the tegument has at least 12 different glycoproteins (B-N) on their surface.
  • the glycoproteins may exist as heterodimers (H/L and E/I) with most existing as monomers.
  • HSV-1 and HSV-2 are responsible for a number of minor, moderate and severe pathologies, including oral and genital ulceration, virally induced blindness, viral encephalitis and disseminated infection of neonates. HSV-1 and HSV-2 are usually transmitted by different routes and affect different areas of the body, but the signs and symptoms that they cause can overlap. Infections caused by HSV-1 represent one of the more widespread infections of the orofacial region and commonly causes herpes labialis, herpetic stomatitis, and keratitis. HSV-2 typically causes genital herpes and is transmitted primarily by direct sexual contact with lesions.
  • HSV infections are transmitted through contact with herpetic lesions, mucosal surfaces, genital secretions, or oral secretions. The average incubation period after exposure is typically 4 days, but may range between 2 and 12 days. HSV particles can infect neuronal prolongations enervating peripheral tissues and establish latency in these cells, namely in the trigeminal ganglia and dorsal root ganglia of the sacral area from where they can sporadically reactivate.
  • HSV infections are lifelong and generally asymptomatic. Without wishing to be bound by any particular theory, it is understood that HSV particles can be shed from infected individuals independent of the occurrence of clinical manifestations. [0175] HSV infections are rarely fatal, but are characterized by blisters that can rupture and become painful. There are few clear differences in clinical presentation based on the type of infecting virus. However, as discussed above, HSV-1 infections tend to be less severe than HSV-2 infections, and patients infected with HSV-2 generally have more outbreaks. A.
  • an HSV (HSV-1 or HSV-2) particle binds to the cell surface using the viral glycoproteins and fuses its envelope with the plasma membrane (see, e.g., FIG.2, Step 1).
  • the viral capsid and tegument proteins are internalized in the cytoplasm (see, e.g., FIG.2, Step 2).
  • the viral capsid Once in the cytoplasm, the viral capsid accumulates in the nucleus and releases viral DNA into the nucleus (see, e.g., FIG.2, Step 3).
  • HSV replicates by three rounds of transcription that yield: ⁇ (immediate early) proteins that mainly regulate viral replication; ⁇ (early) proteins that synthesize and package DNA; and ⁇ (late) proteins, most of which are virion proteins (see, Whitley et.al., Lancet 2001 May 12;357(9267); Taylor et.al., Front Biosci. 2002 Mar 1;7:d752-64; and Ibá ⁇ ez et.al., Front Microbiol. 2018 Oct 11;9:2406; each of which is incorporated herein by reference in its entirety) (see, e.g., FIG.2, Steps 4-6).
  • HSV capsids are assembled within the nucleus of infected cells (see, e.g., FIG..2, Step 7). Once the assembly of viral capsids has been completed in the nucleus, these particles will continue their maturation process in this same compartment through the acquisition of tegument proteins. After leaving the nucleus, additional tegument proteins will be added to the capsids. Meanwhile, the glycoproteins are translated and glycosylated in the endoplasmic reticulum and processed in the trans-Golgi network (TGN) and then directed to multivesicular bodies (see, e.g., FIG.2, Step 8).
  • TGN trans-Golgi network
  • HSV HSV-1 or HSV-2
  • HSV are able to establish a latent infection. After primary infection, HSV either replicates productively in epithelial cells or enters sensory neuron axons and moves to the neuronal cell nucleus.
  • the viral DNA remains as circular, extra-chromosomal DNA, and does not possess any lytic gene expression; however, latency associated transcripts are expressed and then spliced to produce RNA.
  • This general transcriptional silence may allow the virus to remain hidden in the cell by avoiding immune surveillance.
  • technologies e.g., compositions and methods for augmenting, inducing, promoting, enhancing and/or improving an immune response against HSV (e.g., HSV-1 and/or HSV-2) or a component thereof (e.g., a protein or portion thereof).
  • technologies provided herein are designed to augment, induce, promote, enhance and/or improve immunological memory against HSV or a component thereof (e.g., a protein or portion thereof).
  • technologies described herein are designed to act as an immunological boost to a primary composition (e.g., immunogenic composition, e.g., vaccine), such as a composition (e.g., immunogenic composition, e.g., vaccine) directed to an epitope and/or epitopes of HSV (e.g., HSV-1 and/or HSV-2).
  • HSV Genome The genome of HSV-1 and the genome of HSV-2 are both approximately 150 kb long of double- stranded DNA, varying slightly between subtypes and strains. The genome encodes more than 80 genes and has high GC contents: 67 and 69% for HSV-1 and HSV-2, respectively (see, Whitley et.al., Lancet 2001 May 12;357(9267); Taylor et.al., Front Biosci. 2002 Mar 1;7:d752-64; and Jiao et.al., Microbiol Resour Announc.
  • the genome is organized as unique long region (UL) and a unique short region (US).
  • the UL is typically bounded by terminal long (TRL) and internal long (IRL) repeats.
  • the US is typically bounded by terminal short (IRS) and internal short (TRS) repeats.
  • TRL terminal long
  • INL internal long
  • TRS terminal short
  • TRS internal short
  • HSV contains three origins of replication within the genome that are named depending upon their location in either the Long (oriL) or Short (oriS) region of the genome. OriL is found as a single copy in the UL segment, but oriS is located in the repeat region of the Short segment; thus, it is present in the genome in two copies. Both oriL and oriS are palindromic sequences consisting of an AT-rich center region flanked by inverted repeats that contain multiple binding sites of varying affinity for the viral origin binding protein (UL9).
  • UL9 viral origin binding protein
  • oriL or one of the oriS sequences is sufficient for viral replication (see, Whitley et.al., Lancet 2001 May 12;357(9267); Taylor et.al., Front Biosci. 2002 Mar 1;7:d752-64; and Jiao et.al., Microbiol Resour Announc. 2019 Sep; 8(39): e00993-19, which is incorporated herein by reference in its entirety).
  • the viral genome also contains signals that orchestrate proper processing of the newly synthesized genomes for packaging into pre-formed capsids. Progeny genomes are generated in long concatemers that require cleavage into unit-length monomers.
  • the viral genome contains two DNA sequence elements, pac1 and pac2, that ensure proper cleavage and packaging of unit-length progeny genomes. These elements are located within the direct repeats (DR) found within the inverted repeat regions at the ends of the viral genome (see, Whitley et.al., Lancet 2001 May 12;357(9267); Taylor et.al., Front Biosci. 2002 Mar 1;7:d752-64; and Jiao et.al., Microbiol Resour Announc. 2019 Sep; 8(39): e00993-19, which is incorporated herein by reference in its entirety).
  • DR direct repeats
  • ICP0 Infected cell protein 0 (ICP0) of herpes simplex virus 1 (HSV-1) is an ⁇ (immediate-early) protein of herpes simplex virus 1, and is capable of activating HSV-1 gene expression, disrupt nuclear domain (ND) 10 structures, mediate the degradation of cellular proteins, and evade the host cell’s intrinsic and innate antiviral defenses (see., Smith et.al., Future Virol. 2011 Apr; 6(4): 421–429).
  • ICP22 ICP22
  • ICP22 is expressed from an immediate–early (IE) gene during the replication cycle of HSV-1 and HSV-2.
  • ICP22 can generally regulate viral and host gene transcription by changing the phosphorylation status of host RNA polymerase II (RNA pol II) and can also facilitate the nuclear egress complex (NEC) accurately locate to the nuclear membrane to promote nuclear budding (see, Wu et.al., Front Microbiol. 2021 Jun 7;12:668461).
  • RNA pol II RNA polymerase II
  • NEC nuclear egress complex
  • the UL48 gene encodes VP16 or alpha-gene-transactivating factor ( ⁇ -TIF).
  • VP16 is an important transactivator that can activate the transcription of viral immediate-early genes, and in the late stage of viral replication. Additionally, VP16, as a tegument, is involved in viral assembly (see Fan, et.al., Front Microbiol.
  • VP16 released by invading virions binds to the immediate-early (IE) gene promoter to stimulate the transcription of IE genes as a transactivating factor that acts specifically on IE genes (see Fan, et.al., Front Microbiol. 2020; 11: 1910).
  • IE immediate-early
  • VP16 assembles into the tegument to participate in the assembly of virions and promote their maturation (see Fan, et.al., Front Microbiol. 2020; 11: 1910).
  • Glycoproteins [0188] In order to replicate, enveloped HSV must be able to fuse with the membrane of a living cell and deliver their genetic material into its cytoplasm.
  • the HSV viral envelope surrounding the tegument has at least 12 different glycoproteins (gB-gN) on their surface.
  • the glycoproteins may exist as heterodimers (gH/gL and gE/gI) with most existing as monomers.
  • HSV gC, gB, gD, gH, and gL are involved in the process of viral cell entry. Initial attachment is mediated by gC, followed by gD. Then gH/gL pull the virus and the cell membrane together, and then gB triggers the membrane fusion. (Reske et. al., Rev Med Virol. May-Jun 2007; and Arii et. al., Adv Exp Med Biol. 2018;1045:3-21).
  • HSV glycoprotein e.g., gB, gC, gD, gE, gG, gH, gI, and/or gL
  • HSV glycoprotein e.g., gB, gC, gD, gE, gG, gH, gI, and/or gL
  • antigens and antigenic fragments thereof can be useful in preventing or treating HSV, e.g., in HSV antigen constructs and/or HSV compositions (e.g., immunogenic compositions, e.g., vaccines) as further disclosed herein.
  • HSV glycoprotein e.g., gB, gC, gD, gE, gG, gH, gI, and/or gL
  • HSV glycoprotein e.gB, gC, gD, gE, gG, gH, gI, and/or gL
  • Glycoprotein C is a type I membrane glycoprotein and is considered a significant attachment protein and principle viral ligand for binding heparin sulfate proteoglycans (HSPGs) on a cell surface. This binding can occur by gC interaction with HSPG rich regions found on F-actin rich membrane protrusions referred to as filopodia. [0191] Glycoprotein C has also been shown to be involved in regulation of cell entry and infection by increasing pH threshold for acid-induced conformational changes of gB. Low pH induces reversible conformational changes to gB domains I and V, the functional region containing hydrophobic loops important in cell fusion.
  • HSPGs heparin sulfate proteoglycans
  • Glycoprotein C By positively regulating low-pH-induced conformational changes of gB, gC can enhance HSV’s ability to invade cell types, like epithelial cells, that require a low-pH mechanism for invasion. [0192] Glycoprotein C has also been shown to play a role in immune evasion, in addition to its role in attachment. Glycoprotein C is a target for lymphocyte cytotoxicity in certain cell types and is able to bind complement component C3b to inhibit compliment activation. Furthermore, neutralizing epitopes that exist on other HSV glycoproteins, like gB, can be protected by gC, preventing immune responses from blocking fusion. 6.
  • HSV glycoprotein D (gD) is a 46 kDA type I membrane glycoprotein.
  • the N-terminal ectodomain is comprised of 316 amino acids.
  • Glycoprotein D facilitates invasion by interacting with several cell surface receptors, including herpesvirus entry mediator (HVEM), nectin-1 or nectin-2, and heparin sulfate that contain specific modifications. These cellular receptors do not function as co-receptors, as each glycoprotein interaction with a cell’s receptor occurs independently of each other.
  • HVEM herpesvirus entry mediator
  • nectin-1 or nectin-2 heparin sulfate that contain specific modifications.
  • HVEM the first gD receptor identified, belongs to the tumor necrosis factor receptor family and is commonly found on T cells, B cells, dendritic cells, natural killer cells, macrophages, as well as non-immune cell types like neurons and epithelial cells. Within the N-terminus of gD, there is a 37 residue hairpin structure that forms the entire site for binding to HVEM.
  • HVEM cysteine-rich domain 1.
  • this N-terminal extension adopts an extended and flexible conformation.
  • Clinical strains of HSV use nectin-1 for cell entry; however, several mutant strains of HSV utilize nectin-2. Furthermore, heparin sulfate is utilized by HSV-1 but not HSV-2. Glycoprotein D interaction with net-1 has been shown to be essential in some cell types such as neurons, even when other receptors are present on a cell surface. 7.
  • Glycoprotein H (gH)/Glycoprotein L (gL) Complex [0195] Glycoprotein H (gH) is an essential 56kD protein that exists as a heterodimeric complex with 25 kDa glycoprotein L (gL) (complex referred to herein as gH/gL). The gH/gL complex is required for cell fusion and entry. gH/gL does not share any structural similarities with documented fusion proteins and likely does not function as a cofusogen with gB. Instead, gH/gL may act as a regulator of fusion and important component in stabilizing contact between HSV and a cell.
  • Glycoprotein H receives a signal from gD through its H1 domain, and transmits this signal to membrane proximal H3 domain, which in turn propagates that signal to gH’s cytoplasmic tail.
  • gH’s cytoplasmic tail Once gH’s cytoplasmic tail receives this signal, it releases strain on the pre-fusion conformation of gB, which favors attachment of gB’s fusion loop to a cell surface, promoting gB mediated membrane fusion.
  • Mutations in gH’s C-terminal tail have been shown to reduce fusion activity.
  • antibody responses directed towards gH have been shown capable of inhibiting fusion processes mediated by gB-gH-gL.
  • Glycoprotein B is a protein that has an apparent molecular weight of approximately 95-100 kDa and consists of an extended rod or spike-like ectodomain, a hydrophobic membrane proximal region (MPR), a transmembrane region (TMR), and a C-terminal domain (CTD).
  • Glycoprotein B is a class III fusogen. Glycoprotein B ectodomain architecture shares conformational similarity with fusogens from viruses not belonging to the herpesvirdae family. Glycoprotein B is activated through its interaction with gH/gL, but HSV cannot fuse with a target cell through activation of gB alone and requires gB interaction to specific receptors for fusion to be completed.
  • a well-known receptor target of gB is cell-surface heparin sulfate, an interaction that is not essential for HSV fusion, but is known to promote viral adhesion to a cell surface.
  • HSV gB exists in two forms, a pre-fusion and post fusion form. Several changes in the pre-fusion form of gB are thought to lead to its active and post-fusion state. The first change occurs at domain V or at MPR, which allows fusion loops to point towards a cell membrane and away from a viral membrane. This change can produce a compacting intermediate conformation 1 that does not yet attach to a cell membrane surface. The next change occurs at domain III and involves gB adopting an extended intermediate conformation 2 that allows its fusion loop to attach to a cell membrane surface.
  • domain V converts gB to its post-fusion conformation that favors membrane fusion.
  • the post-fusion form of HSV-1 gB has an ectodomain that exists as three protomers that interact to produce a rod-like trimeric structure. Each promoter is comprised of five distinct domains with linker regions that individually form a hairpin shape. Each domain of an individual protomer interacts with the same domain of an adjacent protomer to form the described trimeric structure.
  • Domain I houses an important fusion loop and is commonly referred to as the fusion domain.
  • Domain II facilitates interactions with gH/gL and is referred to as the gH/gL domain.
  • Domain III is comprised of alpha helices that help form the trimeric coil-coil central core of this protein.
  • Domain IV is referred to as the crown domain and sits on top of the post-fusion form; it is believed to bind with cellular receptors. Antibodies that bind to the crown domain can disrupt gB binding to cellular receptors. Domain V consists of a long extension and connects protomers together. 9. Glycoprotein E and glycoprotein I (gE/gI) [0199] Glycoprotein E is approximately 53 kDa and Glycoprotein I is approximately 141 kDa. Both proteins interact to form a heterodimeric complex (complex referred to herein as gE/gI) that plays a role in cell-to-cell spread and virus induced fusion.
  • gE/gI glycoprotein I
  • the gE/gI complex unlike gB, gD, and gH/gL, is not required for fusion and entrance into a cell, but is important for cell-to-cell spread. Disruption of gE/gI formation has effects on HSV proliferation, as this virus relies on cell-to-cell spread for its lytic cycle.
  • the mechanism in which gE/gI facilitate cell-to-cell spread is thought to be reliant on several tegument polypeptides. Cooperation of tegument polypeptides, UL11, UL16, and UL21 may play a role in processing, transport, and biological activity of gE. 10.
  • Glycoprotein G from both HSV-1 (gG1) and HSV-1 (gG2) is the first viral chemokine-binding protein shown to potentiate chemokine function of a cell.
  • Glycoprotein G varies in size significantly between HSV-1 and HSV-2, with a 76 kDa and 43 kDa size, respectively.
  • Glycoprotein G is unique in that its soluble form (SgG2) can have immune modulatory capacity through its extracellular activity.
  • SgG2 binds chemokines through the glycosaminoglycan (GAG)-binding domain of a chemokine without interfering with chemokine’s G protein coupled receptors (GPCRs) binding site.
  • GAG glycosaminoglycan
  • GPCRs G protein coupled receptors
  • SgG2’s interaction with GAG containing proteins allows initiation of lipid raft formation and accumulation, which produces a clustering of chemokine receptors into this micro domain. Clustering of chemokine receptors, in turn, increases local concentration of chemokines on a host cell’s extracellular surface and allows these chemokines to interact with GPCRs. This interaction likely leads to increased immune signaling responses and chemokine stimulation.
  • ICP47 Infected cell protein 47 (ICP47) encoded by gene US12, is a polymorphous protein and could block RNA splicing in early infection, and then, shuttle viral RNA from nucleus to cytoplasm in late infection.
  • ICP47 directly binds antigen-dependent transporter (TAP), limiting antigen trafficking, leading to the occurrence of empty MHC-I (Cheng et.al., Virol J. 2020 Jul 10;17(1):101).
  • TAP antigen-dependent transporter
  • the binding of ICP47 to TAP stabilizes the inward conformation, therefore blocking the translocation pathway points to the endoplasmic reticulum (ER) cavity.
  • ER endoplasmic reticulum
  • VHS virion-host shutoff
  • the virion-host shutoff (VHS) protein is viral protein synthesized with late kinetics and packaged into mature virion particles. Functionally, VHS is a viral RNase that preferentially degrade both host and viral RNA species. VHS has been reported to interfere with dendritic cells (DC) activation during both productive and nonproductive HSV infection (Cotter et.al., J Virol. 2011 Dec; 85(23): 12662–12672.). 13. US3 [0203] All members of the Alphaherpesvirinae subfamily encode a serine/threonine kinase, designated US3.
  • HSV-1 herpes simplex virus type 1
  • nAbs neutralizing antibodies
  • compositions e.g., immunogenic compositions, e.g., vaccines
  • T cell activity in some embodiments, CD4 T cell activity, in some embodiments CD8 T cell activity, in some embodiments, both.
  • compositions e.g., immunogenic compositions, e.g., vaccines
  • CD4 and CD8 epitope(s) of one or more HSV antigens e.g., HSV-1 antigens, HSV-2 antigens, or a combination thereof
  • HSV antigens e.g., HSV-1 antigens, HSV-2 antigens, or a combination thereof
  • B cell antigens and/or epitopes e.g., in addition to one or more B cell antigens and/or epitopes
  • Table 1 Certain HSV Compositions Under Clinical Development Name Platform Antigens Immune Clinical results Response is , is c c c y Failed to reduce shedding c
  • the present disclosure provides the recognition that constructs and/or compositions described herein may be administered as part of regimen with other therapeutic agents.
  • an anti-viral agent can be administered to treat HSV-1 or HSV-2 infection or recurrent episodes.
  • an anti-viral agent is or comprises acyclovir, valacyclovir, famciclovir, or a combination thereof. Table 2 below provides certain information about select anti-viral agents.
  • the present invention provides combinations of nucleotides (e.g., polyribonucleotides) that can be used to express and/or deliver one or more HSV polypeptides or an antigenic portion thereof.
  • nucleotides e.g., polyribonucleotides
  • the present disclosure provides a combination comprising a plurality of polyribonucleotides.
  • the plurality of polyribonucleotides comprise a first set of polyribonucleotides. In some embodiments the plurality of polyribonucleotides comprise a first set of polyribonucleotides and a second set of polyribonucleotides. In some embodiments, a first set of polyribonucleotides encode one or more glycoprotein (GP) polypeptides. In some embodiments, a second set of polyribonucleotides encode one or more T-cell string polypeptides.
  • GP glycoprotein
  • the present disclosure provides the insight that delivering one or more GP polypeptides and one or more T-cell string polypeptides to a subject can induce a more robust immune response to HSV (e.g., HSV-1, HSV-2 or both).
  • HSV e.g., HSV-1, HSV-2 or both
  • GP polypeptides can induce humoral immune responses (e.g., B cell response) to portions of an HSV particle that are surface exposed in a subject prior to infection
  • T-cell string polypeptides can induce cell-mediated immune responses (e.g., T cell response) to HSV molecules (e.g., proteins) produced inside a subject’s cells following infection.
  • a first set of polyribonucleotides can include one or more polyribonucleotides.
  • a polyribonucleotide in a first set can encode a glycoprotein (GP) polypeptide.
  • GP glycoprotein
  • a GP polypeptide comprises an HSV glycoprotein or an antigenic portion thereof.
  • a first set of polyribonucleotides encodes two or more GP polypeptides.
  • a first set of polyribonucleotides comprises two or more polyribonucleotides.
  • two or more GP polypeptides comprise an HSV glycoprotein or an antigenic portion thereof.
  • the two or more GP polypeptides differ.
  • a first set of polyribonucleotides encode three or more GP polypeptides.
  • a first set of polyribonucleotides comprises three or more polyribonucleotides.
  • three or more GP polypeptides comprise an HSV glycoprotein or an antigenic portion thereof. In some embodiments, the three or more GP polypeptides differ.
  • a first set of polyribonucleotides encode four or more GP polypeptides. In some embodiments, a first set of polyribonucleotides comprises four or more polyribonucleotides. In some embodiments, four or more GP polypeptides comprise an HSV glycoprotein or an antigenic portion thereof. In some embodiments, the four or more GP polypeptides differ.
  • a first set of polyribonucleotides encode one or more GP polypeptides, wherein the one or more GP polypeptides comprise an HSV glycoprotein C (gC) or an antigenic portion thereof, an HSV glycoprotein D (gD) or an antigenic portion thereof, an HSV glycoprotein E (gE) or an antigenic portion thereof, an HSV glycoprotein B (gB) or an antigenic portion thereof, an HSV glycoprotein I (gI) or an antigenic portion thereof, an HSV glycoprotein G (gG) or an antigenic portion thereof, an HSV glycoprotein H (gH) or an antigenic portion thereof, an HSV glycoprotein L (gL) or an antigenic portion thereof, or a combination thereof.
  • the one or more GP polypeptides comprise an HSV glycoprotein C (gC) or an antigenic portion thereof, an HSV glycoprotein D (gD) or an antigenic portion thereof, an HSV glycoprotein E (gE) or an antigenic portion thereof, an HSV glycoprotein B (gB) or
  • a first set of polyribonucleotides comprises (i) a polyribonucleotide encoding an HSV gC or an antigenic portion thereof, (ii) a polyribonucleotide encoding an HSV gD or an antigenic portion thereof, (iii) a polyribonucleotide encoding an HSV gE or an antigenic portion thereof, or (iv) a combination thereof.
  • a first set of polyribonucleotides comprises (i) a polyribonucleotide encoding an HSV gB or an antigenic portion thereof, (ii) a polyribonucleotide encoding an HSV gC or an antigenic portion thereof, (iii) a polyribonucleotide encoding an HSV gD or an antigenic portion thereof, or (iv) a combination thereof.
  • a first set of polyribonucleotides comprises (i) a polyribonucleotide encoding an HSV gC or an antigenic portion thereof, (ii) a polyribonucleotide encoding an HSV gD or an antigenic portion thereof, (iii) a polyribonucleotide encoding an HSV gE or an antigenic portion thereof, (iv) a polyribonucleotide encoding an HSV gB or an antigenic portion thereof, or (v) a combination thereof.
  • a first set of polyribonucleotides comprises: (i) a polyribonucleotide that encoding an antigenic portion of HSV gC, (ii) a polyribonucleotide that encoding an antigenic portion of HSV gD, (iii) a polyribonucleotide that encoding an antigenic portion of HSV gE, or (iv) a combination thereof.
  • a first set of polyribonucleotides comprises: (i) a polyribonucleotide that encoding an antigenic portion of HSV gC, (ii) a polyribonucleotide that encoding an antigenic portion of HSV gD, (iii) a polyribonucleotide that encoding an antigenic portion of HSV gE, (iv) a polyribonucleotide that encoding an antigenic portion of HSV gB, or (v) a combination thereof.
  • the first set of polyribonucleotides comprises (i) a polyribonucleotide that encoding an antigenic portion of HSV gC, (ii) a polyribonucleotide that encoding an antigenic portion of HSV gD, and (iii) a polyribonucleotide that encoding an antigenic portion of HSV gE.
  • the first set of polyribonucleotides comprises (i) a polyribonucleotide that encoding an antigenic portion of HSV gC, (ii) a polyribonucleotide that encoding an antigenic portion of HSV gD, (iii) a polyribonucleotide that encoding an antigenic portion of HSV gE, and (iv) a polyribonucleotide that encoding an antigenic portion of HSV gB.
  • the present invention provides one or more glycoprotein (GP) polypeptides each comprising an HSV glycoprotein or an antigenic portion thereof.
  • an HSV glycoprotein or an antigenic portion thereof is an HSV-1 glycoprotein or an antigenic portion thereof. In some embodiments, an HSV glycoprotein or an antigenic portion thereof is an HSV-2 glycoprotein or an antigenic portion thereof. [0225] An overview of exemplary HSV glycoproteins with or without a secretory signal is provided in Table 3 below.
  • Table 3 Exemplary HSV Glycoprotein, Secretory Signal, and Versions HSV-2 Secretory Amino nt Version 1 Version 2 Version 3 Version 4 Glycoprotein signal acid SEQ ID nt nt nt nt SEQ ID NO SEQ ID SEQ ID SEQ ID SEQ ID SEQ ID HSV-2 gE – 327 325 LVVV (SEQ ID NO: Table 4 below, example deoxyribonucleic acid sequences encoding certain HSV gB, gC, gD, and gE glycoproteins are provided in Table 5, and example ribonucleic acid sequences encoding certain HSV gB, gC, gD, and gE glycoproteins are provided in Table 6 below.
  • a first set of polyribonucleotides encode one or more glycoprotein (GP) polypeptides.
  • one or more GP polypeptides each comprise an HSV glycoprotein, a variant thereof, or an antigenic portion thereof.
  • one or more GP polypeptides comprise an HSV glycoprotein, a variant thereof, or an antigenic portion thereof as set forth in Table 4.
  • a GP polypeptide comprises an HSV glycoprotein C (gC) or antigenic portions thereof.
  • a GP polypeptide comprises an antigenic portion of HSV gC.
  • an antigenic portion of HSV gC comprises an amino acid sequence that is at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to the amino acid sequence of SEQ ID NO: 1 or a portion thereof.
  • an antigenic portion of HSV gC has an amino acid sequence that is identical to the amino acid sequence of SEQ ID NO: 1.
  • an antigenic portion of HSV gC comprises an amino acid sequence that is at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to the amino acid sequence of SEQ ID NO: 260 or a portion thereof.
  • an antigenic portion of HSV gC has an amino acid sequence that is identical to the amino acid sequence of SEQ ID NO: 260.
  • At least one polyribonucleotide of the first set of polyribonucleotides comprises a ribonucleic acid sequence that is at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to the ribonucleic acid sequence of any one of SEQ ID NO: 16-19, 147 and 274-281.
  • a GP polypeptide comprises an HSV glycoprotein D (gD) or antigenic portions thereof.
  • a GP polypeptide comprises an antigenic portion of HSV gD.
  • an antigenic portion of HSV gD comprises an amino acid sequence that is at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to the amino acid sequence of SEQ ID NO: 2 or a portion thereof.
  • an antigenic portion of HSV gD has an amino acid sequence that is identical to the amino acid sequence of SEQ ID NO: 2.
  • At least one polyribonucleotide of the first set of polyribonucleotides comprises a ribonucleic acid sequence that is at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to the ribonucleic acid sequence of any one of SEQ ID NO: 20-23, 143, and 286.
  • a GP polypeptide comprises an HSV glycoprotein E (gE) or antigenic portions thereof.
  • a polypeptide comprises an HSV gE antigens or antigenic portions thereof.
  • an antigenic portion of HSV gE comprises an amino acid sequence that is at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to the amino acid sequence of SEQ ID NO: 3 or a portion thereof.
  • an antigenic portion of HSV gE has an amino acid sequence that is identical to the amino acid sequence of SEQ ID NO: 3.
  • At least one polyribonucleotide of the first set of polyribonucleotides comprises a ribonucleic acid sequence that is at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to the ribonucleic acid sequence of any one of SEQ ID NO: 24-27 and 282-285.
  • the present disclosure provides a combination comprising a plurality of polyribonucleotides.
  • a plurality of polyribonucleotides comprise a first set of polyribonucleotides.
  • a first set of polyribonucleotides encode one or more glycoprotein (GP) polypeptides.
  • at least one polyribonucleotide of the polyribonucleotides in the first set encodes a GP polypeptide that comprises an HSV glycoprotein (gB) variant thereof, or one or more antigenic portions thereof.
  • GP glycoprotein
  • At least one polyribonucleotide of the polyribonucleotides in the first set encodes a GP polypeptide that comprises an antigenic portion of HSV glycoprotein (gB) or an antigenic portion of a variant of HSV gB.
  • a GP polypeptide comprises an HSV glycoprotein B (gB), variant thereof, or antigenic portions thereof.
  • a GP polypeptide comprises an antigenic portion of HSV gB.
  • an HSV gB comprises an amino acid sequence that is at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to the amino acid sequence of any one of SEQ ID NO: 379 or a portion thereof.
  • an antigenic portion of HSV gB comprises an amino acid sequence that is at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to the amino acid sequence of any one of SEQ ID NO: 380 or a portion thereof.
  • an HSV gB comprises an amino acid sequence that is at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to the amino acid sequence of any one of SEQ ID NO: 381 or a portion thereof.
  • an antigenic portion of HSV gB comprises an amino acid sequence that is at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to the amino acid sequence of any one of SEQ ID NO: 382 or a portion thereof.
  • an HSV gB comprises an amino acid sequence that is at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to the amino acid sequence of any one of SEQ ID NO: 383 or a portion thereof.
  • HSV gB comprises an amino acid sequence that is at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to the amino acid sequence of any one of SEQ ID NO: 384 or a portion thereof.
  • an HSV gB comprises an amino acid sequence that is identical to the amino acid of SEQ ID NO: 379.
  • an HSV gB comprises an amino acid sequence that is identical to the amino acid of SEQ ID NO: 380.
  • an HSV gB has an amino acid sequence that is identical to the amino acid of SEQ ID NO: 381. In some embodiments, an HSV gB has an amino acid sequence that is identical to the amino acid of SEQ ID NO: 382. In some embodiments, an HSV gB has an amino acid sequence that is identical to the amino acid of SEQ ID NO: 383. In some embodiments, an HSV gB has an amino acid sequence that is identical to the amino acid of SEQ ID NO: 384.
  • HSV compositions e.g., immunogenic compositions, e.g., vaccines
  • HSV gB HSV-1 gB, HSV-2 gB, or both.
  • HSV gB can be unstable.
  • cell entry of enveloped viruses requires specialized viral proteins that mediate fusion with the host membrane.
  • the viral proteins, including gB undergo substantial structural rearrangements from a metastable prefusion conformation to a stable postfusion conformation (FIG.16).
  • the term “stable”, when applied to glycoprotein B, means that glycoprotein B maintains one of more aspects of a physical structure (e.g., maintains a specific conformation) and/or activity for a specific period of time.
  • a stable glycoprotein B has been modified (e.g., certain mutations) so that its structure is stabilized.
  • a stable glycoprotein B structure is maintained for a specific period of time.
  • a stable glycoprotein B is in a stable prefusion conformation.
  • a stable glycoprotein B is in a stable postfusion conformation. In some embodiments, a stable glycoprotein B maintains a biological relevant activity (e.g., antigenic potential). In some embodiments, a stable glycoprotein B is in a stable prefusion conformation and maintains antigenic potential. [0240] The present disclosure encompasses a recognition that stabilization of gB or antigenic portions thereof can be useful or advantageous for eliciting an immune response.
  • the present disclosure further provides the recognition that stabilization of HSV gB or antigenic portions thereof can be particularly advantageous for use, e.g., in preventing or treating HSV, e.g., in HSV antigen constructs and/or HSV compositions (e.g., immunogenic compositions, e.g., vaccines) as further disclosed herein. Accordingly, the present disclosure provides certain mutations that can stabilize HSV gB or antigenic portions thereof.
  • a polyribonucleotide encoding a GP polypeptide that comprises an HSV glycoprotein B (gB), variant thereof, or one or more antigenic portions thereof.
  • an HSV gB or antigenic portion thereof comprises one or more mutations that stabilize the HSV gB or antigenic portion thereof relative to a comparable HSV gB or antigenic portion thereof that does not comprise the one or more mutations.
  • one or more mutations are one or more amino acid substitutions.
  • one or more amino acid substitutions comprise 120C, 181C, 238C, 251C, 259C, 290C, 291C, 391C, 526C, 571C, 610C, 630C, 636C, 676C, 677C, 680C, 714C, 718C, 725C, 758C, and combinations thereof, wherein the numbering is with reference to SEQ ID NO: 379.
  • an HSV gB comprises an amino acid sequence according to SEQ ID NO: 379 or antigenic portion thereof, except that the HSV gB or antigenic portion thereof comprises one or more mutations comprising 120C, 181C, 238C, 251C, 259C, 290C, 291C, 391C, 526C, 571C, 610C, 630C, 636C, 676C, 677C, 680C, 714C, 718C, 725C, 758C, and combinations thereof.
  • an HSV gB has an amino acid sequence that (i) is at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the amino acid SEQ ID NO: 379 and (ii) comprises one or more mutations, where the one or more mutations comprise 120C, 181C, 238C, 251C, 259C, 290C, 291C, 391C, 526C, 571C, 610C, 630C, 636C, 676C, 677C, 680C, 714C, 718C, 725C, 758C, and combinations thereof.
  • an HSV gB has an amino acid sequence that comprises one or more mutations, where the one or more mutations comprise: (a) 120C and 677C, (b) 181C and 725C, (c) 238C and 610C, (d) 251C and 718C, (e) 259C and 758C, (f) 290C and 680C, (g) 291C and 636C, (h) 391C and 526C, (i) 571C and 676C, (j) 571C and 680C, and/or (k) 630C and 714C, where the numbering is with reference to SEQ ID NO: 379.
  • an HSV gB has an amino acid sequence that (i) is at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the amino acid SEQ ID NO: 379 and (ii) comprises one or more mutations, where the one or more mutations comprise: (a) 120C and 677C, (b) 181C and 725C, (c) 238C and 610C, (d) 251C and 718C, (e) 259C and 758C, (f) 290C and 680C, (g) 291C and 636C, (h) 391C and 526C, (i) 571C and 676C, (j) 571C and 680C, and/or (k) 630C and 714C, where the numbering is with reference to SEQ ID NO: 379.
  • an HSV gB or antigenic portion thereof does not comprise an 516P mutation, where the numbering is with reference to SEQ ID NO: 379.
  • the present disclosure provides a plurality of polyribonucleotides, wherein the plurality of polyribonucleotides comprises a first set of polyribonucleotides that comprises: (i) a polyribonucleotide that encoding an antigenic portion of HSV gC as provided herein, (ii) a polyribonucleotide that encoding an antigenic portion of HSV gD as provided herein, (iii) a polyribonucleotide that encoding an antigenic portion of HSV gE as provided herein, and (iv) a polyribonucleotide that encoding an antigenic portion of HSV gB or variant thereof as provided herein.
  • a first set of polyribonucleotides further comprises a polyribonucleotide encoding an HSV glycoprotein G (gG) or an antigenic portion thereof, a polyribonucleotide encoding an HSV glycoprotein H (gH) or an antigenic portion thereof, a polyribonucleotide encoding an HSV glycoprotein I (gI) or an antigenic portion thereof, a polyribonucleotide encoding an HSV glycoprotein L (gL) or an antigenic portion thereof, or a combination thereof.
  • G HSV glycoprotein G
  • I HSV glycoprotein I
  • gL HSV glycoprotein L
  • combinations provided herein further comprises a second set of polyribonucleotides.
  • a second set of polyribonucleotides encode one or more T-cell string polypeptides.
  • one or more T-cell string polypeptides comprises one or more HSV T-cell antigens or antigenic portions thereof.
  • one or more HSV T-cell antigens comprises (i) one or more HSV RS1 polypeptides or antigenic portions thereof, (ii) one or more HSV RL2 polypeptides or antigenic portions thereof, (iii) one or more HSV UL1 polypeptides or antigenic portions thereof, (iv) one or more HSV UL5 polypeptides or antigenic portions thereof, (v) one or more HSV UL9 polypeptides or antigenic portions thereof, (vi) one or more HSV UL19 polypeptides or antigenic portions thereof, (vii) one or more HSV UL21 polypeptides or antigenic portions thereof, (viii) one or more HSV UL25 polypeptides or antigenic portions thereof, (ix) one or more HSV UL27 polypeptides or antigenic portions thereof, (x) one or more HSV UL29 polypeptides or antigenic portions thereof, (xi) one or more HSV UL30 polypeptides or antigenic
  • Example antigen amino acid sequences are shown in Table 7 below.
  • Table 7 Example amino acid antigen sequences SEQ Antigen Strain Amino Acid Sequence ID NO 476 UL1 HG52 MGFVCLFGLVVMGAWGAWGGSQATEYVLRSVIAKEVGDILRVPCMRTPADDVS F R G G V A G L D G G V A G L D G G V A G L D G A G L D G A P S F A L R Y T T GPULIEGQLGENNELRLTRDALEPCTVGHRRYFIFGGGYVYFEEYAYSHQLSRADVT TVSTFIDLNITMLEDHEFVPLEVYTRHEIKDSGLLDYTEVQRRNQLHDLRFADIDTVI RADANAAMFAGLCAFFEGMGDLGRAVGKVVMGVVGGVVSAVSGVSSFMSNPFGAL G R A P S F A L R Y T T T R A G Y T G V D V A D G E R E I N L E H R A D V A D G E
  • a T-cell string polypeptide includes a plurality of HSV T-cell antigens or antigenic portions thereof (e.g., a plurality of HSV antigens that are or include one or more T cell and/or B cell antigens for HSV).
  • T cell antigens include, e.g., CD4 T cell antigens and/or CD8 T cells.
  • an HSV antigen is a T cell antigen.
  • an HSV antigen is a B cell antigen.
  • an HSV T-cell antigen comprises at least one of UL1, UL21, UL27, UL29, UL39, UL40, UL46, UL47, UL48, UL49, RS1, RL2, UL5, UL9, UL19, UL25, UL30, UL52, US1, US7, US8, UL22, US10, US12, UL26, UL50, and/or UL54 or antigenic portion thereof.
  • a T-cell polypeptide comprises a plurality of (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19 of) UL1, UL21, UL27, UL29, UL39, UL40, UL46, UL47, UL48, UL49, RS1, RL2, UL5, UL9, UL19, UL25, UL30, UL52, US1, US7, US8, UL22, US10, US12, UL26, UL50, and/or UL54 or antigenic portions thereof.
  • a T-cell polypeptide comprises and/or encodes UL54, UL29, UL39, UL9, UL30a (UL30.1), UL40, UL5a (UL5.1), UL21, and/or UL46 or fragments thereof.
  • a T-cell polypeptide comprises and/or encodes UL54, UL29, UL40, and/or UL47 or fragments thereof.
  • a T-cell polypeptide comprises and/or encodes at least UL47 or fragments thereof.
  • a T-cell polypeptide comprises and/or encodes at least UL40 or fragments thereof.
  • a T-cell polypeptide comprises and/or encodes one or more of RL2, UL54, UL9, UL39, UL29, UL5, UL40, UL30, UL49, UL46 and/or UL21 or fragments thereof.
  • the UL1 open reading frame encodes HSV gL (also referred to herein as UL1 polypeptide).
  • an HSV antigen e.g., a T-cell or B cell antigen for HSV
  • a UL1 polypeptide or fragment thereof has at least 80% sequence identity with a UL1 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity).
  • UL1 polypeptides known in the art include UL1 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains.
  • a UL1 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 476.
  • an HSV antigen e.g., a T cell or B cell antigen for HSV
  • a UL21 polypeptide or fragment thereof has at least 80% sequence identity with a UL21 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity).
  • UL21 polypeptides known in the art include UL21 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains.
  • a UL21 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 477, 478 and/or 479.
  • the UL27 open reading frame encodes HSV gB (also referred to herein as UL27 polypeptide).
  • an HSV antigen e.g., a T cell or B cell antigen for HSV
  • a UL27 polypeptide or fragment thereof has at least 80% sequence identity with a UL27 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity).
  • UL27 polypeptides known in the art include UL27 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains.
  • a UL27 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 480, 481, 482 and/or 483.
  • an HSV antigen e.g., a T cell or B cell antigen for HSV
  • a UL29 polypeptide or fragment thereof has at least 80% sequence identity with a UL29 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity).
  • UL29 polypeptides known in the art include UL29 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains.
  • a UL29 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 484, 485, and/or 486.
  • an HSV antigen e.g., a T cell or B cell antigen for HSV
  • a UL39 polypeptide or fragment thereof has at least 80% sequence identity with a UL39 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity).
  • UL39 polypeptides known in the art include UL39 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains.
  • a UL39 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 487, 488 and/or 489.
  • an HSV antigen e.g., a T cell or B cell antigen for HSV
  • a UL40 polypeptide or fragment thereof has at least 80% sequence identity with a UL40 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity).
  • UL40 polypeptides known in the art include UL40 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains.
  • a UL40 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 490, 491 and/or 492.
  • an HSV antigen e.g., a T cell or B cell antigen for HSV
  • a UL46 polypeptide or fragment thereof has at least 80% sequence identity with a UL46 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity).
  • UL46 polypeptides known in the art include UL46 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains.
  • a UL46 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 493, 494 and/or 495.
  • an HSV antigen e.g., a T cell or B cell antigen for HSV
  • a UL47 polypeptide or fragment thereof has at least 80% sequence identity with a UL47 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity).
  • UL47 polypeptides known in the art include UL47 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains.
  • a UL47 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 496, 497 and/or 498.
  • an HSV antigen e.g., a T cell or B cell antigen for HSV
  • a UL48 polypeptide or fragment thereof has at least 80% sequence identity with a UL48 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity).
  • UL48 polypeptides known in the art include UL48 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains.
  • a UL48 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 499, 500 and/or 501.
  • an HSV antigen e.g., a T cell or B cell antigen for HSV
  • a UL49 polypeptide or fragment thereof has at least 80% sequence identity with a UL49 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity).
  • UL49 polypeptides known in the art include UL49 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains.
  • a UL49 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 502, 503 and/or 504.
  • an HSV antigen e.g., a T cell or B cell antigen for HSV
  • RS1 polypeptide or fragment thereof is or includes a RS1 polypeptide or fragment thereof.
  • a RS1 polypeptide or fragment thereof has at least 80% sequence identity with a RS1 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity).
  • RS1 polypeptides known in the art include RS1 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains.
  • a RS1 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 505, 506 and/or 507.
  • an HSV antigen e.g., a T cell or B cell antigen for HSV
  • a RL2 polypeptide or fragment thereof has at least 80% sequence identity with a RL2 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity).
  • RL2 polypeptides known in the art include RL2 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains.
  • a RL2 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 508, 509 and/or 510.
  • an HSV antigen e.g., a T cell or B cell antigen for HSV
  • a UL5 polypeptide or fragment thereof has at least 80% sequence identity with a UL5 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity).
  • UL5 polypeptides known in the art include UL5 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains.
  • a UL5 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 511, 512 and/or 513.
  • an HSV antigen e.g., a T cell or B cell antigen for HSV
  • a UL9 polypeptide or fragment thereof has at least 80% sequence identity with a UL9 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity).
  • UL9 polypeptides known in the art include UL9 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains.
  • a UL9 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 514, 515 and/or 516.
  • an HSV antigen e.g., a T cell or B cell antigen for HSV
  • a UL19 polypeptide or fragment thereof has at least 80% sequence identity with a UL19 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity).
  • UL19 polypeptides known in the art include UL19 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains.
  • a UL19 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 517, 518 and/or 519.
  • an HSV antigen e.g., a T cell or B cell antigen for HSV
  • a UL25 polypeptide or fragment thereof has at least 80% sequence identity with a UL25 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity).
  • UL25 polypeptides known in the art include UL25 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains.
  • a UL25 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 520, 521 and/or 522.
  • an HSV antigen e.g., a T cell or B cell antigen for HSV
  • a UL30 polypeptide or fragment thereof has at least 80% sequence identity with a UL30 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity).
  • UL30 polypeptides known in the art include UL30 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains.
  • a UL30 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 523, 524 and/or 525.
  • an HSV antigen e.g., a T cell or B cell antigen for HSV
  • a UL52 polypeptide or fragment thereof has at least 80% sequence identity with a UL52 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity).
  • UL52 polypeptides known in the art include UL52 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains.
  • a UL52 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 526, 527 and/or 528.
  • an HSV antigen e.g., a T cell or B cell antigen for HSV
  • a US1 polypeptide or fragment thereof has at least 80% sequence identity with a US1 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity).
  • US1 polypeptides known in the art include US1 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains.
  • a US1 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 532, 533, 534, 535 and/or 536.
  • the US7 open reading frame encodes HSV gI (also referred to herein as US7 polypeptide).
  • an HSV antigen e.g., a T cell or B cell antigen for HSV
  • a US7 polypeptide or fragment thereof has at least 80% sequence identity with a US7 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity).
  • US7 polypeptides known in the art include US7 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains.
  • a US7 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 537, 538, 539 and/or 540.
  • the US8 open reading frame encodes HSV gE (also referred to herein as US8 polypeptide).
  • an HSV antigen is (e.g., a T cell or B cell antigen for HSV) or includes a US8 polypeptide or fragment thereof.
  • a US8 polypeptide or fragment thereof has at least 80% sequence identity with a US8 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity).
  • US8 polypeptides known in the art include US8 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains.
  • a US8 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 541, 542, 543 and/or 544.
  • the UL22 open reading frame encodes HSV gH (also referred to herein as UL22 polypeptide).
  • an HSV antigen is (e.g., a T cell or B cell antigen for HSV) or includes a UL22 polypeptide or fragment thereof.
  • a UL22 polypeptide or fragment thereof has at least 80% sequence identity with a UL22 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity).
  • UL22 polypeptides known in the art include UL22 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains.
  • a UL22 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 545, 546, 547 and/or 548.
  • an HSV antigen e.g., a T cell or B cell antigen for HSV
  • a US10 polypeptide or fragment thereof has at least 80% sequence identity with a US10 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity).
  • US10 polypeptides known in the art include US10 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains.
  • a US10 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 551, 552, 553 and/or 554.
  • an HSV antigen e.g., a T cell or B cell antigen for HSV
  • a US12 polypeptide or fragment thereof has at least 80% sequence identity with a US12 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity).
  • US12 polypeptides known in the art include US12 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains.
  • a US12 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NO: 555.
  • an HSV antigen e.g., a T cell or B cell antigen for HSV
  • a UL26 polypeptide or fragment thereof has at least 80% sequence identity with a UL26 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity).
  • UL26 polypeptides known in the art include UL26 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains.
  • a UL26 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 558, 559, 560, and/or 561.
  • an HSV antigen e.g., a T cell or B cell antigen for HSV
  • a UL50 polypeptide or fragment thereof has at least 80% sequence identity with a UL50 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity).
  • UL50 polypeptides known in the art include UL50 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains.
  • a UL50 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 556 and/or 557.
  • an HSV antigen e.g., a T cell or B cell antigen for HSV
  • a UL54 polypeptide or fragment thereof has at least 80% sequence identity with a UL54 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity).
  • UL54 polypeptides known in the art include UL54 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains.
  • a UL54 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 529, 530, and/or 531.
  • at least one of the polypeptides in the second set of polynucleotides encode one or more T-cell string polypeptides.
  • a T-cell string polypeptide can include one or more HSV antigens including one or more T cell antigens (e.g., CD4 and/or CD8 T cell antigens) for HSV of the present disclosure and one or more HSV antigens that is not a T cell antigen of the present disclosure.
  • a polyribonucleotide encoding a T-cell string polypeptide can include and/or encode one or more HSV antigens including one or more B cell antigens for HSV of the present disclosure and one or more HSV antigens that is not a B cell antigen of the present disclosure.
  • a polyribonucleotide encoding a T-cell string polypeptide can include and/or encode one or more HSV antigens including one or more T cell antigens for HSV of the present disclosure and one or more HSV antigens that is a B cell antigen for HSV (e.g., an antigen that is or includes a B cell epitope disclosed herein or otherwise known in the art).
  • a polyribonucleotide encoding a T-cell string polypeptide can include and/or encode one or more HSV antigens including one or more T cell antigens for HSV of the present disclosure and one or more HSV antigens selected from HSV glycoproteins or fragments thereof.
  • a polyribonucleotide encoding a T-cell string polypeptide can include and/or encode one or more HSV antigens including one or more T cell antigens for HSV of the present disclosure and one or more HSV antigens selected from an HSV gD protein or an antigenic fragment thereof, an HSV gB protein or an antigenic fragment thereof, an HSV gE protein or an antigenic fragment thereof, an HSV gG protein or an antigenic fragment thereof, an HSV gI protein or an antigenic fragment thereof, an HSV gH protein or an antigenic fragment thereof, an HSV gL protein or an antigenic fragment thereof, an HSV ICP4 protein or an antigenic fragment thereof, or an ICP8 protein or an antigenic fragment thereof.
  • a polyribonucleotide encoding a T-cell string polypeptide can be present in a composition for delivery of the HSV antigen construct to a subject.
  • an HSV antigen construct can be present in a composition for delivery of one or more HSV antigens and/or epitopes to a subject.
  • a polyribonucleotide encoding a T-cell string polypeptide can be or include an RNA molecule that encodes one or more antigens and/or epitopes.
  • compositions for delivery of a polyribonucleotide encoding a T-cell string polypeptide advantageously include, for example, one or more B cell antigens for HSV and one or more T cell antigens (e.g., CD4 and/or CD8 T cell antigens) for HSV.
  • B cell antigens for HSV and one or more T cell antigens (e.g., CD4 and/or CD8 T cell antigens) for HSV.
  • T cell antigens e.g., CD4 and/or CD8 T cell antigens
  • combination of B cell antigens and T cell antigens can be advantageous in promoting immune system defenses against HSV at multiple lifecycle points include without limitation prior to cellular entry and after cellular entry.
  • the present disclosure provides an insight that many prior strategies for developing pharmaceutical compositions (e.g., immunogenic compositions, e.g., vaccines) for treatment of and/or protection from viral infection have focused primarily, or even almost exclusively, on development of neutralizing antibodies that target surface glycoproteins.
  • the present disclosure identifies a problem with such strategies including, for example, that they may fail to appreciate value or even criticality of ensuring that an induced immune response includes significant T cell activity (in some embodiments, CD4 T cell activity, in some embodiments CD8 T cell activity, in some embodiments, both).
  • the present disclosure provides an insight that consideration of expression of HSV proteins (e.g., at particular periods of the HSV life cycle and/or in particular tissues or compartments of an infected subject) can improve composition (e.g., immunogenic composition, e.g., vaccine) effectiveness.
  • composition e.g., immunogenic composition, e.g., vaccine
  • the present disclosure provides technologies for identifying, selecting, and/or characterizing HSV protein sequences (e.g., HSV-1 protein sequences, HSV-2 protein sequences, or a combination thereof), and combinations thereof, particularly useful for inclusion in pharmaceutical compositions (e.g., immunogenic compositions, e.g., vaccines) as described herein.
  • compositions e.g., immunogenic compositions, e.g., vaccines
  • CD4 and CD8 antigen(s) of one or more HSV proteins e.g., HSV-1 proteins, HSV-2 proteins, or a combination thereof
  • HSV proteins e.g., HSV-1 proteins, HSV-2 proteins, or a combination thereof
  • HSV antigen constructs e.g., HSV-1 antigen constructs, HSV-2 antigen constructs, or a combination thereof
  • compositions e.g., pharmaceutical compositions, e.g., immunogenic compositions, e.g., vaccines
  • antigen constructs that induce both neutralizing antibodies and T cells (e.g., CD4 and/or CD8 T cells).
  • neutralizing antibodies and T cells e.g., CD4 and/or CD8 T cells
  • the present disclosure provides such polyribonucleotides and compositions that induce particularly strong neutralizing antibody responses and/or particularly diverse T cell responses (e.g., targeting multiple T cell antigens). [0295] In some embodiments, the present disclosure provides such polyribonucleotides and compositions that induce robust B cell responses. In some embodiments, a B cell response includes the production of a diverse, specific repertoire of antibodies. [0296] In some embodiments, the present disclosure provides such polyribonucleotides and compositions that induce T cell and B cell responses to HSV antigens and/or epitopes.
  • the present disclosure provides the recognition, for example, that polyribonucleotides and compositions comprising RNA molecules as described herein (e.g., encoding for one or more HSV (e.g., HSV-1 and/or HSV-2) antigens and/or epitopes) may result in a higher degree of antigen presentation to various immune system components and/or pathways.
  • administration of such constructs or compositions may induce T cell and/or B cell responses.
  • the present disclosure provides the insight that, e.g., in some embodiments in which T cell and B cell responses are induced in a subject, the subject may have a more sustained, long-term immune response.
  • Such an immune response can be beneficial, e.g., for preventing HSV (e.g., HSV-1 and/or HSV-2) reactivation with a single administration, which may increase vaccination rates and subject compliance as compared with presently available vaccines that require dosing every few years.
  • HSV e.g., HSV-1 and/or HSV-2
  • constructs and compositions comprising RNA molecules as described herein (e.g., encoding for one or more HSV (e.g., HSV-1, HSV- 2, or a combination thereof) antigens and/or epitopes) can provide more diverse protection (e.g., protection against HSV (e.g., HSV-1 and/or HSV-2) variants) because, without wishing to be bound to any particular theory, the constructs and compositions can induce multiple immune system responses.
  • the present disclosure also provides the recognition that, by administering polyribonucleotides and compositions that encode HSV (e.g., HSV-1 and/or HSV-2) antigens and/or epitopes, the polyribonucleotides and compositions described herein avoid administering HSV (e.g., HSV-1 and/or HSV-2) virions, which may infect the subject, go into latency, and reactivate to cause a flare-up.
  • HSV e.g., HSV-1 and/or HSV-2
  • the present disclosure provides an insight (and also identifies a source of a problem in some prior HSV vaccination strategies) that, in some embodiments, particularly effective pharmaceutical compositions (e.g., immunogenic compositions, e.g., vaccines) alter one or more characteristics of the innate immune system.
  • compositions including, for example, compositions that comprise RNA construct(s) encoding HSV (e.g., HSV-1 and/or HSV-2) protein(s) (e.g., HSV antigens or HSV epitopes) as described herein.
  • the present disclosure provides particular pharmaceutical composition (e.g., immunogenic composition, e.g., vaccine) formats including, for example, RNA pharmaceutical compositions (e.g., immunogenic compositions, e.g., vaccines) comprising particular elements and/or sequences useful for vaccination.
  • RNA pharmaceutical compositions e.g., immunogenic compositions, e.g., vaccines
  • the present disclosure provides a variety of insights and technologies related to such HSV (e.g., HSV-1 and/or HSV-2) antigen constructs and compositions (e.g., RNA vaccine).
  • provided compositions e.g.
  • compositions include an RNA active encoding one or more HSV (e.g., HSV-1 and/or HSV-2) polypeptides or antigenic fragments thereof; in some embodiments such RNA active is a modified RNA format in that its uridine residues are substituted with uridine analog(s) such as pseudouridine; alternatively or additionally, in some embodiments, such RNA active includes particular elements (e.g., cap, 5’UTR, 3’UTR, polyA tail, etc) and/or characteristics (e.g., codon optimization) identified, selected, characterized, and/or demonstrated to achieve significant (e.g., elevated) translatability (e.g., in vitro) and/or expression (i.e., in a subject to whom it has been administered) of encoded protein(s).
  • HSV e.g., HSV-1 and/or HSV-2
  • RNA active is a modified RNA format in that its uridine residues are substituted with uridine analog(s) such as pseudo
  • such RNA active includes particular elements and/or characteristics identified, selected, characterized, and/or demonstrated to achieve significant RNA stability and/or efficient manufacturing, particularly at large scale (e.g., 0.1-10 g, 10-500 g, 500 g-1 kg, 750 g-1.5 kg; those skilled in the art will appreciate that different products may be manufactured at different scales, e.g., depending on patient population size).
  • such RNA manufacturing scale may be within a range of about 0.01 g/hr RNA to about 1 g/hr RNA, 1 g/hr RNA to about 100 g/hr RNA, about 1 g RNA/hr to about 20 g RNA/hr, or about 100 g RNA/hr to about 10,000 g RNA/hr. In some embodiments, such RNA manufacturing scale may be tens or hundreds of milligrams to tens or hundreds of grams (or more) of RNA per batch.
  • such RNA manufacturing scale may allow a batch size within a range of about 0.01 g to about 500 g RNA, about 0.01 g to about 10 g RNA, about 1 g to about 10 g RNA, about 10 g to about 500 g RNA, about 10 g to about 300 g RNA, about 10 g to about 200 g RNA or about 30 g to about 60 g RNA.
  • the present disclosure provides technologies for rapid development of a pharmaceutical composition (e.g., immunogenic composition, e.g., HSV vaccine) for delivering particular HSV (e.g., HSV-1 and/or HSV-2) antigen constructs to a subject.
  • a pharmaceutical composition e.g., immunogenic composition, e.g., HSV vaccine
  • particular HSV e.g., HSV-1 and/or HSV-2 antigen constructs to a subject.
  • the present disclosure provides, for example, nucleic acid constructs encoding HSV (e.g., HSV-1 and/or HSV-2) antigens as described herein, expressed HSV (e.g., HSV-1 and/or HSV-2) proteins, and various methods of production and/or use relating thereto, as well as compositions developed therewith and methods relating thereto.
  • the present disclosure provides technologies for preventing, characterizing, treating, and/or monitoring HSV (e.g., HSV-1 and/or HSV-2) outbreaks and/or infections including, as noted, various nucleic acid constructs and encoded proteins, as well as agents (e.g., antibodies) that bind to such proteins, and compositions that comprise and/or deliver them.
  • HSV e.g., HSV-1 and/or HSV-2
  • agents e.g., antibodies
  • technologies provided herein are designed to augment, induce, promote, enhance and/or improve immunological memory against HSV (e.g., HSV-1 and/or HSV-2) or a component thereof (e.g., a protein or fragment thereof).
  • technologies described herein are designed to act as an immunological boost to a primary composition (e.g., immunogenic composition, e.g., vaccine), such as a composition (e.g., immunogenic composition, e.g., vaccine) directed to an antigen and/or epitopes of HSV (e.g., HSV-1 and/or HSV-2).
  • compositions of the present disclosure comprise one or more polynucleotide constructs (e.g., one or more string constructs) that encode one or more antigens from HSV (e.g., HSV-1 and/or HSV-2).
  • HSV e.g., HSV-1 and/or HSV-2
  • the present disclosure provides compositions comprising nucleic acids encoding such HSV (e.g., HSV-1 and/or HSV-2) antigens; those skilled in the art will appreciate from context when reference to a particular polynucleotide (e.g., a DNA or RNA) as “encoding” such antigens in fact is referencing a coding strand or its complement.
  • compositions e.g., immunogenic compositions, e.g., vaccines
  • a subject e.g., a patient
  • related technologies e.g., methods
  • pharmaceutical compositions e.g., immunogenic compositions, e.g., vaccines
  • deliver particular HSV-1 antigen constructs to a subject e.g., a patient
  • related technologies e.g., methods
  • the present disclosure provides pharmaceutical compositions (e.g., immunogenic compositions, e.g., vaccines) that deliver particular HSV-2 antigen constructs to a subject (e.g., a patient) and related technologies (e.g., methods).
  • pharmaceutical compositions e.g., immunogenic compositions, e.g., vaccines
  • the present disclosure provides pharmaceutical compositions (e.g., immunogenic compositions, e.g., vaccines) that deliver particular HSV-1 and HSV-2 antigen constructs to a subject (e.g., a patient) and related technologies (e.g., methods).
  • the present disclosure further provides the recognition that some HSV antigens are common to both HSV-1 and HSV-2.
  • HSV antigens include sequences conserved between HSV-1 and HSV-2.
  • HSV-1 antigens have, e.g., at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to comparable HSV-2 antigens.
  • the present disclosure provides some HSV antigen constructs particularly useful in effective vaccination.
  • HSV antigen constructs are HSV-1 antigen construct, HSV-2 antigen constructs, or a combination thereof.
  • Antigens utilized in accordance with the present disclosure are or include HSV (e.g., HSV-1 and/or HSV-2) components (e.g., antigenic fragments thereof, including epitopes that may comprise non-amino acid, e.g., carbohydrate moieties), which components induce immune responses when administered to humans (or other animals such as rodents and non-human primates susceptible to HSV (e.g., HSV-1 and/or HSV-2) infection).
  • HSV e.g., HSV-1 and/or HSV-2
  • components e.g., antigenic fragments thereof, including epitopes that may comprise non-amino acid, e.g., carbohydrate moieties
  • antigens utilized in provided pharmaceutical compositions include both B-cell and T-cell antigens and/or epitopes, as described herein.
  • delivered antigens include both B-cell and T cell (e.g., CD4 and/or CD8 T cell) antigens and/or epitopes, optionally together in a single antigen polypeptide.
  • antigens utilized in provided pharmaceutical compositions include T cell antigens and/or epitopes.
  • antigens utilized in provided pharmaceutical compositions include B cell, CD4 T cell and CD8 T cell epitopes.
  • the present disclosure defines particularly useful epitopes for inclusion in HSV (e.g., HSV-1 and/or HSV-2) compositions (e.g., immunogenic compositions, e.g., vaccines), and/or provides antigens that include them.
  • HSV HSV-1 and/or HSV-2
  • compositions e.g., immunogenic compositions, e.g., vaccines
  • antigens that include them.
  • Exemplary antigens and/or epitopes for use in compositions described herein included those provided in, e.g., Tables 7 and 8 herein and antigenic fragments thereof.
  • exemplary antigens disclosed in Tables 7 and 8, and/or fragments and/or epitopes thereof can be useful for compositions described herein.
  • a provided pharmaceutical composition e.g., immunogenic composition, e.g., HSV (e.g., HSV-1 and/or HSV-2) vaccine
  • comprises or delivers e.g., causes expression of in a recipient organism, for example by administration of a nucleic acid construct, such as an RNA construct as described herein, that encodes it
  • an antigen that is or comprises one or more epitopes (e.g., one or more B-cell and/or one or more T-cell antigens and/or epitopes) of an HSV (e.g., HSV-1 and/or HSV-2) protein.
  • a pharmaceutical composition described herein induces a relevant immune response effective against HSV (e.g., by targeting an HSV-1 protein, an HSV-2 protein, or a combination thereof).
  • a provided pharmaceutical composition e.g., immunogenic composition, e.g., HSV (e.g., HSV-1 and/or HSV-2) vaccine
  • HSV e.g., HSV-1 and/or HSV-2
  • a provided pharmaceutical composition comprises or delivers an antigen that is or comprises a full-length HSV (e.g., HSV-1 and/or HSV-2) protein.
  • a provided pharmaceutical composition e.g., immunogenic composition, e.g., HSV (e.g., HSV-1 and/or HSV-2) vaccine
  • HSV e.g., HSV-1 and/or HSV-2 vaccine
  • an antigen that is or comprises a fragment of an HSV (e.g., HSV-1 and/or HSV-2) protein that is less than a full-length HSV (e.g., HSV-1 and/or HSV-2) protein.
  • a provided pharmaceutical composition e.g., immunogenic composition, e.g., HSV (e.g., HSV-1 and/or HSV-2) vaccine
  • HSV e.g., HSV-1 and/or HSV-2
  • a chimeric polypeptide that is or comprises part or all of an HSV (e.g., HSV-1 and/or HSV-2) protein and one or more heterologous polypeptide elements.
  • an antigen that is included in and/or delivered by a provided pharmaceutical composition is or comprises one or more peptide fragments of an HSV (e.g., HSV-1 and/or HSV-2) antigen; in some such embodiments, each of the one or more peptide fragments includes at least one epitope (e.g., one or more B cell epitopes and/or one or more T cell epitopes), for example as may be predicted, selected, assessed and/or characterized as described herein.
  • a T-cell string polypeptide that is included in and/or delivered by a provided pharmaceutical composition e.g., immunogenic composition, e.g., HSV (e.g., HSV-1 and/or HSV-2) vaccine
  • a provided pharmaceutical composition e.g., immunogenic composition, e.g., HSV (e.g., HSV-1 and/or HSV-2) vaccine
  • HSV e.g., HSV-1 and/or HSV-2
  • HSV e.g., HSV-1 and/or HSV-2
  • a single polypeptide antigen may include a plurality of such fragments, e.g., presented as a string of antigens or fragments thereof as described herein (e.g., in that a single polypeptide includes a plurality of amino acid sequences derived from distinct HSV antigens or fragments thereof, optionally separated by or otherwise associated with amino acid linkers or other intervening or terminal amino acid sequences).
  • a single RNA antigen construct may include a plurality of sequences encoding HSV antigens, e.g., presented as a string of antigen encoding sequences as described herein (e.g., in that a single RNA molecule includes a plurality of nucleic acid sequences encoding distinct HSV antigens or fragments thereof, optionally separated by or otherwise associated with nucleic acid linkers or other intervening or terminal nucleic acid sequences).
  • one or more HSV e.g., HSV-1 and/or HSV-2
  • antigens or antigenic fragments thereof may be linked with one or more sequences with which it is linked in nature.
  • such sequence(s) may be or comprise one or more heterologous elements (e.g., one or more elements, not naturally found in the relevant HSV (e.g., HSV-1 and/or HSV-2) such as a polypeptide or antigenic fragment thereof not naturally found to be directly linked to the relevant HSV (e.g., HSV-1 and/or HSV-2) antigen(s)).
  • heterologous elements e.g., one or more elements, not naturally found in the relevant HSV (e.g., HSV-1 and/or HSV-2) such as a polypeptide or antigenic fragment thereof not naturally found to be directly linked to the relevant HSV (e.g., HSV-1 and/or HSV-2) antigen(s)
  • an antigen peptide provided and/or utilized in accordance with the present disclosure may include one or more linker elements and/or one or more membrane association elements and/or one or more secretion elements, etc.
  • an antigenic polypeptide may comprise a plurality of HSV (e.g., HSV-1 and/or HSV-2) protein fragments or epitopes separated from one another by linkers.
  • HSV e.g., HSV-1 and/or HSV-2
  • an HSV e.g., HSV-1 and/or HSV-2 polypeptide, or fragment or epitope thereof, utilized in a construct as described herein (or encoded by a polyribonucleotide describe herein
  • a utilized antigen may include one or more sequence variations found in circulating strains or predicted to arise, e.g., in light of assessments of sequence conservation and/or evolution of HSV (e.g., HSV-1 and/or HSV-2) polypeptides over time and/or across strains.
  • a utilized antigen may include one or more sequence variations selected, for example, to impact stability, folding, processing and/or display of the antigen or any epitope thereof.
  • an HSV e.g., HSV-1 and/or HSV-2
  • an antigen as described herein shows at least 70%, 71%, 72%, 73%, 74%, 75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% amino acid sequence identity with a relevant corresponding reference (e.g., wild type) polypeptide, fragment or epitope.
  • a relevant corresponding reference e.g., wild type
  • an HSV e.g., HSV-1 and/or HSV-2
  • an antigen as described herein shows at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence homology (i.e., identity or conservative substitution as is understood in the art) amino acid sequence identity with a relevant corresponding reference (e.g., wild type) protein, fragment or epitope.
  • sequence homology i.e., identity or conservative substitution as is understood in the art
  • an HSV e.g., HSV-1 and/or HSV-2
  • a relevant corresponding reference e.g., wild type polypeptide, fragment or epitope.
  • assessments of degree of conservation may consider the physiochemical difference between two amino acids as described, for example, in WO2014/180569, which is incorporated herein by reference in its entirety. It is well known in molecular evolution that amino acids that interchange frequently are likely to have chemical and physical similarities whereas amino acids that interchange rarely are likely to have different physico-chemical properties. The likelihood for a given substitution to occur in nature compared with the likelihood for this substitution to occur by chance can measured by log-odds matrices.
  • substitutions with positive T scores would have a lower likelihood of altering immunogenicity.
  • substitutions with negative T scores reflect substitutions that are unlikely to occur in nature and hence correspond to two amino acids that have significantly different physico-chemical properties. Such substitutions would have a greater chance of altering immunogenicity.
  • presence of negative T score substitutions within a sequence may indicate that it would be relatively less useful in a composition antigen as described herein.
  • an HSV antigen construct includes and/or encodes a plurality of HSV antigens (e.g., a plurality of HSV antigens that are or include one or more T cell antigens for HSV) provided in Table 7, or fragments thereof.
  • a plurality of HSV antigens e.g., a plurality of HSV antigens that are or include one or more T cell antigens for HSV
  • an HSV antigen construct can include and/or encode at least one HSV antigen provided in Table 7, or fragments thereof.
  • an HSV antigen construct can include and/or encode at least one T cell antigen for HSV provided in Table 7, or fragments thereof.
  • an HSV antigen construct can include and/or encode a plurality of (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19) HSV antigens provided in Table 7, or fragments thereof.
  • an HSV antigen construct can include and/or encode a plurality of (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19) T cell antigens for HSV selected from HSV antigens provided in Table 7, or fragments thereof.
  • an antigen utilized in accordance with the present disclosure includes HSV (e.g., HSV-1 and/or HSV-2) protein sequences identified and/or characterized by one or more of: 1) HLA-I or HLA-II binding (e.g., to HLA allele(s) present in a relevant population) 2) HLA ligandomics data, optionally confirmed by mass spectrometry 3) Relatively high expression 4) Sequence conservation 5) Surface exposure 6) Serum reactivity 7) Immunogenicity (e.g., presence of one or more B-cell and/or T-cell antigens and/or epitopes; evidence of ability to induce sterile protection in model systems including, e.g., humans, non-human primates, and/or mice).
  • HSV e.g., HSV-1 and/or HSV-2
  • HLA-II binding e.g., to HLA allele(s) present in a relevant population
  • HLA ligandomics data optionally confirmed by mass spectrometry
  • HLA-I and/or HLA-II binding is experimentally assessed; in some embodiments it is predicted.
  • predicted HLA-I or HLA-II binding is assessed using an algorithm such as neonmhc 1 and/or neonmhc2, which predict and/or characterize likelihood of MHC class I and MHC class II binding, respectively.
  • an MHC-peptide presentation prediction algorithm or MHC-peptide presentation predictor is or comprises NetMHCpan or NetMHCIIpan.
  • a hidden Markov model approach may be utilized for MHC-peptide presentation prediction and/or characterization.
  • the peptide prediction model MARIA may be utilized.
  • NetMHCpan is not utilized to predict or characterize likelihood of MHC binding for peptides as described herein.
  • the peptide prediction model MARIA may be utilized.
  • NetMHCIIpan is not utilized to predict or characterize likelihood of MHC binding for peptides as described herein.
  • neither NetMHCpan nor NetMHCIIpan is utilized to predict or characterize likelihood of MHC binding for peptides as described herein.
  • an MHC-peptide presentation prediction algorithm or MHC-peptide presentation predictor is or comprises RECON ® (Real-time Epitope Computation for ONcology), which offers high quality MHC-peptide presentation prediction based on expression, processing and binding capabilities. See, for example, Abelin et al., Immunity 21:315, 2017; Abelin et al., Immunity 15:766, 2019. [0329] In some embodiments, HLA binding and/or ligandomics assessments will consider the geographic region of subjects to be immunized. For example, in some embodiments, HLA allelic diversity will be considered.
  • antigen(s) included in a provided pharmaceutical composition will be or comprise peptides (e.g., epitopes) expected or determined, when considered together, to bind to a significant percentage (e.g., at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or more) of HLA alleles expected or known to be present in a relevant region or population.
  • antigen(s) included in a provided pharmaceutical composition will be or comprise peptides expected or determined, when considered together, to bind to the most prevalent (e.g., the 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 most prevalent, or at least 1, 2, 3, 4, or 5 of the 10 most prevalent, etc.) HLA alleles expected or known to be present in a relevant region or population).
  • expression level is experimentally determined (e.g., in a model system or in infected humans). In some embodiments, expression level is a reported level (e.g., in a published or presented report).
  • expression level is assessed as RNA (e.g., via RNASeq). In some embodiments (and typically preferably), expression levels is assessed as protein.
  • sequence conservation is assessed, for example, using publicly available sequence evaluation software (such as, for example, multiple sequence alignment programs MAFFT, Clustal Omega, etc.). In some embodiments, sequence conservation is determined by consultation with published resources (e.g., sequences). In some embodiments, sequence conservation includes consideration of currently or recently detected strains (e.g., in an active outbreak).
  • surface exposure is assessed by reference to publicly available database and/or software.
  • serum reactivity is assessed by contacting serum samples from infected individuals with polypeptides including sequences of interest (e.g., as may be displayed via, for example, phage display or peptide array, etc.; see, for example, Whittemore et al “ A General Method to Discover Epitopes from Sera” PlosOne, 2016; https://doi.org/10.1371/journal.pone.0157462).
  • serum reactivity is assessed by consultation with literature reports and or database data indicating serum-recognized sequences.
  • assessment of immunoreactivity and/or of presence of an epitope may be or comprise consultation with the Immune Epitope Database (IEDB) which those skilled in the art will be aware is a freely available resource funded by NIAID that catalogs experimental data on antibody and T cell epitopes (see iedb.org).
  • IEDB Immune Epitope Database
  • antigen(s) utilized in accordance with the present disclosure are characterized by dendritic cell presentation which, in turn may be indicative of HLA binding and/or of immunogenicity.
  • antigen(s) utilized in accordance with the present disclosure are or comprises sequences (e.g., epitopes, fragments, complete proteins) of HSV proteins found in the HSV envelope.
  • antigen(s) utilized in accordance with the present disclosure are or comprises sequences (e.g., epitopes, fragments, complete proteins) of HSV proteins found in the HSV tegument.
  • sequences e.g., epitopes, fragments, complete proteins
  • the present disclosure provides an insight that, in some embodiments, it may be desirable to include two or more different epitopes, optionally from two or more different HSV (e.g., HSV-1 and/or HSV-2) proteins, in pharmaceutical composition (e.g., immunogenic composition, e.g., vaccine) compositions, which can be useful in the treatment of HSV.
  • Table 8 Exemplary antigen fragment Antigen Sequence (Amino Acid) SEQ ID Fragment NO: RL2.1 CTDEIAPPLRCQSFPCLHPFCIPCMKTWIPLRNTCPLCNTPVAYLIVGVTASGSFSTIPIVN 562 Fragment DPRTRVEAEAAVRAGTAVDFIWTGNPRTAPRSLS RL2.2 LPIAGVSSVVALAPYVNKTVTGDCLPVLDMETGHIGAYVVLVDQTGNVADLLRAAAPAWS 563 Fragment RRTLLPEHARNCVRPPDYPTPPASEWNSLWMTPVGNMLFDQGTLVG RS1 RAAAWMRQVPDPEDVRVVILYSPLPGEDLAAGRAGGGPPPEWSAERGGLSCLLAALGNR 564 Fragment LCGPATAAWAGNWTGAPDVSALGAQGVLLLSTRDLAFAGAVEFLGLLAGACDRRLIVVN AVRAADWPADGPVVSRQHAYLACEVLPAVQCAVRWPAARDLRRTVLASGRVFGPGV
  • a polyribonucleotide provided herein encodes one or more of an RL2 polypeptide or antigenic fragment thereof, an RS1 polypeptide or antigenic fragment thereof, a UL54 polypeptide or antigenic fragment thereof, a UL29 polypeptide or antigenic fragment thereof, a UL39 polypeptide or antigenic fragment thereof, a UL49 polypeptide or antigenic fragment thereof, a UL9 polypeptide or antigenic fragment thereof, a UL30 polypeptide or antigenic fragment thereof, a UL40 polypeptide or antigenic fragment thereof, a UL5 polypeptide or antigenic fragment thereof, a UL52 polypeptide or antigenic fragment thereof, a UL1 polypeptide or antigenic fragment thereof, a UL19 polypeptide or antigenic fragment thereof, a UL21 polypeptide or antigenic fragment thereof, a UL27 polypeptide or antigenic fragment thereof, a UL46 polypeptide or antigenic fragment thereof, a UL47
  • a polyribonucleotide provided herein encodes a polypeptide comprising one or more HSV RL2 polypeptides or antigenic fragments thereof, one or more HSV RS1 polypeptides or antigenic fragments thereof, and one or more HSV UL54 polypeptides or antigenic fragments thereof.
  • a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an RL2 polypeptide or antigenic fragment thereof, a linker, an RL2 polypeptide or antigenic fragment thereof, a linker, an RS1 polypeptide or antigenic fragment thereof, a linker, a UL54 polypeptide or antigenic fragment thereof, and a linker.
  • a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an UL54 polypeptide or antigenic fragment thereof, a linker, an RS1 polypeptide or antigenic fragment thereof, a linker, an RL2 polypeptide or antigenic fragment thereof, a linker, a RL2 polypeptide or antigenic fragment thereof, and a linker.
  • a polyribonucleotide provided herein encodes a polypeptide comprising one or more HSV UL29 polypeptides or antigenic fragments thereof, one or more HSV UL39 polypeptides or antigenic fragments thereof, one or more HSV UL49 polypeptides or antigenic fragments thereof, and one or more HSV UL9 polypeptides or antigenic fragments thereof.
  • a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an UL29 polypeptide or antigenic fragment thereof, a linker, an UL39 polypeptide or antigenic fragment thereof, a linker, an UL49 polypeptide or antigenic fragment thereof, a linker, a UL9 polypeptide or antigenic fragment thereof, and a linker.
  • a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an UL9 polypeptide or antigenic fragment thereof, a linker, an UL49 polypeptide or antigenic fragment thereof, a linker, an UL39 polypeptide or antigenic fragment thereof, a linker, a UL29 polypeptide or antigenic fragment thereof, and a linker.
  • a polyribonucleotide provided herein encodes a polypeptide comprising one or more HSV UL30 polypeptides or antigenic fragments thereof, one or more HSV UL40 polypeptides or antigenic fragments thereof, one or more HSV UL5 polypeptides or antigenic fragments thereof, and one or more HSV UL52 polypeptides or antigenic fragments thereof.
  • a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an UL30 polypeptide or antigenic fragment thereof, a linker, an UL30 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, a UL5 polypeptide or antigenic fragment thereof, a linker, a UL5 polypeptide or antigenic fragment thereof, a linker, a UL52 polypeptide or antigenic fragment thereof, and a linker.
  • a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an UL52 polypeptide or antigenic fragment thereof, a linker, an UL5 polypeptide or antigenic fragment thereof, a linker, an UL5 polypeptide or antigenic fragment thereof, a linker, a UL40 polypeptide or antigenic fragment thereof, a linker, a UL30 polypeptide or antigenic fragment thereof, a linker, a UL30 polypeptide or antigenic fragment thereof, and a linker.
  • a polyribonucleotide provided herein encodes a polypeptide comprising one or more HSV UL1 polypeptides or antigenic fragments thereof, one or more HSV UL19 polypeptides or antigenic fragments thereof, one or more HSV UL21 polypeptides or antigenic fragments thereof, one or more HSV UL27 polypeptides or antigenic fragments thereof, one or more HSV UL46 polypeptides or antigenic fragments thereof, one or more HSV UL47 polypeptides or antigenic fragments thereof, one or more UL48 polypeptides or antigenic fragments thereof, and one or more HSV UL25 polypeptides or antigenic fragments thereof.
  • a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an HSV UL1 polypeptide or antigenic fragment thereof, a linker, an HSV UL19 polypeptide or antigenic fragment thereof, a linker, an HSV UL21 polypeptide or antigenic fragment thereof, a linker, an HSV UL27 polypeptide or antigenic fragment thereof, a linker, an HSV UL27 polypeptide or antigenic fragment thereof, a linker, an HSV UL46 polypeptide or antigenic fragment thereof, a linker, an HSV UL47 polypeptide or antigenic fragment thereof, a linker, an HSV UL25 polypeptide or antigenic fragment thereof, a linker, an HSV UL48 polypeptide or antigenic fragment thereof, and a linker.
  • a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, nucleotide sequences that encode an HSV-1 gD secretory signal, an HSV UL48 polypeptide or antigenic fragment thereof, a linker, an HSV UL25 polypeptide or antigenic fragment thereof, a linker, an HSV UL47 polypeptide or antigenic fragment thereof, a linker, an HSV UL46 polypeptide or antigenic fragment thereof, a linker, an HSV UL27 polypeptide or antigenic fragment thereof, a linker, an HSV UL27 polypeptide or antigenic fragment thereof, a linker, an HSV UL21 polypeptide or antigenic fragment thereof, a linker, an HSV UL19 polypeptide or antigenic fragment thereof, a linker, an HSV UL1 polypeptide or antigenic fragment thereof, and a linker, and a MITD.
  • a polyribonucleotide provided herein encodes a polypeptide comprising, one or more HSV US1 polypeptides or antigenic fragments thereof, one or more HSV US8 polypeptides or antigenic fragments thereof, one or more HSV US12 polypeptides or antigenic fragments thereof, one or more HSV UL50 polypeptides or antigenic fragments thereof, one or more HSV UL26 polypeptides or antigenic fragments thereof, and one or more HSV US10 polypeptides or antigenic fragments thereof.
  • a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an US1 polypeptide or antigenic fragment thereof, a linker, an US1 polypeptide or antigenic fragment thereof, a linker, an US8 polypeptide or antigenic fragment thereof, a linker, a US8 polypeptide or antigenic fragment thereof, a linker, a US8 polypeptide or antigenic fragment thereof, a linker, a US12 polypeptide or antigenic fragment thereof, a linker, a UL50 polypeptide or antigenic fragment thereof, a linker, a UL26 polypeptide or antigenic fragment thereof, a linker, a UL26 polypeptide or antigenic fragment thereof, a linker, a US10 polypeptide or antigenic fragment thereof, and a linker.
  • a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an UL26 polypeptide or antigenic fragment thereof, a linker, an UL26 polypeptide or antigenic fragment thereof, a linker, an US10 polypeptide or antigenic fragment thereof, a linker, a UL50 polypeptide or antigenic fragment thereof, a linker, a US12 polypeptide or antigenic fragment thereof, a linker, a US8 polypeptide or antigenic fragment thereof, a linker, a US8 polypeptide or antigenic fragment thereof, a linker, a US8 polypeptide or antigenic fragment thereof, a linker, a US8 polypeptide or antigenic fragment thereof, a linker, a US8 polypeptide or antigenic fragment thereof, a linker, a US1 polypeptide or antigenic fragment thereof, a linker, a US1 polypeptide or antigenic fragment thereof, and a linker.
  • a polyribonucleotide provided herein encodes a polypeptide comprising one or more HSV RL2 polypeptides or antigenic fragments thereof, one or more HSV UL54 polypeptides or antigenic fragments thereof, one or more HSV UL47 polypeptides or antigenic fragments thereof, one or more HSV UL46 polypeptides or antigenic fragments thereof, and one or more HSV UL21 polypeptides or antigenic fragments thereof.
  • a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an RL2 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, a UL46 polypeptide or antigenic fragment thereof, a linker, a UL21 polypeptide or antigenic fragment thereof, and a linker.
  • a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an UL21 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, an RL2 polypeptide or antigenic fragment thereof, and a linker.
  • a polyribonucleotide provided herein encodes a polypeptide comprising one or more HSV UL29 polypeptides or antigenic fragments thereof, one or more HSV UL39 polypeptides or antigenic fragments thereof, one or more HSV UL9 polypeptides or antigenic fragments thereof, one or more HSV UL5 polypeptides or antigenic fragments thereof, one or more HSV UL40 polypeptides or antigenic fragments thereof, and one or more HSV UL30 polypeptides or antigenic fragments thereof.
  • a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an UL29 polypeptide or antigenic fragment thereof, a linker, an UL39 polypeptide or antigenic fragment thereof, a linker, an UL9 polypeptide or antigenic fragment thereof, a linker, an UL5.1 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL30.1 polypeptide or antigenic fragment thereof, and a linker.
  • a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an UL30.1 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL5.1 polypeptide or antigenic fragment thereof, a linker, an UL9 polypeptide or antigenic fragment thereof, a linker, an UL39 polypeptide or antigenic fragment thereof, a linker, an UL29 polypeptide or antigenic fragment thereof, and a linker.
  • a polyribonucleotide provided herein encodes a polypeptide comprising one or more HSV RL2 polypeptides or antigenic fragments thereof, one or more HSV UL54 polypeptides or antigenic fragments thereof, one or more HSV UL9 polypeptides or antigenic fragments thereof, one or more HSV UL39 polypeptides or antigenic fragments thereof, and one or more HSV UL5 polypeptides or antigenic fragments thereof.
  • a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an RL2.1 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, an UL9 polypeptide or antigenic fragment thereof, a linker, a UL39 polypeptide or antigenic fragment thereof, a linker, a UL5.1 polypeptide or antigenic fragment thereof, and a linker.
  • a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an UL5.1 polypeptide or antigenic fragment thereof, a linker, an UL39 polypeptide or antigenic fragment thereof, a linker, an UL9 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, an RL2.1 polypeptide or antigenic fragment thereof, and a linker.
  • a polyribonucleotide provided herein encodes a polypeptide comprising one or more HSV UL47 polypeptides or antigenic fragments thereof, one or more HSV UL46 polypeptides or antigenic fragments thereof, one or more HSV UL21 polypeptides or antigenic fragments thereof, one or more HSV UL5.2 polypeptides or antigenic fragments thereof, one or more HSV UL40 polypeptides or antigenic fragments thereof, one or more HSV UL30.1 polypeptides or antigenic fragments thereof, and one or more HSV UL29 polypeptides or antigenic fragments thereof.
  • a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL21 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL30.1 polypeptide or antigenic fragment thereof, a linker, an UL29 polypeptide or antigenic fragment thereof, and a linker.
  • a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an UL29 polypeptide or antigenic fragment thereof, a linker, an UL30.1 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL21 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, and a linker.
  • a polyribonucleotide provided herein encodes a polypeptide comprising one or more HSV RL2.1 polypeptides or antigenic fragments thereof, one or more HSV UL54 polypeptides or antigenic fragments thereof, one or more HSV UL5.2 polypeptides or antigenic fragments thereof, one or more HSV UL40 polypeptides or antigenic fragments thereof, one or more HSV UL47 polypeptides or antigenic fragments thereof, and one or more HSV UL46 polypeptides or antigenic fragments thereof.
  • a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, a RL2.1 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, and a linker.
  • a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, a RL2.1 polypeptide or antigenic fragment thereof, and a linker.
  • a polyribonucleotide provided herein encodes a polypeptide comprising one or more HSV RL2.1 polypeptides or antigenic fragments thereof, one or more HSV RS1 polypeptides or antigenic fragments thereof, and one or more HSV UL54 polypeptides or antigenic fragments thereof.
  • a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, a RL2.1 polypeptide or antigenic fragment thereof, a linker, a RS1 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, and a linker.
  • a polyribonucleotide provided herein encodes a polypeptide comprising one or more HSV UL29 polypeptides or antigenic fragments thereof, one or more HSV UL39 polypeptides or antigenic fragments thereof, and one or more HSV UL9 polypeptides or antigenic fragments thereof.
  • a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an UL29 polypeptide or antigenic fragment thereof, a linker, an UL39 polypeptide or antigenic fragment thereof, a linker, an UL9 polypeptide or antigenic fragment thereof, and a linker.
  • a polyribonucleotide provided herein encodes a polypeptide comprising one or more HSV UL5.1 polypeptides or antigenic fragments thereof, one or more HSV UL5.2 polypeptides or antigenic fragments thereof, one or more HSV UL40 polypeptides or antigenic fragments thereof, and one or more HSV UL30.1 polypeptides or antigenic fragments thereof.
  • a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an UL5.1 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL30.1 polypeptide or antigenic fragment thereof, and a linker.
  • a polyribonucleotide provided herein encodes a polypeptide comprising one or more HSV UL47 polypeptides or antigenic fragments thereof, one or more HSV UL46 polypeptides or antigenic fragments thereof, and one or more HSV UL21 polypeptides or antigenic fragments thereof.
  • a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL21 polypeptide or antigenic fragment thereof, and a linker.
  • a polyribonucleotide provided herein encodes a polypeptide comprising one or more HSV UL29 polypeptides or antigenic fragments thereof, one or more HSV UL39 polypeptides or antigenic fragments thereof, one or more HSV UL9 polypeptides or antigenic fragments thereof, one or more HSV UL5.1 polypeptides or antigenic fragments thereof, one or more HSV UL5.2 polypeptides or antigenic fragments thereof, one or more HSV UL40 polypeptides or antigenic fragments thereof, and one or more HSV UL30.1 polypeptides or antigenic fragments thereof.
  • a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an UL29 polypeptide or antigenic fragment thereof, a linker, an UL39 polypeptide or antigenic fragment thereof, a linker, an UL9 polypeptide or antigenic fragment thereof, a linker, an UL5.1 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL30.1 polypeptide or antigenic fragment thereof, and a linker.
  • a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an UL30.1 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL5.1 polypeptide or antigenic fragment thereof, a linker, an UL9 polypeptide or antigenic fragment thereof, a linker, an UL39 polypeptide or antigenic fragment thereof, a linker, an UL29 polypeptide or antigenic fragment thereof, and a linker.
  • a polyribonucleotide provided herein encodes a polypeptide comprising one or more HSV RL2.1 polypeptides or antigenic fragments thereof, one or more HSV RS1 polypeptides or antigenic fragments thereof, one or more HSV UL54 polypeptides or antigenic fragments thereof, one or more HSV UL47 polypeptides or antigenic fragments thereof, one or more HSV UL46 polypeptides or antigenic fragments thereof, and one or more HSV UL21 polypeptides or antigenic fragments thereof.
  • a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, a RL2.1 polypeptide or antigenic fragment thereof, a linker, a RS1 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL21 polypeptide or antigenic fragment thereof, and a linker.
  • a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an UL21 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, a RS1 polypeptide or antigenic fragment thereof, a linker, a RL2.1 polypeptide or antigenic fragment thereof, and a linker.
  • a polyribonucleotide provided herein encodes a polypeptide comprising one or more HSV UL5.1 polypeptides or antigenic fragments thereof, one or more HSV UL5.2 polypeptides or antigenic fragments thereof, one or more HSV UL40 polypeptides or antigenic fragments thereof, one or more HSV UL30.1 polypeptides or antigenic fragments thereof, one or more HSV UL47 polypeptides or antigenic fragments thereof, one or more HSV UL46 polypeptides or antigenic fragments thereof, and one or more HSV UL21 polypeptides or antigenic fragments thereof.
  • a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an UL5.1 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL30.1 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL21 polypeptide or antigenic fragment thereof, and a linker.
  • a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an UL21 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL30.1 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL5.1 polypeptide or antigenic fragment thereof, and a linker.
  • a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is an RL2 polypeptide or antigenic fragment thereof.
  • an RL2 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of CTDEIAPPLRCQSFPCLHPFCIPCMKTWIPLRNTCPLCNTPVAYLIVGVTASGSFSTIPIVNDPRTRVEAEAAVRAGTAVDFIWTGNPR TAPRSLS (SEQ ID NO: 562).
  • an RL2 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of LPIAGVSSVVALAPYVNKTVTGDCLPVLDMETGHIGAYVVLVDQTGNVADLLRAAAPAWSRRTLLPEHARNCVRPPDYPTPPASEWN SLWMTPVGNMLFDQGTLVG (SEQ ID NO: 563).
  • a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is an RS1 polypeptide or antigenic fragment thereof.
  • an RS1 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of RAAAWMRQVPDPEDVRVVILYSPLPGEDLAAGRAGGGPPPEWSAERGGLSCLLAALGNRLCGPATAAWAGNWTGAPDVSALGAQ GVLLLSTRDLAFAGAVEFLGLLAGACDRRLIVVNAVRAADWPADGPVVSRQHAYLACEVLPAVQCAVRWPAARDLRRTVLASGRVF GPGVFARVEAAHARLYPDAPPLRLCRGANVRYRVRTRFGPDTLVPMSPREYRRAVLPALDGRAAAS (SEQ ID NO: 564).
  • a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is a UL54 polypeptide or antigenic fragment thereof.
  • a UL54 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of ETLVAHGPSLYRTFAANPRAASTAKAMRDCVLRQENLIEALASADETLAWCKMCIHHNLPLRPQDPIIGTAAAVLENLATRLRPFLQC YLKARGLCGLDDLCSRRRLSDIKDIASFVLVILARLANRVERGVSEIDYTTVGVGAGETMHFYIPGACMAGLIEILDTHRQECSSRVCE LTASHTIAPLYVHGKYFYCNSLF (SEQ ID NO: 565).
  • a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is a UL29 polypeptide or antigenic fragment thereof.
  • HSV T-cell antigens or antigenic portions thereof e.g., HSV-1 and/or HSV- 2
  • a UL29 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of REDIETIAFIKRFSLDYGAINFINLAPNNVSELAMYYMANQILRYCDHSTYFINTLTAVIAGSRRPPSVQAAAAWAPQGGAGLEAGAR ALMDSLDAHPGAWTSMFASCNLLRPVMAARPMVVLGLSISKYYGMAGNDRVFQAGNWASLLGGKNACPLLIFDRTRKFVL (SEQ ID NO: 566).
  • a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is a UL39 polypeptide or antigenic fragment thereof.
  • a UL39 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of RTFGSAPRLTEDDFGLLNYALAEMRRLCLDLPPVPPNAYTPYHLREYATRLVNGFKPLVRRSARLYRILGVLVHLRIRTREASFEEWM RSKEVDLDFGLTERLREHEAQLMILAQALNPYDCLIHSTPNTLVERGLQSALKYEEFYLKRFGGHYMESVFQMYTRIAGFLA (SEQ ID NO: 567).
  • a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is a UL49 polypeptide or antigenic fragment thereof.
  • a UL49 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of KMTRGAPKASATPATDPARGRRPAQADSAVLLDAPAPTASGRTKTPAQGLAKKLHFSTAPPSPTAPWTPRVAGFNKRVFCAAVG (SEQ ID NO: 568).
  • a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is a UL9 polypeptide or antigenic fragment thereof.
  • a UL9 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of LLNNYDVLVLDEVMSTLGQLYSPTMQQLGRVDALMLRLLRTCPRIIAMDATANAQLVDFLCSLRGEKNVHVVIGEYAMPGFSARRCL FLPRLGPEVLQAALRRRGPAGGAPPPDAPPDATFFGELEARLAGGDNVCIFSSTVSFAEVVARFCRQFTDRVLLLHSLTPPGDVTTWG RYRVVIYTTVVTVGLSFDPPHFDSMFAYVKPMNYGPDMVSVYQSLGRVRTLRKGELLIYMDGSGARSEPV (SEQ ID NO: 569).
  • a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is a UL30 polypeptide or antigenic fragment thereof.
  • HSV T-cell antigens or antigenic portions thereof e.g., HSV-1 and/or HSV- 2
  • a UL30 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of ISCLLYDLSTTALEHILLFSLGSCDLPESHLSDLASRGLPAPVVLEFDSEFEMLLAFMTFVKQYGPEFVTGYNIINFDWPFVLTKLTEIYK VPLDGYGRMNGRGVFRVWDIGQSHF (SEQ ID NO: 570).
  • a UL30 polypeptide or antigenic fragment thereof comprises an amino acid sequence of GLLPCLHVAATVTTIGREMLLATRAYVHARWAEFDQLLADFPEAAGMRAPGPYSM (SEQ ID NO: 571).
  • a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is a UL40 polypeptide or antigenic fragment thereof.
  • a UL40 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of TSQCPDINHLRSLSILNRWLETELVFVGDEEDVSKLSEGELGFYRFLFAFLSAADDLVTENLGGLSGLFEQKDILHYYVEQECIEVVHS RVYNIIQLVLFHNNDQARRAYVARTINHPAIRVKVDWLEARVRECDSIPEKFILMILIEGVFFAASFAAIAYLRTNNLLR (SEQ ID NO: 572).
  • a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is a UL5 polypeptide or antigenic fragment thereof.
  • a UL5 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of HEFGNLMKVLEYGLPITEEHMQFVDRFVVPESYITNPANLPGWTRLFSSHKEVSAYMAKLHAYLKVTREGEFVVFTLPVLTFVSVKEF DEYRRL (SEQ ID NO: 573).
  • a UL5 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of ELFGEVFESAPFSTYVDNVIFRGCELLTGSPRGGLMSVALQTDNYTLMGYTYTRVFAFAEELRRRHATAGVAEFLEESPLPYIVLRDQ HGFMSVVNTNI (SEQ ID NO: 574).
  • a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is a UL52 polypeptide or antigenic fragment thereof.
  • a UL52 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of SVAAPVEVTALYATDGCVITSSLALLTNCLLGAEPLYIFSYDAYRSDAPNGPTGAPTEQERFEGSRALYRDAGGLNGDSFRVTFCLLGT EVGVTHHPKGRTRPMFVCRFERADDVAVLQDALGRGTPLLPAHVTATLDLEATFALHANIIMALTVAIVHNAPARIGSGSTAPLYEPG ESMRSVV (SEQ ID NO: 575).
  • a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is a UL1 polypeptide or antigenic fragment thereof.
  • a UL1 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of RTPADDVSWRYEAPSVIDYARIDGIFLRYHCPGLDTFLWDRHAQRAYLVNPFLFAAGFLEDLSHSVFPADTQETT (SEQ ID NO: 576).
  • a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is a UL19 polypeptide or antigenic fragment thereof.
  • a UL19 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of DGRLLHNTQARAADAADDRPHRPADWTVHHKIYYYVLVPAFSRGRCCTAGVRFDRVYATLQNMVVPEIAPGEECPSDPVTDPAHPL HPANLVANTVKRMFHN (SEQ ID NO: 577).
  • a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is a UL21 polypeptide or antigenic fragment thereof.
  • a UL21 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of SPTQKLAVYYYLIHRERRMSPFPALVRLVGRYIQRHGLYVPAPDEPTLADAMNGL (SEQ ID NO: 578).
  • a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is a UL27 polypeptide or antigenic fragment thereof.
  • a UL27 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of NYTEGIAVVFKENIAPYKFKATMYYKDVTVSQVWFGHRYSQFMGIFEDRAPVPFEEV (SEQ ID NO: 579).
  • a UL27 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of SVYPYDEFVLATGDFVYMSPFYGYREGSH (SEQ ID NO: 580).
  • a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is a UL46 polypeptide or antigenic fragment thereof.
  • a UL46 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of GLASDPHYDYIRHYASAAKQALGEVELSGGQLSRAILAQYWKYLQTVVPSGLDIPDDPAGDCDPSLHVLLRPTLLPKLLVRAPFKSGA AAAKYAAAVAGLRDAAHRLQQYMFFMRPADPSRPSTDTALRLSELLAYVSVLYHWASWMLWTADKYV (SEQ ID NO: 581).
  • a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is a UL47 polypeptide or antigenic fragment thereof.
  • a UL47 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of GPDAAVFRSSLGSLLYWPGVRALLGRDCRVAARYAGRMTYIATGALLARFNPGAVKCVLPREAAFAGRVL (SEQ ID NO: 582).
  • a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is a UL48 polypeptide or antigenic fragment thereof.
  • a UL48 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of ALFNRLLDDLGFSAGPALCTMLDTWNEDLFSGFPTNADMYRECKFLSTLPSDVIDWGDAHVPERSPIDIRAHGDVAFPTLPATRDEL PSYYEAMAQFFRGELRA (SEQ ID NO: 583).
  • a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is a UL25 polypeptide or antigenic fragment thereof.
  • a UL25 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of FLWEDQTLLRATANTITALAVLRRLLANGNVYADRLDNRLQLGMLIPGAVPAEAIARGASGLDSGAIKSGDNNLEALCVNYVLPLYQA DPTVELTQLFPGLAALCL (SEQ ID NO: 584).
  • a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is a US1 polypeptide or antigenic fragment thereof.
  • a US1 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of DDASDGWLVDTPPRKSKRPRINLRLTSSPDRRAGVVFPEV (SEQ ID NO: 585).
  • a US1 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of PASLPGIAHAHRRSARQAQMRSGAAWTLDLHYIRQCVNQL (SEQ ID NO: 586).
  • a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is a US8 polypeptide or antigenic fragment thereof.
  • an US8 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of ILSPTAPSVYPHSEGRKSRRPLTTFGSGSPGRRHSQASYPSVLW (SEQ ID NO: 587).
  • a US8 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of GLAWLASTVNLEFQHASPQHAGLYLCVVYVDDHIHAWGHMTISTAAQYRNAVVEQHLPQRQPEPVEPTRPHVRAPHPAPSARGPL RL (SEQ ID NO: 588).
  • a US8 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of KLLWAAEPLDACGPLRPSWVALWPPRRVLETVVDAACMRAPEPLAIAYSPPFPAGDEGLYSELAWRDRVAVVNESLVIYGALETDSG LYTLSVVGLSDEARQVASVVLVVEPAP (SEQ ID NO: 589).
  • a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is a US12 polypeptide or antigenic fragment thereof.
  • an US12 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of EDREAARTAVTDPELPLLCPPDV (SEQ ID NO: 590).
  • a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is an UL50 polypeptide or antigenic fragment thereof.
  • an UL50 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of ANGATVIQPSLRVLRAADGPEACYVLGRSSLNARGLLVMPTRWPSGHACAFVVCNLTGVPVTL (SEQ ID NO: 591).
  • a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is an UL26 polypeptide or antigenic fragment thereof.
  • an UL26 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of APLPDRAVPIYVAGFLALYDSGDPGELALDPDTVRAALPPENPLPINVDHRARCEVGRVLAVVNDPRGPFFVGLIACVQLERVLETAAS AAIFERRGPALSREERLLYLITNYLPSVSLSTKRRGDEVPPDRTLFAHVALCAIGRRLGTIVTYDTSLDAA (SEQ ID NO: 592).
  • an UL26 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of HYPPPPAHPYPGMLFAGPSPLEAQIAALVGAIAADRQAGGLPAAAGDHGIRGSAKRRRHEVEQPEYDCG (SEQ ID NO: 593).
  • a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is an US10 polypeptide or antigenic fragment thereof.
  • an US10 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of SSPRQRTYVLPRVGIHNAPASDTRAPKRANSRHRADRPPESPGSELYPLNAQALAHLQMLPADHRAFFRTVIEVSRLCALNTHDPPPP LAGARVGQEAQLVHTQWLRANRESSPLWPWRTAAMNFIAAAAPCVQTHRHMHDLLMACAFWC (SEQ ID NO: 594).
  • an HSV e.g., HSV-1 and/or HSV-2
  • T-cell antigen for use in accordance with the present disclosure is an intermediate early protein or an antigenic fragment thereof.
  • a polyribonucleotide provided herein encodes one or more HSV (e.g., HSV-1 and/or HSV-2) antigens, wherein at least one HSV T-cell antigen comprises an intermediate early protein or an antigenic fragment thereof.
  • a polyribonucleotide provided herein encodes one or more HSV (e.g., HSV-1 and/or HSV-2) T-cell antigens, wherein each of the HSV T-cell antigens comprises an intermediate early protein or an antigenic fragment thereof.
  • an HSV-2 T-cell antigen for use in accordance with the present disclosure is an intermediate early protein or an antigenic fragment thereof.
  • a polyribonucleotide provided herein encodes one or more HSV-2 antigens, wherein at least one HSV-2 antigen comprises an intermediate early protein or an antigenic fragment thereof. In some embodiments, a polyribonucleotide provided herein encodes one or more HSV-2 antigens, wherein each of the HSV-2 antigens comprises an intermediate early protein or an antigenic fragment thereof.
  • an HSV-2 T-cell antigen for use in accordance with the present disclosure comprises an RL2 polypeptide or antigenic fragment thereof, an RS1 polypeptide or antigenic fragment thereof, a UL54 polypeptide or antigenic fragment thereof, or a combination thereof.
  • a polyribonucleotide provided herein encodes one or more of an RL2 polypeptide or antigenic fragment thereof, an RS1 protein or antigenic fragment thereof, and a UL54 protein or antigenic fragment thereof.
  • an HSV e.g., HSV-1 and/or HSV-2
  • T-cell antigen for use in accordance with the present disclosure is an early protein or an antigenic fragment thereof.
  • a polyribonucleotide provided herein encodes one or more HSV (e.g., HSV-1 and/or HSV-2) T-cell antigens, wherein at least one HSV T-cell antigen comprises an early protein or an antigenic fragment thereof.
  • a polyribonucleotide provided herein encodes one or more HSV (e.g., HSV-1 and/or HSV-2) T-cell antigens, wherein each of the HSV T-cell antigens comprises an early protein or an antigenic fragment thereof.
  • HSV e.g., HSV-1 and/or HSV-2
  • an HSV (e.g., HSV-1, HSV-2, or a combination) T-cell antigen for use in accordance with the present disclosure is an early protein or an antigenic fragment thereof.
  • a polyribonucleotide provided herein encodes one or more HSV T-cell antigens, wherein at least one HSV T-cell antigen comprises an early protein or an antigenic fragment thereof.
  • a polyribonucleotide provided herein encodes one or more HSV T-cell antigens, wherein each of the HSV T-cell antigens comprises an early protein or an antigenic fragment thereof.
  • the HSV T-cell antigen is a HSV-2 T-cell antigen.
  • an HSV (e.g., HSV-1, HSV-2, or a combination) T-cell antigen for use in accordance with the present disclosure comprises a UL29 polypeptide or antigenic fragment thereof, a UL39 polypeptide or antigenic fragment thereof, a UL49 polypeptide or antigenic fragment thereof, a UL9 polypeptide or antigenic fragment thereof, or a combination thereof.
  • a polyribonucleotide encoding a T-cell string encodes a UL29 polypeptide or antigenic fragment thereof, a UL39 polypeptide or antigenic fragment thereof, a UL49 polypeptide or antigenic fragment thereof, and a UL9 polypeptide or antigenic fragment thereof.
  • an HSV e.g., HSV-1, HSV-2, or a combination
  • a polyribonucleotide encoding a T-cell string encodes one or more of a UL30 polypeptide or antigenic fragment thereof, a UL40 polypeptide or antigenic fragment thereof, a UL5 polypeptide or antigenic fragment thereof, and a UL52 polypeptide or antigenic fragment thereof.
  • an HSV e.g., HSV-1, HSV-2, or a combination
  • a polyribonucleotide encoding a T-cell string encodes one or more of a UL29 polypeptide or antigenic fragment thereof, a UL39 polypeptide or antigenic fragment thereof, a UL49 polypeptide or antigenic fragment thereof, a UL9 polypeptide or antigenic fragment thereof, a UL30 polypeptide or antigenic fragment thereof, a UL40 polypeptide or antigenic fragment thereof, a UL5 polypeptide or antigenic fragment thereof, and a UL52 polypeptide or antigenic fragment thereof.
  • an HSV (e.g., HSV-1 and/or HSV-2) antigen for use in accordance with the present disclosure is a late protein or an antigenic fragment thereof.
  • a polyribonucleotide provided herein encodes one or more HSV (e.g., HSV-1 and/or HSV-2) antigens, wherein at least one HSV antigen comprises a late protein or an antigenic fragment thereof.
  • a polyribonucleotide provided herein encodes one or more HSV (e.g., HSV-1 and/or HSV-2) antigens, wherein each of the HSV antigens comprises a late protein or an antigenic fragment thereof.
  • an HSV (e.g., HSV-1, HSV-2, or a combination) T-cell antigen for use in accordance with the present disclosure is a late protein or an antigenic fragment thereof.
  • a polyribonucleotide provided herein encodes one or more HSV antigens, wherein at least one HSV antigen comprises a late protein or an antigenic fragment thereof.
  • a polyribonucleotide provided herein encodes one or more HSV antigens, wherein each of the HSV antigens comprises a late protein or an antigenic fragment thereof.
  • an HSV T-cell antigen is an HSV-2 T-cell antigen.
  • an HSV e.g., HSV-1, HSV-2, or a combination
  • a polyribonucleotide encoding a T-cell string encodes one or more of a UL1 polypeptide or antigenic fragment thereof, a UL19 polypeptide or antigenic fragment thereof, a UL21 polypeptide or antigenic fragment thereof, a UL27 polypeptide or antigenic fragment thereof, a UL46 polypeptide or antigenic fragment thereof, a UL47 polypeptide or antigenic fragment thereof, a UL48 polypeptide or antigenic fragment thereof, and a UL25 polypeptide or antigenic fragment thereof.
  • Exemplary polyribonucleotide sequences encoding antigenic fragments are examples of a UL1 polypeptide or antigenic fragment thereof, a UL19 polypeptide or antigenic fragment thereof, a UL21 polypeptide or antigenic fragment thereof, a UL27 polypeptide or antigenic fragment thereof, a UL46 polypeptide or antigenic fragment thereof, a UL47 polypeptide or antigenic fragment thereof, a UL48 polypeptide
  • Table 9 Exemplary polyribonucleotide sequences encoding antigenic fragments SEQ ID Antigen Optimized Nucleotide Sequence NO c c g a a a g c gc c c g a a a g c gc uccaggcgauacuggaacuccugcuccugccucuggcgagaucgccccuccuaauucuacaagaagc gccagcgagagccggcacagaugauaa 611 US7 [HSV-2 Opt3 gguccuaccgugucucucgugucugacagccugguagacgccggugcugugggcccucagggcuuc a a cc cg a ac gg g u a c c u a u c c u a u g c a u g c a u g c a
  • a first set of polyribonucleotides further comprises a polyribonucleotide encoding an HSV glycoprotein B (gB) , variant thereof, or an antigenic portion thereof, a polyribonucleotide encoding an HSV glycoprotein G (gG) or an antigenic portion thereof, a polyribonucleotide encoding an HSV glycoprotein H (gH) or an antigenic portion thereof, a polyribonucleotide encoding an HSV glycoprotein I (gI) or an antigenic portion thereof, a polyribonucleotide encoding an HSV glycoprotein L (gL) or an antigenic portion thereof, or a combination thereof.
  • gB HSV glycoprotein B
  • additional polyribonucleotides are used with the first and/or second set of polyribonucleotides.
  • an additional polyribonucleotide encodes a polypeptide comprising ICP0, ICP4, ICP22, VP16, ICP47, VHS, or US3.
  • an additional set of polyribonucleotides encodes a set of polypeptides, wherein the set of polypeptides comprises ICP0, ICP4, ICP22, VP16, ICP47, VHS, US3, or any combination thereof.
  • an additional set of polyribonucleotides encodes a set of polypeptides, wherein the set of polypeptides comprises ICP0, ICP4, or both.
  • a first set of polyribonucleotides comprises: a polyribonucleotide encoding an HSV glycoprotein B (gB), variant thereof, or an antigenic portion thereof; a polyribonucleotide encoding an HSV glycoprotein C (gC) , variant thereof, or an antigenic portion thereof; and a polyribonucleotide encoding an HSV glycoprotein D (gD) , variant thereof, or an antigenic portion thereof.
  • an additional set of polyribonucleotides encodes a set of polypeptides, wherein the set of polypeptides comprises ICP0, ICP4, or both.
  • a combination provided herein comprises a polyribonucleotide encoding an HSV glycoprotein B (gB), variant thereof, or an antigenic portion thereof; a polyribonucleotide encoding an HSV glycoprotein C (gC) , variant thereof, or an antigenic portion thereof; a polyribonucleotide encoding an HSV glycoprotein D (gD) , variant thereof, or an antigenic portion thereof; a polyribonucleotide encoding an HSV ICP0, variant thereof, or an antigenic portion thereof; and a polyribonucleotide encoding an HSV ICP4, variant thereof, or an antigenic portion thereof.
  • a combination provided herein comprises a polyribonucleotide encoding an antigenic portion HSV glycoprotein B (gB), variant thereof, or an antigenic portion thereof; a polyribonucleotide encoding an HSV glycoprotein C (gC) , variant thereof, or an antigenic portion thereof; a polyribonucleotide encoding an HSV glycoprotein D (gD) , variant thereof, or an antigenic portion thereof; a polyribonucleotide encoding an HSV ICP0, variant thereof, or an antigenic portion thereof; and a polyribonucleotide encoding an HSV ICP4, variant thereof, or an antigenic portion thereof.
  • gB HSV glycoprotein B
  • gC HSV glycoprotein C
  • gD HSV glycoprotein D
  • a combination provided herein comprises a polyribonucleotide encoding an antigenic portion HSV glycoprotein B (gB); a polyribonucleotide encoding an antigenic portion of HSV glycoprotein C (gC); a polyribonucleotide encoding an HSV glycoprotein D (gD) or an antigenic portion thereof; a polyribonucleotide encoding an antigenic portion of HSV ICP0; and a polyribonucleotide encoding an antigenic portion of HSV ICP4.
  • a combination provided herein comprises a polyribonucleotide encoding an antigenic portion HSV-2 glycoprotein B (gB); a polyribonucleotide encoding an antigenic portion of HSV-2 glycoprotein C (gC); a polyribonucleotide encoding an HSV-2 glycoprotein D (gD) or an antigenic portion thereof; a polyribonucleotide encoding an antigenic portion of HSV-2 ICP0; and a polyribonucleotide encoding an antigenic portion of HSV-2 ICP4. D.
  • polyribonucleotides encoding polypeptides (e.g., GP polypeptide) comprising (i) an HSV glycoprotein or antigenic portion thereof and (ii) a secretory signal.
  • polyribonucleotides encoding polypeptides (e.g., T-cell string polypeptides) comprising (i) one or more HSV T-cell antigens or antigenic portions thereof and (ii) secretory signal.
  • a GP polypeptide comprises a secretory signal.
  • a GP polypeptide comprises (i) an HSV gC or antigenic portion thereof and (ii) a secretory signal. In some embodiments, a GP polypeptide comprises (i) an HSV gD or antigenic portion thereof and (ii) a secretory signal. In some embodiments, a GP polypeptides comprises (i) an HSV gD or antigenic portion thereof and (ii) a secretory signal. In some embodiments, a GP polypeptide comprises (i) an HSV gB, variant thereof, or one or more antigenic portions thereof and (ii) a secretory signal. [0435] In some embodiments, a T-cell string polypeptide comprises a secretory signal.
  • a T-cell string polypeptide comprises (i) one or more HSV T-cell antigens or antigenic portions thereof and (ii) a secretory signal.
  • a secretory signal is functional in mammalian cells.
  • a utilized secretory signal is a heterologous secretory signal.
  • a secretory signal comprises or consists of a human secretory signal.
  • a secretory signal comprises or consists of an IL2 secretory signal.
  • a secretory signal comprises or consists of a viral secretory signal.
  • a viral secretory signal comprises or consists of an HSV secretory signal (e.g., an HSV-1 or HSV-2 secretory signal).
  • a secretory signal comprises or consists of an HSV-1 secretory signal.
  • a secretory signal comprises or consists of an HSV-2 secretory signal.
  • an HSV secretory signal comprises or consists of an HSV glycoprotein D (gD) secretory signal (e.g., an HSV-1 or HSV-2 gD secretory signal).
  • gD HSV glycoprotein D
  • an HSV secretory signal comprises or consists of an HSV-1 gD secretory signal.
  • an HSV-1 gD secretory signal comprises one or more additional amino acids. In some embodiments, an HSV-1 gD secretory signal comprises KY at the C terminus of the signal sequence. In some embodiments, an HSV secretory signal comprises or consists of an HSV-2 gD secretory signal. In some embodiments, an HSV-2 gD secretory signal comprises one or more additional amino acids. In some embodiments, an HSV-2 gD secretory signal comprises KY, KYA, KYAL (SEQ ID NO: 842), or KYALA (SEQ ID NO: 841) at the C terminus of the signal sequence.
  • an HSV secretory signal comprises or consists of an HSV glycoprotein C (gC) secretory signal (e.g., an HSV-1 or HSV-2 gC secretory signal). In some embodiments, an HSV secretory signal comprises or consists of an HSV-2 gC secretory signal. [0440] In some embodiments, an HSV secretory signal comprises or consists of an HSV glycoprotein E (gE) secretory signal (e.g., an HSV-1 or HSV-2 gE secretory signal). In some embodiments, an HSV secretory signal comprises or consists of an HSV-1 gE secretory signal.
  • gE HSV glycoprotein E secretory signal
  • an HSV secretory signal comprises or consists of an HSV-2 gE secretory signal. In some embodiments, an HSV-2 gE secretory signal comprises one or more additional amino acids. In some embodiments, an HSV-2 gE secretory signal comprises RTS. In some embodiments, an HSV-2 secretory signal comprises A20V, A21V, A22V substitutions. [0441] In some embodiments, an HSV secretory signal comprises or consists of an HSV glycoprotein B (gB) secretory signal (e.g., an HSV-1 or HSV-2 gB secretory signal). In some embodiments, an HSV secretory signal comprises or consists of an HSV-1 gB secretory signal.
  • gB HSV glycoprotein B
  • an HSV secretory signal comprises or consists of an HSV-1 gB secretory signal.
  • an HSV-1 gB secretory signal comprises one or more additional amino acids.
  • an HSV-1 gB secretory signal comprises AP at the C terminus of the signal sequence.
  • an HSV secretory signal comprises or consists of an HSV-2 gB secretory signal.
  • an HSV secretory signal comprises or consists of an HSV glycoprotein I (gI) secretory signal (e.g., an HSV-1 or HSV-2 gI secretory signal).
  • an HSV secretory signal comprises or consists of an HSV-1 gI secretory signal.
  • an HSV-1 gI secretory signal comprises one or more additional amino acids.
  • an HSV secretory signal comprises or consists of an HSV- 2 gI secretory signal.
  • an HSV-2 gI secretory signal comprises an additional leucine residue at the C terminus of the signal sequence.
  • a secretory signal comprises or consists of an Ebola spike glycoprotein (EboZ).
  • EboZ secretory signal comprises one or more additional amino acids.
  • an EboZ secretory signal comprises IP at the C terminus of the signal sequence.
  • a secretory signal is characterized by a length of about 15 to 30 amino acids.
  • a secretory signal is positioned at the N-terminus of a polyribonucleotide.
  • a secretory signal preferably allows transport of a polyribonucleotide with which it is associated into a defined cellular compartment, preferably a cell surface, endoplasmic reticulum (ER) or endosomal- lysosomal compartment.
  • polyribonucleotides comprising an HSV antigen do not comprise a secretory signal.
  • polyribonucleotides comprising an HSV antigen further comprise a codon initiation start site.
  • a secretory signal is one listed in Table 10, or a secretory signal having 1, 2, 3, 4, or 5 amino acid differences relative thereto.
  • a secretory signal is selected from those included in the Table 10 below and/or those encoded by the sequences in Table 11 and/or Table 12 below.
  • a polypeptide (e.g., a GP polypeptide or a T-cell string polypeptide) described herein includes a transmembrane region.
  • a polyribonucleotide described herein encodes a polypeptide (e.g., a GP polypeptide or a T-cell string polypeptide) that comprises a transmembrane region.
  • a transmembrane region is located at the N-terminus of a polypeptide (e.g., a GP polypeptide or a T-cell string polypeptide).
  • a transmembrane region is located at the C-terminus of a polypeptide (e.g., a GP polypeptide or a T-cell string polypeptide). In some embodiments, a transmembrane region is not located at the N-terminus or C-terminus of a polypeptide (e.g., a GP polypeptide or a T-cell string polypeptide). In some embodiments, a polypeptide does not include a transmembrane region. [0449] Transmembrane regions are known in the art, any of which can be utilized in a polypeptide described herein.
  • a transmembrane region comprises or is a transmembrane region of Hemagglutinin (HA) of Influenza virus, Env of HIV-1, equine infectious anaemia virus (EIAV), murine leukaemia virus (MLV), mouse mammary tumor virus, G protein of vesicular stomatitis virus (VSV-G), Rabies virus, or a seven transmembrane domain receptor.
  • HA Hemagglutinin
  • EIAV equine infectious anaemia virus
  • MMV murine leukaemia virus
  • VSV-G G protein of vesicular stomatitis virus
  • Rabies virus or a seven transmembrane domain receptor.
  • a polypeptide comprises an HSV transmembrane region.
  • an HSV transmembrane region is an HSV-1 or HSV-2 transmembrane region.
  • an HSV transmembrane region is an HSV-2 gD transmembrane region. In some embodiments, an HSV transmembrane region is an HSV-2 gC transmembrane region. In some embodiments, an HSV transmembrane region is an HSV-2 gE transmembrane region. In some embodiments, an HSV transmembrane region comprises or consists of an HSV gD transmembrane region, e.g., comprising or consisting of an amino acid sequence of GLIAGAVGGSLLAALVICGIVYWMRRHTQKAPKRIRLPHIR (SEQ ID NO: 468).
  • a utilized transmembrane region is a heterologous transmembrane region.
  • a heterologous transmembrane region comprises or consists of a human transmembrane region.
  • a human transmembrane region comprises or consists of a human decay accelerating factor glycosylphosphatidylinositol (hDAF-GPI) anchor region.
  • hDAF-GPI anchor region comprises or consists of an amino acid sequence of PNKGSGTTSGTTRLLSGHTCFTLTGLLGTLVTMGLLT (SEQ ID NO: 469).
  • an HSV transmembrane region comprises or consists of an HSV gB transmembrane region, e.g., comprising or consisting of an amino acid sequence of MSNPFGALAVGLLVLAGLAAAFFAFRYVMRL (SEQ ID NO: 470) or MSNPFGALAVGLLVLAGLVAAFFAFRYVLQL (SEQ ID NO: 471).
  • a viral transmembrane region comprises or consists of a vesicular stomatitis virus G (VSV-G) transmembrane region.
  • a VSV-G transmembrane region comprises or consists of an amino acid sequence of IASFFFIIGLIIGLFLVLRVGIYLCIKLKHTKKRQIYTDIEMN (SEQ ID NO: 245).
  • Example transmembrane regions are provided in the following Table 13: Table 13: Example transmembrane regions SEQ Transmembrane Region Sequence (Amino Acid) ID 468 HSV-1 gD GLIAGAVGGSLLAALVICGIVYWMRRHTQKAPKRIRLPHIR 469 hDAF-GPI anchor region PNKGSGTTSGTTRLLSGHTCFTLTGLLGTLVTMGLLT [0453] Provided herein are polypeptides (e.g., GP polypeptides and/or T-cell string polypeptides) that comprise a trafficking signal.
  • polypeptides e.g., GP polypeptides and/or T-cell string polypeptides
  • a trafficking signal is an MHC Class I Trafficking Signal (MITD).
  • MITD MHC Class I Trafficking Signal
  • at least one of the one or more GP polypeptides comprise a MITD.
  • at least one of the one or more GP polypeptides comprise (i) an HSV gC or antigenic portion thereof and (ii) a MITD.
  • at least one of the one or more GP polypeptides comprise (i) an HSV gD or antigenic portion thereof and (ii) a MITD.
  • at least one of the one or more GP polypeptides comprise (i) an HSV gD or antigenic portion thereof and (ii) a MITD.
  • At least one of the one or more GP polypeptides comprise (i) an HSV gB, variant thereof, or one or more antigenic portions thereof and (ii) a MITD.
  • at least one of the one or more T-cell string polypeptides comprises a MITD.
  • a MITD comprises or consists of an amino acid sequence that is at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to IVGIVAGLAVLAVVVIGAVVATVMCRRKSSGGKGGSYSQAASSDSAQGSDVSLTA (SEQ ID NO: 473).
  • a MITD comprises or consists of an amino acid sequence of IVGIVAGLAVLAVVVIGAVVATVMCRRKSSGGKGGSYSQAASSDSAQGSDVSLTA (SEQ ID NO: 473).
  • G. Multimerization Regions Provided herein are polypeptides (e.g., GP polypeptides and/or T-cell string polypeptides) that comprise one or more multimerization regions.
  • a multimerization region is a heterologous multimerization region.
  • a heterologous multimerization region comprises a dimerization, trimerization or tetramerization region.
  • At least one of the one or more GP polypeptides comprise one or more multimerization regions. In some embodiments, at least one of the one or more GP polypeptides comprise (i) an HSV gC or antigenic portion thereof and (ii) one or more multimerization regions. In some embodiments, at least one of the one or more GP polypeptides comprise (i) an HSV gD or antigenic portion thereof and (ii) one or more multimerization regions. In some embodiments, at least one of the one or more GP polypeptides comprise (i) an HSV gD or antigenic portion thereof and (ii) one or more multimerization regions.
  • At least one of the one or more GP polypeptides comprise (i) an HSV gB, variant thereof, or one or more antigenic portions thereof and (ii) one or more multimerization regions.
  • at least one of the one or more T-cell string polypeptides comprises one or more multimerization regions.
  • a multimerization region is one described in WO2017/081082, which is incorporated herein by reference in its entirety (e.g., SEQ ID NOs: 1116-1167, or fragments or variants thereof).
  • Example trimerization and tetramerization regions include, but are not limited to, engineered leucine zippers, fibritin foldon domain from enterobacteria phage T4, GCN4pll, GCN4-pll, and p53.
  • a multimerization region comprising or consisting of the amino acid sequence GYIPEAPRDGQAYVRKDGEWVLLSTFL (SEQ ID NO: 474).
  • a GP polypeptide comprising an HSV-2 gB or antigenic portion thereof, as described herein, is able to form a trimeric complex.
  • a GP polypeptide comprising an HSV-2 gB or antigenic portion thereof, as described herein may further comprise a multimerization region allowing formation of a multimeric complex, such as for example a trimeric complex of an HSV-2 gB described herein.
  • a multimerization region allowing formation of a multimeric complex comprises a trimerization region, for example, a trimerization region described herein.
  • a GP polypeptide comprising an HSV-2 gB or antigenic portion thereof, as described herein further includes a T4-fibritin-derived “foldon” trimerization region, for example, to increase its immunogenicity.
  • a polypeptide (e.g., a GP polypeptide and/or T-cell string polypeptide) includes one or more linkers. In some embodiments, at least one of the one or more T-cell string polypeptides comprises a secretory signal. [0464] In some embodiments, a linker is or comprises 2, 3, 4, 5, 6, 7, 8, 9, 10 or more amino acids. In some embodiments, a linker is or comprises no more than about 30, 25, 20, 15, 10 or fewer amino acids. A linker can include any amino acid sequence and is not limited to any particular amino acids. In some embodiments, a linker comprises one or more glycine (G) amino acids.
  • G glycine
  • a linker comprises one or more serine (S) amino acids.
  • a linker comprises a glycine-serine linker.
  • a “glycine-serine linker” as used herein refers to a linker that comprises predominantly (e.g., 80% or more) glycine and serine amino acids.
  • a linker includes amino acids selected based on a cleavage predictor to generate highly-cleavable linkers.
  • a linker is or comprises SGGGGSGGGGS (SEQ ID NO: 475).
  • a linker is or comprises GSPGSGSGS (SEQ ID NO: 476).
  • a linker is or comprises GGSGGGGSGG (SEQ ID NO: 477). In some embodiments, a linker is one presented in Table 14. In some embodiments, a linker is or comprises a sequence as set forth in WO2017/081082, which is incorporated herein by reference in its entirety (see SEQ ID NOs: 1509-1565, or a fragment or variant thereof). [0466] In some embodiments, a GP polypeptide described herein comprises a linker between a C-terminal region or fragment thereof and a transmembrane region. In some embodiments, a GP polypeptide described herein comprises a linker after a minor repeat sequence.
  • a T-cell string polypeptide comprises one or more linkers. In some embodiments, a T-cell string polypeptide comprises a one or more linkers separating one or more HSV T-cell antigens or antigenic portions thereof.
  • Example linkers are provided in the following Table 14: Table 14: Example linkers SEQ ID NO: Sequence (Amino Acid) 475 SGGGGSGGGGS [0468]
  • a polyribonucleotide encodes a GP polypeptide, wherein the GP polypeptide comprises an HSV glycoprotein or antigenic portion thereof and a secretory signal.
  • a polyribonucleotide encodes a GP polypeptide, wherein the GP polypeptide comprises an antigenic portion of an HSV glycoprotein and a secretory signal.
  • Example polyribonucleotide constructs encoding a gC, gD, or gE as described herein are provided in Table 15 below.
  • Example combinations of an antigenic portion of HSV-2 gC and a secretory signal are provided in Table 16 below, along with example corresponding amino acid sequences.
  • a GP polypeptide as described herein (or encoded by a polyribonucleotide as described herein) comprises an antigenic portion of HSV-2 gD and a secretory signal.
  • Example combinations of an antigenic portion of HSV-2 gD and a secretory signal are provided in Table 16 below, along with example corresponding amino acid sequences.
  • a GP polypeptide as described herein (or encoded by a polyribonucleotide as described herein) comprises an antigenic portion of HSV-2 gE and a secretory signal.
  • Example combinations of an antigenic portion of HSV-2 gE and a secretory signal are provided in Table 16 below, along with example corresponding amino acid sequences.
  • Example GP polypeptides comprising a secretory signal and an antigenic portions of an HSV-2 glycoprotein SEQ ID Secretory Antigenic NO: signal fragment Sequence (Amino acids) T IR P R W R SS PR RY Q Y G T A IP S DP PP Q T PP G YP P S LE A G PE G A PL P Q T YYPGNRAEFVWFEDGRRVFDPAQIHTQTQENPDGFSTVSTVTSAAVGGQGPP RTFTCQLTWHRDSVSFSRRNASGTASVLPRPTITMEFTGDHAVCTAGCVPEG VTFAWFLGDDSSPAEKVAVASQTSCGRPGTATIRSTLPVSYEQTEYICRLAGYP S LA G G YY T TF GI S C V E T G G C P V D A L L E P A G G YY T TF GI SN N N L TC G C RL 69 IL2 HSV-2 gD MRMQLLLLIALSLAL
  • a pol ribonucleotide encoding a T-cell string can include, in ).
  • a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 595.
  • a polyribonucleotide encodes a T-cell string that comprises amino acids 26- 693 of amino acid sequence SEQ ID NO: 595.
  • a polyribonucleotide encoding a T-cell string can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an UL29 polypeptide or antigenic ).
  • a polyribonucleotide encodes a polypeptide having an amino acid 26- can YG RAY FL FAFLSAADDLVTENLGGLSGLFEQKDILHYYVEQECIEVVHSRVYNIIQLVLFHNNDQARRAYVARTINHPAIRVKVDWLEARVRECD SIPEKFILMILIEGVFFAASFAAIAYLRTNNLLRGGSGGGGSGGHEFGNLMKVLEYGLPITEEHMQFVDRFVVPESYITNPANLPGWTR LFSSHKEVSAYMAKLHAYLKVTREGEFVVFTLPVLTFVSVKEFDEYRRLGGSGGGGSGGELFGEVFESAPFSTYVDNVIFRGCELLTGS PRGGLMSVALQTDNYTLMGYTYTRVFAFAEELRRRHATAGVAEFLEESPLPYIVLRDQHGFMSVVNTNIGGSGGGGSGGSVAAPVE VTALYATDGCVITSSLALLTNCLLGAEPLYIFSYDAYRSD
  • a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 597. [0479] In some embodiments, a polyribonucleotide encodes a T-cell string that comprises amino acids 26- 792 of amino acid sequence SEQ ID NO: 597.
  • a polyribonucleotide encoding a T-cell string polypeptide can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an UL1 polypeptide or antigenic fragment thereof, a linker, an UL19 polypeptide or antigenic fragment thereof, a linker, an UL21 polypeptide or antigenic fragment thereof, a linker, a UL27 polypeptide or fragment thereof, a linker, a UL27 polypeptide or fragment thereof, a linker, a UL46 polypeptide or fragment thereof, a linker, a UL47 polypeptide or fragment thereof, a linker, a UL25 polypeptide or fragment thereof, a linker, a UL48 polypeptide or fragment thereof, a linker, and a MITD (see FIG.18D).
  • such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGRTPADDVSWRYEAPSVIDYARIDGIFLRYHCPGLDTFLWDRHAQRAYLVNPFLFAAGFLEDL SHSVFPADTQETTGGSGGGGSGGDGRLLHNTQARAADAADDRPHRPADWTVHHKIYYYVLVPAFSRGRCCTAGVRFDRVYATLQ NMVVPEIAPGEECPSDPVTDPAHPLHPANLVANTVKRMFHNGGSGGGGSGGSPTQKLAVYYYLIHRERRMSPFPALVRLVGRYIQR HGLYVPAPDEPTLADAMNGLGGSGGGGSGGNYTEGIAVVFKENIAPYKFKATMYYKDVTVSQVWFGHRYSQFMGIFEDRAPVPFEE VGGSGGGGSGGSVYPYDEFVLATGDFVYMSPFYGYREGSHG
  • a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 598.
  • a polyribonucleotide encodes a T-cell string that comprises amino acids 26- 858 of amino acid sequence SEQ ID NO: 598.
  • a polyribonucleotide encoding a T-cell string polypeptide can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an UL54 polypeptide or antigenic fragment thereof, a linker, an RS1 polypeptide or antigenic fragment thereof, a linker, an RL2 polypeptide or antigenic fragment thereof, a linker, a RL2 polypeptide or fragment thereof, a linker, and a MITD.
  • such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGETLVAHGPSLYRTFAANPRAASTAKAMRDCVLRQENLIEALASADETLAWCKMCIHHNLPLR PQDPIIGTAAAVLENLATRLRPFLQCYLKARGLCGLDDLCSRRRLSDIKDIASFVLVILARLANRVERGVSEIDYTTVGVGAGETMHFYI PGACMAGLIEILDTHRQECSSRVCELTASHTIAPLYVHGKYFYCNSLFGGSGGGGSGGRAAAWMRQVPDPEDVRVVILYSPLPGEDL AAGRAGGGPPPEWSAERGGLSCLLAALGNRLCGPATAAWAGNWTGAPDVSALGAQGVLLLSTRDLAFAGAVEFLGLLAGACDRRLI VVNAVRAADWPADGPVVSRQHAYLACEVLPAVQCAVRWPAARDLRRTVLASGRVFGPGV
  • a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 599. [0483] In some embodiments, a polyribonucleotide encodes a T-cell string that comprises amino acids 26- 692 of amino acid sequence SEQ ID NO: 599.
  • a polyribonucleotide encoding a T-cell string polypeptide can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an UL9 polypeptide or antigenic fragment thereof, a linker, an UL49 polypeptide or antigenic fragment thereof, a linker, an UL39 polypeptide or antigenic fragment thereof, a linker, a UL29 polypeptide or fragment thereof, a linker, and a MITD.
  • such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGLLNNYDVLVLDEVMSTLGQLYSPTMQQLGRVDALMLRLLRTCPRIIAMDATANAQLVDFLCS LRGEKNVHVVIGEYAMPGFSARRCLFLPRLGPEVLQAALRRRGPAGGAPPPDAPPDATFFGELEARLAGGDNVCIFSSTVSFAEVVAR FCRQFTDRVLLLHSLTPPGDVTTWGRYRVVIYTTVVTVGLSFDPPHFDSMFAYVKPMNYGPDMVSVYQSLGRVRTLRKGELLIYMD GSGARSEPVGGSGGGGSGGKMTRGAPKASATPATDPARGRRPAQADSAVLLDAPAPTASGRTKTPAQGLAKKLHFSTAPPSPTAPW TPRVAGFNKRVFCAAVGGGSGGGGSGGRTFGSAPRLTEDDFGLLNYALAEMRRLCLD
  • a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 600. [0485] In some embodiments, a polyribonucleotide encodes a T-cell string that comprises amino acids 26- 722 of amino acid sequence SEQ ID NO: 600.
  • a polyribonucleotide encoding a T-cell string polypeptide can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an UL52 polypeptide or antigenic fragment thereof, a linker, an UL5 polypeptide or antigenic fragment thereof, a linker, an UL5 polypeptide or antigenic fragment thereof, a linker, a UL40 polypeptide or fragment thereof, a linker, a UL30 polypeptide or fragment thereof, a linker, a UL30 polypeptide or fragment thereof, a linker, and a MITD.
  • such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGSVAAPVEVTALYATDGCVITSSLALLTNCLLGAEPLYIFSYDAYRSDAPNGPTGAPTEQERFEG SRALYRDAGGLNGDSFRVTFCLLGTEVGVTHHPKGRTRPMFVCRFERADDVAVLQDALGRGTPLLPAHVTATLDLEATFALHANIIM ALTVAIVHNAPARIGSGSTAPLYEPGESMRSVVGGSGGGGSGGHEFGNLMKVLEYGLPITEEHMQFVDRFVVPESYITNPANLPGWT RLFSSHKEVSAYMAKLHAYLKVTREGEFVVFTLPVLTFVSVKEFDEYRRLGGSGGGGSGGELFGEVFESAPFSTYVDNVIFRGCELLTG SPRGGLMSVALQTDNYTLMGYTYTRVFAEELRRRHATAGVAEFLEESPLPY
  • a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 601. [0487] In some embodiments, a polyribonucleotide encodes a T-cell string that comprises amino acids 26- 792 of amino acid sequence SEQ ID NO: 601.
  • a polyribonucleotide encoding a T-cell string polypeptide can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an UL48 polypeptide or antigenic fragment thereof, a linker, an UL25 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, a UL46 polypeptide or fragment thereof, a linker, a UL27 polypeptide or fragment thereof, a linker, a UL27 polypeptide or fragment thereof, a linker, a UL21 polypeptide or fragment thereof, a linker, a UL19 polypeptide or fragment thereof, a linker, a UL1 polypeptide or fragment thereof, a linker, and a MITD.
  • such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGALFNRLLDDLGFSAGPALCTMLDTWNEDLFSGFPTNADMYRECKFLSTLPSDVIDWGDAHV PERSPIDIRAHGDVAFPTLPATRDELPSYYEAMAQFFRGELRAGGSGGGGSGGFLWEDQTLLRATANTITALAVLRRLLANGNVYAD RLDNRLQLGMLIPGAVPAEAIARGASGLDSGAIKSGDNNLEALCVNYVLPLYQADPTVELTQLFPGLAALCLGGSGGGGSGGGPDAA VFRSSLGSLLYWPGVRALLGRDCRVAARYAGRMTYIATGALLARFNPGAVKCVLPREAAFAGRVLGGSGGGGSGGGLASDPHYDYIR HYASAAKQALGEVELSGGQLSRAILAQYWKYLQTVVPSGLDIPDDPAGDCDPSLHVLLRPTLLPKLL
  • a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 602. [0489] In some embodiments, a polyribonucleotide encodes a T-cell string that comprises amino acids 26- 858 of amino acid sequence SEQ ID NO: 602.
  • a polyribonucleotide encoding a T-cell string polypeptide can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-2 gD secretory signal, an RL2 polypeptide or antigenic fragment thereof, a linker, an RL2 polypeptide or antigenic fragment thereof, a linker, an RS1 polypeptide or antigenic fragment thereof, a linker, a UL54 polypeptide or fragment thereof, a linker, and a MITD.
  • such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGRLTSGVGTAALLVVAVGLRVVCACTDEIAPPLRCQSFPCLHPFCIPCMKTWIPLRNTCPLCNTPVAYLIVGVTASGSFSTIPIVNDP RTRVEAEAAVRAGTAVDFIWTGNPRTAPRSLSGGSGGGGSGGLPIAGVSSVVALAPYVNKTVTGDCLPVLDMETGHIGAYVVLVDQ TGNVADLLRAAAPAWSRRTLLPEHARNCVRPPDYPTPPASEWNSLWMTPVGNMLFDQGTLVGGGSGGGGSGGRAAAWMRQVPD PEDVRVVILYSPLPGEDLAAGRAGGGPPPEWSAERGGLSCLLAALGNRLCGPATAAWAGNWTGAPDVSALGAQGVLLLSTRDLAFA GAVEFLGLLAGACDRRLIVVNAVRAADWPADGPVVSRQHAYLACEVLPAVQCAVRWPAARDLRR
  • a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 603. [0491] In some embodiments, a polyribonucleotide encodes a T-cell string that comprises amino acids 26- 693 of amino acid sequence SEQ ID NO: 603.
  • a polyribonucleotide encoding a T-cell string polypeptide can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-2 gD secretory signal, an UL54 polypeptide or antigenic fragment thereof, a linker, an RS1 polypeptide or antigenic fragment thereof, a linker, an RL2 polypeptide or antigenic fragment thereof, a linker, a RL2 polypeptide or fragment thereof, a linker, and a MITD.
  • such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGRLTSGVGTAALLVVAVGLRVVCAETLVAHGPSLYRTFAANPRAASTAKAMRDCVLRQENLIEALASADETLAWCKMCIHHNLPLR PQDPIIGTAAAVLENLATRLRPFLQCYLKARGLCGLDDLCSRRRLSDIKDIASFVLVILARLANRVERGVSEIDYTTVGVGAGETMHFYI PGACMAGLIEILDTHRQECSSRVCELTASHTIAPLYVHGKYFYCNSLFGGSGGGGSGGRAAAWMRQVPDPEDVRVVILYSPLPGEDL AAGRAGGGPPPEWSAERGGLSCLLAALGNRLCGPATAAWAGNWTGAPDVSALGAQGVLLLSTRDLAFAGAVEFLGLLAGACDRRLI VVNAVRAADWPADGPVVSRQHAYLACEVLPAVQCAVRWPAARDLRRTVLASGRVFGPGV
  • a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 604. [0493] In some embodiments, a polyribonucleotide encodes a T-cell string that comprises amino acids 26- 692 of amino acid sequence SEQ ID NO: 604.
  • a polyribonucleotide encoding a T-cell string polypeptide can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an US1 polypeptide or antigenic fragment thereof, a linker, an US1 polypeptide or antigenic fragment thereof, a linker, an US8 polypeptide or antigenic fragment thereof, a linker, a US8 polypeptide or fragment thereof, a linker, a US8 polypeptide or fragment thereof, a linker, a US12 polypeptide or fragment thereof, a linker, a UL50 polypeptide or fragment thereof, a linker, a UL26 polypeptide or fragment thereof, a linker, a UL26 polypeptide or fragment thereof, a linker, a US10 polypeptide or fragment thereof, a linker, and a MITD.
  • such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGDDASDGWLVDTPPRKSKRPRINLRLTSSPDRRAGVVFPEVGGSGGGGSGGPASLPGIAHAH RRSARQAQMRSGAAWTLDLHYIRQCVNQLGGSGGGGSGGILSPTAPSVYPHSEGRKSRRPLTTFGSGSPGRRHSQASYPSVLWGG SGGGGSGGGLAWLASTVNLEFQHASPQHAGLYLCVVYVDDHIHAWGHMTISTAAQYRNAVVEQHLPQRQPEPVEPTRPHVRAPHP APSARGPLRLGGSGGGGSGGKLLWAAEPLDACGPLRPSWVALWPPRRVLETVVDAACMRAPEPLAIAYSPPFPAGDEGLYSELAWR DRVAVVNESLVIYGALETDSGLYTLSVVGLSDEARQVASVVLVVEPAPGGSGG
  • a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 605. [0495] In some embodiments, a polyribonucleotide encodes a T-cell string that comprises amino acids 26- 905 of amino acid sequence SEQ ID NO: 605.
  • a polyribonucleotide encoding a T-cell string polypeptide can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an UL26 polypeptide or antigenic fragment thereof, a linker, an UL26 polypeptide or antigenic fragment thereof, a linker, an US10 polypeptide or antigenic fragment thereof, a linker, a UL50 polypeptide or fragment thereof, a linker, a US12 polypeptide or fragment thereof, a linker, a US8 polypeptide or fragment thereof, a linker, a US8 polypeptide or fragment thereof, a linker, a US8 polypeptide or fragment thereof, a linker, a US8 polypeptide or fragment thereof, a linker, a US1 polypeptide or fragment thereof, a linker, a US1 polypeptide or fragment thereof, a linker, and a MITD.
  • such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGHYPPPPAHPYPGMLFAGPSPLEAQIAALVGAIAADRQAGGLPAAAGDHGIRGSAKRRRHEVE QPEYDCGGGSGGGGSGGAPLPDRAVPIYVAGFLALYDSGDPGELALDPDTVRAALPPENPLPINVDHRARCEVGRVLAVVNDPRGPF FVGLIACVQLERVLETAASAAIFERRGPALSREERLLYLITNYLPSVSLSTKRRGDEVPPDRTLFAHVALCAIGRRLGTIVTYDTSLDAA GGSGGGGSGGSSPRQRTYVLPRVGIHNAPASDTRAPKRANSRHRADRPPESPGSELYPLNAQALAHLQMLPADHRAFFRTVIEVSRL CALNTHDPPPPLAGARVGQEAQLVHTQWLRANRESSPLWPWRTAAMNFIAAAAPCVQ
  • a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 606. [0497] In some embodiments, a polyribonucleotide encodes a T-cell string that comprises amino acids 26- 905 of amino acid sequence SEQ ID NO: 606.
  • a polyribonucleotide encoding a T-cell string can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an RL2 polypeptide or antigenic fragment thereof, a linker, an RL2 polypeptide or antigenic fragment thereof, a linker, an RS1 polypeptide or antigenic fragment thereof, a linker, a UL54 polypeptide or fragment thereof, a linker, and an HSV-1 transmembrane region.
  • such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGCTDEIAPPLRCQSFPCLHPFCIPCMKTWIPLRNTCPLCNTPVAYLIVGVTASGSFSTIPIVNDP RTRVEAEAAVRAGTAVDFIWTGNPRTAPRSLSGGSGGGGSGGLPIAGVSSVVALAPYVNKTVTGDCLPVLDMETGHIGAYVVLVDQ TGNVADLLRAAAPAWSRRTLLPEHARNCVRPPDYPTPPASEWNSLWMTPVGNMLFDQGTLVGGGSGGGGSGGRAAAWMRQVPD PEDVRVVILYSPLPGEDLAAGRAGGGPPPEWSAERGGLSCLLAALGNRLCGPATAAWAGNWTGAPDVSALGAQGVLLLSTRDLAFA GAVEFLGLLAGACDRRLIVVNAVRAADWPADGPVVSRQHAYLACEVLPAVQCAVRWPAARDL
  • a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 607. [0499] In some embodiments, a polyribonucleotide encodes a T-cell string that comprises amino acids 26- 693 of amino acid sequence SEQ ID NO: 607.
  • a polyribonucleotide encoding a T-cell string polypeptide can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an RL2 polypeptide or antigenic fragment thereof, a linker, an RL2 polypeptide or antigenic fragment thereof, a linker, an RS1 polypeptide or antigenic fragment thereof, a linker, a UL54 polypeptide or fragment thereof, a linker, and a VSV-G transmembrane region.
  • such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGCTDEIAPPLRCQSFPCLHPFCIPCMKTWIPLRNTCPLCNTPVAYLIVGVTASGSFSTIPIVNDP RTRVEAEAAVRAGTAVDFIWTGNPRTAPRSLSGGSGGGGSGGLPIAGVSSVVALAPYVNKTVTGDCLPVLDMETGHIGAYVVLVDQ TGNVADLLRAAAPAWSRRTLLPEHARNCVRPPDYPTPPASEWNSLWMTPVGNMLFDQGTLVGGGSGGGGSGGRAAAWMRQVPD PEDVRVVILYSPLPGEDLAAGRAGGGPPPEWSAERGGLSCLLAALGNRLCGPATAAWAGNWTGAPDVSALGAQGVLLLSTRDLAFA GAVEFLGLLAGACDRRLIVVNAVRAADWPADGPVVSRQHAYLACEVLPAVQCAVRWPAARDL
  • a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 608. [0501] In some embodiments, a polyribonucleotide encodes a T-cell string that comprises amino acids 26- 693 of amino acid sequence SEQ ID NO: 608.
  • such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGCTDEIAPPLRCQSFPCLHPFCIPCMKTWIPLRNTCPLCNTPVAYLIVGVTASGSFSTIPIVNDP RTRVEAEAAVRAGTAVDFIWTGNPRTAPRSLSGGSGGGGSGGETLVAHGPSLYRTFAANPRAASTAKAMRDCVLRQENLIEALASA DETLAWCKMCIHHNLPLRPQDPIIGTAAAVLENLATRLRPFLQCYLKARGLCGLDDLCSRRRLSDIKDIASFVLVILARLANRVERGVS EIDYTTVGVGAGETMHFYIPGACMAGLIEILDTHRQECSSRVCELTASHTIAPLYVHGKYFYCNSLFGGSGGGGSGGGPDAAVFRSSL GSLLYWPGVRALLGRDCRVAARYAGRMTYIATGALLARFNPGAVKCVLPREAAFAGR
  • a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 633. [0503] In some embodiments, a polyribonucleotide encodes a polypeptide having amino acids 26-651 of amino acid sequence SEQ ID NO: 633.
  • a polyribonucleotide encoding a T-cell string polypeptide can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an UL21 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, an RL2 polypeptide or antigenic fragment thereof, a linker, and a MITD.
  • such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGSPTQKLAVYYYLIHRERRMSPFPALVRLVGRYIQRHGLYVPAPDEPTLADAMNGLGGSGGGG SGGGLASDPHYDYIRHYASAAKQALGEVELSGGQLSRAILAQYWKYLQTVVPSGLDIPDDPAGDCDPSLHVLLRPTLLPKLLVRAPFK SGAAAAKYAAAVAGLRDAAHRLQQYMFFMRPADPSRPSTDTALRLSELLAYVSVLYHWASWMLWTADKYVGGSGGGGSGGGPDA AVFRSSLGSLLYWPGVRALLGRDCRVAARYAGRMTYIATGALLARFNPGAVKCVLPREAAFAGRVLGGSGGGGSGGETLVAHGPSLY RTFAANPRAASTAKAMRDCVLRQENLIEALASADETLAWCKMCIHHNLPLRPQD
  • a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 634. [0505] In some embodiments, a polyribonucleotide encodes a polypeptide having amino acids 26-651 of amino acid sequence SEQ ID NO: 634.
  • a polyribonucleotide encoding a T-cell string polypeptide can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an UL29 polypeptide or antigenic fragment thereof, a linker, an UL39 polypeptide or antigenic fragment thereof, a linker, an UL9 polypeptide or antigenic fragment thereof, a linker, an UL5.1 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL30.1 polypeptide or antigenic fragment thereof, a linker, and a MITD.
  • such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGREDIETIAFIKRFSLDYGAINFINLAPNNVSELAMYYMANQILRYCDHSTYFINTLTAVIAGSR RPPSVQAAAAWAPQGGAGLEAGARALMDSLDAHPGAWTSMFASCNLLRPVMAARPMVVLGLSISKYYGMAGNDRVFQAGNWASL LGGKNACPLLIFDRTRKFVLGGSGGGGSGGRTFGSAPRLTEDDFGLLNYALAEMRRLCLDLPPVPPNAYTPYHLREYATRLVNGFKPL VRRSARLYRILGVLVHLRIRTREASFEEWMRSKEVDLDFGLTERLREHEAQLMILAQALNPYDCLIHSTPNTLVERGLQSALKYEEFYL KRFGGHYMESVFQMYTRIAGFLAGGSGGGGSGGLLNNYDVLVLDEVMSTLGQLYSPTMQQQ
  • a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 635. [0507] In some embodiments, a polyribonucleotide encodes a polypeptide having amino acids 26-1156 of amino acid sequence SEQ ID NO: 635.
  • a polyribonucleotide encoding a T-cell string polypeptide can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an UL30.1 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL5.1 polypeptide or antigenic fragment thereof, a linker, an UL9 polypeptide or antigenic fragment thereof, a linker, an UL39 polypeptide or antigenic fragment thereof, a linker, an UL29 polypeptide or antigenic fragment thereof, a linker, and a MITD.
  • such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGTSQCPDINHLRSLSILNRWLETELVFVGDEEDVSKLSEGELGFYRFLFAFLSAADDLVTENLG GLSGLFEQKDILHYYVEQECIEVVHSRVYNIIQLVLFHNNDQARRAYVARTINHPAIRVKVDWLEARVRECDSIPEKFILMILIEGVFFA ASFAAIAYLRTNNLLRGGSGGGGSGGISCLLYDLSTTALEHILLFSLGSCDLPESHLSDLASRGLPAPVVLEFDSEFEMLLAFMTFVKQY GPEFVTGYNIINFDWPFVLTKLTEIYKVPLDGYGRMNGRGVFRVWDIGQSHFGGSGGGGSGGELFGEVFESAPFSTYVDNVIFRGCE LLTGSPRGGLMSVALQTDNYTLMGYTYTRVFAEE
  • a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 636. [0509] In some embodiments, a polyribonucleotide encodes a polypeptide having amino acids 26-1156 of amino acid sequence SEQ ID NO: 636.
  • a polyribonucleotide encoding a T-cell string polypeptide can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an RL2.1 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, an UL9 polypeptide or antigenic fragment thereof, a linker, a UL39 polypeptide or antigenic fragment thereof, a linker, a UL5.1 polypeptide or antigenic fragment thereof, a linker, and a MITD.
  • such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGCTDEIAPPLRCQSFPCLHPFCIPCMKTWIPLRNTCPLCNTPVAYLIVGVTASGSFSTIPIVNDP RTRVEAEAAVRAGTAVDFIWTGNPRTAPRSLSGGSGGGGSGGETLVAHGPSLYRTFAANPRAASTAKAMRDCVLRQENLIEALASA DETLAWCKMCIHHNLPLRPQDPIIGTAAAVLENLATRLRPFLQCYLKARGLCGLDDLCSRRRLSDIKDIASFVLVILARLANRVERGVS EIDYTTVGVGAGETMHFYIPGACMAGLIEILDTHRQECSSRVCELTASHTIAPLYVHGKYFYCNSLFGGSGGGGSGGLLNNYDVLVLD EVMSTLGQLYSPTMQQLGRVDALMLRLLRTCPRIIAMDATANAQLVDFLC
  • a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 637. [0511] In some embodiments, a polyribonucleotide encodes a polypeptide having amino acids 26-880 of amino acid sequence SEQ ID NO: 637.
  • a polyribonucleotide encoding a T-cell string polypeptide can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an UL5.1 polypeptide or antigenic fragment thereof, a linker, an UL39 polypeptide or antigenic fragment thereof, a linker, an UL9 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, an RL2.1 polypeptide or antigenic fragment thereof, a linker, and a MITD.
  • such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGHEFGNLMKVLEYGLPITEEHMQFVDRFVVPESYITNPANLPGWTRLFSSHKEVSAYMAKLHA YLKVTREGEFVVFTLPVLTFVSVKEFDEYRRLGGSGGGGSGGRTFGSAPRLTEDDFGLLNYALAEMRRLCLDLPPVPPNAYTPYHLRE YATRLVNGFKPLVRRSARLYRILGVLVHLRIRTREASFEEWMRSKEVDLDFGLTERLREHEAQLMILAQALNPYDCLIHSTPNTLVER GLQSALKYEEFYLKRFGGHYMESVFQMYTRIAGFLAGGSGGGGSGGLLNNYDVLVLDEVMSTLGQLYSPTMQQLGRVDALMLRLLR TCPRIIAMDATANAQLVDFLCSLRGEKNVHVVIGEYAMP
  • a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 638. [0513] In some embodiments, a polyribonucleotide encodes a polypeptide having amino acids 26-880 of amino acid sequence SEQ ID NO: 638.
  • a polyribonucleotide encoding a T-cell string polypeptide can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL21 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL30.1 polypeptide or antigenic fragment thereof, a linker, an UL29 polypeptide or antigenic fragment thereof, a linker, and a MITD.
  • such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGGPDAAVFRSSLGSLLYWPGVRALLGRDCRVAARYAGRMTYIATGALLARFNPGAVKCVLPRE AAFAGRVLGGSGGGGSGGGLASDPHYDYIRHYASAAKQALGEVELSGGQLSRAILAQYWKYLQTVVPSGLDIPDDPAGDCDPSLHV LLRPTLLPKLLVRAPFKSGAAAAKYAAAVAGLRDAAHRLQQYMFFMRPADPSRPSTDTALRLSELLAYVSVLYHWASWMLWTADKY VGGSGGGGSGGSPTQKLAVYYYLIHRERRMSPFPALVRLVGRYIQRHGLYVPAPDEPTLADAMNGLGGSGGGGSGGELFGEVFESA PFSTYVDNVIFRGCELLTGSPRGGLMSVALQTDNYTLMGYTYTRVFAEELRRRH
  • a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 639. [0515] In some embodiments, a polyribonucleotide encodes a polypeptide having amino acids 26-927 of amino acid sequence SEQ ID NO: 639.
  • a polyribonucleotide encoding a T-cell string polypeptide can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an UL29 polypeptide or antigenic fragment thereof, a linker, an UL30.1 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL21 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, and a MITD.
  • such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGREDIETIAFIKRFSLDYGAINFINLAPNNVSELAMYYMANQILRYCDHSTYFINTLTAVIAGSR RPPSVQAAAAWAPQGGAGLEAGARALMDSLDAHPGAWTSMFASCNLLRPVMAARPMVVLGLSISKYYGMAGNDRVFQAGNWASL LGGKNACPLLIFDRTRKFVLGGSGGGGSGGISCLLYDLSTTALEHILLFSLGSCDLPESHLSDLASRGLPAPVVLEFDSEFEMLLAFMTF VKQYGPEFVTGYNIINFDWPFVLTKLTEIYKVPLDGYGRMNGRGVFRVWDIGQSHFGGSGGGGSGGTSQCPDINHLRSLSILNRWL ETELVFVGDEEDVSKLSEGELGFYRFLFAFLSAADDLVTENLGGLSGLFEQKD
  • a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 640. [0517] In some embodiments, a polyribonucleotide encodes a polypeptide having amino acids 26-927 of amino acid sequence SEQ ID NO: 640.
  • a polyribonucleotide encoding a T-cell string polypeptide can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, a RL2.1 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, and a MITD.
  • such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGCTDEIAPPLRCQSFPCLHPFCIPCMKTWIPLRNTCPLCNTPVAYLIVGVTASGSFSTIPIVNDP RTRVEAEAAVRAGTAVDFIWTGNPRTAPRSLSGGSGGGGSGGETLVAHGPSLYRTFAANPRAASTAKAMRDCVLRQENLIEALASA DETLAWCKMCIHHNLPLRPQDPIIGTAAAVLENLATRLRPFLQCYLKARGLCGLDDLCSRRRLSDIKDIASFVLVILARLANRVERGVS EIDYTTVGVGAGETMHFYIPGACMAGLIEILDTHRQECSSRVCELTASHTIAPLYVHGKYFYCNSLFGGSGGGGSGGELFGEVFESAP FSTYVDNVIFRGCELLTGSPRGGLMSVALQTDNYTLMGYTYTRVFAEELRRR
  • a polyribonucleotide encoding a T-cell string polypeptide can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, a RL2.1 polypeptide or antigenic fragment thereof, a linker, and a MITD.
  • such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGGLASDPHYDYIRHYASAAKQALGEVELSGGQLSRAILAQYWKYLQTVVPSGLDIPDDPAGDC DPSLHVLLRPTLLPKLLVRAPFKSGAAAAKYAAAVAGLRDAAHRLQQYMFFMRPADPSRPSTDTALRLSELLAYVSVLYHWASWMLW TADKYVGGSGGGGSGGGPDAAVFRSSLGSLLYWPGVRALLGRDCRVAARYAGRMTYIATGALLARFNPGAVKCVLPREAAFAGRVL GGSGGGGSGGTSQCPDINHLRSLSILNRWLETELVFVGDEEDVSKLSEGELGFYRFLFAFLSAADDLVTENLGGLSGLFEQKDILHYY VEQKDILHYY VEQKDILHYY VEQKDILHYY VEQKDILHYY VEQKDILHYY
  • a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 642. [0521] In some embodiments, a polyribonucleotide encodes a polypeptide having amino acids 26-874 of amino acid sequence SEQ ID NO: 642.
  • a polyribonucleotide encoding a T-cell string polypeptide can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, a RL2.1 polypeptide or antigenic fragment thereof, a linker, a RS1 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, and a MITD.
  • such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGCTDEIAPPLRCQSFPCLHPFCIPCMKTWIPLRNTCPLCNTPVAYLIVGVTASGSFSTIPIVNDP RTRVEAEAAVRAGTAVDFIWTGNPRTAPRSLSGGSGGGGSGGRAAAWMRQVPDPEDVRVVILYSPLPGEDLAAGRAGGGPPPEWS AERGGLSCLLAALGNRLCGPATAAWAGNWTGAPDVSALGAQGVLLLSTRDLAFAGAVEFLGLLAGACDRRLIVVNAVRAADWPADG PVVSRQHAYLACEVLPAVQCAVRWPAARDLRRTVLASGRVFGPGVFARVEAAHARLYPDAPPLRLCRGANVRYRVRTRFGPDTLVP MSPREYRRAVLPALDGRAAASGGSGGGGSGGETLVAHGPSLYRTFAANPRAASTAKAMRDCVLRQEN
  • a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 643. [0523] In some embodiments, a polyribonucleotide encodes a polypeptide having amino acids 26-587 of amino acid sequence SEQ ID NO: 643.
  • a polyribonucleotide encoding a T-cell string polypeptide can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an UL29 polypeptide or antigenic fragment thereof, a linker, an UL39 polypeptide or antigenic fragment thereof, a linker, an UL9 polypeptide or antigenic fragment thereof, a linker, and a MITD.
  • such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGREDIETIAFIKRFSLDYGAINFINLAPNNVSELAMYYMANQILRYCDHSTYFINTLTAVIAGSR RPPSVQAAAAWAPQGGAGLEAGARALMDSLDAHPGAWTSMFASCNLLRPVMAARPMVVLGLSISKYYGMAGNDRVFQAGNWASL LGGKNACPLLIFDRTRKFVLGGSGGGGSGGRTFGSAPRLTEDDFGLLNYALAEMRRLCLDLPPVPPNAYTPYHLREYATRLVNGFKPL VRRSARLYRILGVLVHLRIRTREASFEEWMRSKEVDLDFGLTERLREHEAQLMILAQALNPYDCLIHSTPNTLVERGLQSALKYEEFYL KRFGGHYMESVFQMYTRIAGFLAGGSGGGGSGGLLNNYDVLVLDEVMSTLGQLYSPTMQQQ
  • a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 644. [0525] In some embodiments, a polyribonucleotide encodes a polypeptide having amino acids 26-638 of amino acid sequence SEQ ID NO: 644.
  • a polyribonucleotide encoding a T-cell string polypeptide can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an UL5.1 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL30.1 polypeptide or antigenic fragment thereof, a linker, and a MITD.
  • such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGHEFGNLMKVLEYGLPITEEHMQFVDRFVVPESYITNPANLPGWTRLFSSHKEVSAYMAKLHA YLKVTREGEFVVFTLPVLTFVSVKEFDEYRRLGGSGGGGSGGELFGEVFESAPFSTYVDNVIFRGCELLTGSPRGGLMSVALQTDNYT LMGYTYTRVFAFAEELRRRHATAGVAEFLEESPLPYIVLRDQHGFMSVVNTNIGGSGGGGSGGTSQCPDINHLRSLSILNRWLETEL VFVGDEEDVSKLSEGELGFYRFLFAFLSAADDLVTENLGGLSGLFEQKDILHYYVEQECIEVVHSRVYNIIQLVLFHNNDQARRAYVAR TINHPAIRVKVDWLEARVRECDSIPEKFILMILIEG
  • a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 645. [0527] In some embodiments, a polyribonucleotide encodes a polypeptide having amino acids 26-543 of amino acid sequence SEQ ID NO: 645.
  • a polyribonucleotide encoding a T-cell string polypeptide can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL21 polypeptide or antigenic fragment thereof, a linker, and a MITD.
  • such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGGPDAAVFRSSLGSLLYWPGVRALLGRDCRVAARYAGRMTYIATGALLARFNPGAVKCVLPRE AAFAGRVLGGSGGGGSGGGLASDPHYDYIRHYASAAKQALGEVELSGGQLSRAILAQYWKYLQTVVPSGLDIPDDPAGDCDPSLHV LLRPTLLPKLLVRAPFKSGAAAAKYAAAVAGLRDAAHRLQQYMFFMRPADPSRPSTDTALRLSELLAYVSVLYHWASWMLWTADKY VGGSGGGGSGGSPTQKLAVYYYLIHRERRMSPFPALVRLVGRYIQRHGLYVPAPDEPTLADAMNGLGGSGGGGSGGIVGIVAGLAVL AVVVIGAVVATVMCRRKSSGGKGGSYSQAASSDSAQGSDVSLTA (SEQ ID
  • a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 646. [0529] In some embodiments, a polyribonucleotide encodes a polypeptide having amino acids 26-335 of amino acid sequence SEQ ID NO: 646.
  • a polyribonucleotide encoding a T-cell string polypeptide can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an UL29 polypeptide or antigenic fragment thereof, a linker, an UL39 polypeptide or antigenic fragment thereof, a linker, an UL9 polypeptide or antigenic fragment thereof, a linker, an UL5.1 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL30.1 polypeptide or antigenic fragment thereof, a linker, and a MITD.
  • such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGREDIETIAFIKRFSLDYGAINFINLAPNNVSELAMYYMANQILRYCDHSTYFINTLTAVIAGSR RPPSVQAAAAWAPQGGAGLEAGARALMDSLDAHPGAWTSMFASCNLLRPVMAARPMVVLGLSISKYYGMAGNDRVFQAGNWASL LGGKNACPLLIFDRTRKFVLGGSGGGGSGGRTFGSAPRLTEDDFGLLNYALAEMRRLCLDLPPVPPNAYTPYHLREYATRLVNGFKPL VRRSARLYRILGVLVHLRIRTREASFEEWMRSKEVDLDFGLTERLREHEAQLMILAQALNPYDCLIHSTPNTLVERGLQSALKYEEFYL KRFGGHYMESVFQMYTRIAGFLAGGSGGGGSGGLLNNYDVLVLDEVMSTLGQLYSPTMQQQ
  • a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 647. [0531] In some embodiments, a polyribonucleotide encodes a polypeptide having amino acids 26-1156 of amino acid sequence SEQ ID NO: 647.
  • a polyribonucleotide encoding a T-cell string polypeptide can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an UL30.1 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL5.1 polypeptide or antigenic fragment thereof, a linker, an UL9 polypeptide or antigenic fragment thereof, a linker, an UL39 polypeptide or antigenic fragment thereof, a linker, an UL29 polypeptide or antigenic fragment thereof, a linker, and a MITD.
  • such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGISCLLYDLSTTALEHILLFSLGSCDLPESHLSDLASRGLPAPVVLEFDSEFEMLLAFMTFVKQYG PEFVTGYNIINFDWPFVLTKLTEIYKVPLDGYGRMNGRGVFRVWDIGQSHFGGSGGGGSGGTSQCPDINHLRSLSILNRWLETELVF VGDEEDVSKLSEGELGFYRFLFAFLSAADDLVTENLGGLSGLFEQKDILHYYVEQECIEVVHSRVYNIIQLVLFHNNDQARRAYVARTI NHPAIRVKVDWLEARVRECDSIPEKFILMILIEGVFFAASFAAIAYLRTNNLLRGGSGGGGSGGELFGEVFESAPFSTYVDNVIFRGCE LLTGSPRGGLMSVALQTDNYTLMGYTYTRVFAFA
  • a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 648. [0533] In some embodiments, a polyribonucleotide encodes a polypeptide having amino acids 26-1156 of amino acid sequence SEQ ID NO: 648.
  • a polyribonucleotide encoding a T-cell string polypeptide can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, a RL2.1 polypeptide or antigenic fragment thereof, a linker, a RS1 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL21 polypeptide or antigenic fragment thereof, a linker, and a MITD.
  • such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGCTDEIAPPLRCQSFPCLHPFCIPCMKTWIPLRNTCPLCNTPVAYLIVGVTASGSFSTIPIVNDP RTRVEAEAAVRAGTAVDFIWTGNPRTAPRSLSGGSGGGGSGGRAAAWMRQVPDPEDVRVVILYSPLPGEDLAAGRAGGGPPPEWS AERGGLSCLLAALGNRLCGPATAAWAGNWTGAPDVSALGAQGVLLLSTRDLAFAGAVEFLGLLAGACDRRLIVVNAVRAADWPADG PVVSRQHAYLACEVLPAVQCAVRWPAARDLRRTVLASGRVFGPGVFARVEAAHARLYPDAPPLRLCRGANVRYRVRTRFGPDTLVP MSPREYRRAVLPALDGRAAASGGSGGGGSGGETLVAHGPSLYRTFAANPRAASTAKAMRDCVLRQEN
  • a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 649. [0535] In some embodiments, a polyribonucleotide encodes a polypeptide having amino acids 26-897 of amino acid sequence SEQ ID NO: 649.
  • a polyribonucleotide encoding a T-cell string polypeptide can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an UL21 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, a RS1 polypeptide or antigenic fragment thereof, a linker, a RL2.1 polypeptide or antigenic fragment thereof, a linker, and a MITD.
  • such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGSPTQKLAVYYYLIHRERRMSPFPALVRLVGRYIQRHGLYVPAPDEPTLADAMNGLGGSGGGG SGGGLASDPHYDYIRHYASAAKQALGEVELSGGQLSRAILAQYWKYLQTVVPSGLDIPDDPAGDCDPSLHVLLRPTLLPKLLVRAPFK SGAAAAKYAAAVAGLRDAAHRLQQYMFFMRPADPSRPSTDTALRLSELLAYVSVLYHWASWMLWTADKYVGGSGGGGSGGGPDA AVFRSSLGSLLYWPGVRALLGRDCRVAARYAGRMTYIATGALLARFNPGAVKCVLPREAAFAGRVLGGSGGGGSGGETLVAHGPSLY RTFAANPRAASTAKAMRDCVLRQENLIEALASADETLAWCKMCIHHNLPLRPQD
  • a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 650. [0537] In some embodiments, a polyribonucleotide encodes a polypeptide having amino acids 26-897 of amino acid sequence SEQ ID NO: 650.
  • a polyribonucleotide encoding a T-cell string polypeptide can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an UL5.1 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL30.1 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL21 polypeptide or antigenic fragment thereof, a linker, and a MITD.
  • such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGHEFGNLMKVLEYGLPITEEHMQFVDRFVVPESYITNPANLPGWTRLFSSHKEVSAYMAKLHA YLKVTREGEFVVFTLPVLTFVSVKEFDEYRRLGGSGGGGSGGELFGEVFESAPFSTYVDNVIFRGCELLTGSPRGGLMSVALQTDNYT LMGYTYTRVFAFAEELRRRHATAGVAEFLEESPLPYIVLRDQHGFMSVVNTNIGGSGGGGSGGTSQCPDINHLRSLSILNRWLETEL VFVGDEEDVSKLSEGELGFYRFLFAFLSAADDLVTENLGGLSGLFEQKDILHYYVEQECIEVVHSRVYNIIQLVLFHNNDQARRAYVAR TINHPAIRVKVDWLEARVRECDSIPEKFILMILIEG
  • a polyribonucleotide encoding a T-cell string polypeptide can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an UL21 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL30.1 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL5.1 polypeptide or antigenic fragment thereof, a linker, and a MITD.
  • such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGSPTQKLAVYYYLIHRERRMSPFPALVRLVGRYIQRHGLYVPAPDEPTLADAMNGLGGSGGGG SGGGLASDPHYDYIRHYASAAKQALGEVELSGGQLSRAILAQYWKYLQTVVPSGLDIPDDPAGDCDPSLHVLLRPTLLPKLLVRAPFK SGAAAAKYAAAVAGLRDAAHRLQQYMFFMRPADPSRPSTDTALRLSELLAYVSVLYHWASWMLWTADKYVGGSGGGGSGGGPDA AVFRSSLGSLLYWPGVRALLGRDCRVAARYAGRMTYIATGALLARFNPGAVKCVLPREAAFAGRVLGGSGGGGSGGISCLLYDLSTT ALEHILLFSLGSCDLPESHLSDLASRGLPAPVVLEFDSEFEMLLAFMTFVK
  • a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 692. [0541] In some embodiments, a polyribonucleotide encodes a polypeptide having amino acids 26-853 of amino acid sequence SEQ ID NO: 692.
  • an RNA construct provided herein comprises a polyribonucleotide that encodes one or more GP polypeptides.
  • an RNA construct provided herein comprises a polyribonucleotide that encodes an HSV-2 gC or antigenic portion thereof.
  • an RNA construct provided herein comprises a polyribonucleotide that encodes an HSV-2 gD or antigenic portion thereof.
  • an RNA construct provided herein comprises a polyribonucleotide that encodes an HSV-2 gE or antigenic portion thereof. In some embodiments, an RNA construct provided herein comprises a polyribonucleotide that encodes an HSV-2 gB, variant, or antigenic portion thereof. In some embodiments, an RNA construct provided herein comprise a polyribonucleotide that encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof. [0545] In some embodiments, an RNA construct provided herein comprises a polyribonucleotide that encodes an antigenic portion of HSV-2 gC.
  • an RNA construct provided herein comprises a polyribonucleotide that encodes an antigenic portion of HSV-2 gD. In some embodiments, an RNA construct provided herein comprises a polyribonucleotide that encodes an antigenic portion of HSV-2 gE. In some embodiments, an RNA construct provided herein comprises a polyribonucleotide that encodes an antigenic portion of HSV-2 gB. In some embodiments, an RNA construct provided herein comprise a polyribonucleotide that encodes at least one T-cell string polypeptide that comprises one or more antigenic portions of HSV T-cell antigens.
  • an RNA construct provided herein comprises a polyribonucleotide that encodes an antigenic portion of HSV-2 gC and a secretory signal. In some embodiments, an RNA construct provided herein comprises a polyribonucleotide that encodes an antigenic portion of HSV-2 gD and a secretory signal. In some embodiments, an RNA construct provided herein comprises a polyribonucleotide that encodes an antigenic portion of HSV-2 gE and a secretory signal. In some embodiments, an RNA construct provided herein comprises a polyribonucleotide that encodes an antigenic portion of HSV-2 gB and a secretory signal.
  • an RNA construct provided herein comprise a polyribonucleotide that encodes at least one T-cell string polypeptide that comprises one or more antigenic portions of HSV T-cell antigens and a secretory signal.
  • polyribonucleotides described herein can comprise a nucleotide sequence that encodes a 5’UTR and/or a 3’ UTR.
  • polynucleotides described herein can comprise a nucleotide sequence that encodes a polyA tail.
  • polyribonucleotides described herein may comprise a 5’ cap, which may be incorporated during transcription, or joined to a polyribonucleotide post- transcription.
  • 5' Cap A structural feature of RNAs is cap structure at five-prime end (5’).
  • Natural eukaryotic RNA comprises a 7-methylguanosine cap linked to the RNA via a 5 ⁇ to 5 ⁇ -triphosphate bridge resulting in cap0 structure (m7GpppN).
  • RNA capping is well researched and is described, e.g., in Decroly E et al. (2012) Nature Reviews 10: 51-65; and in Ramanathan A. et al., (2016) Nucleic Acids Res; 44(16): 7511–7526, the entire contents of each of which is hereby incorporated by reference.
  • a 5’-cap structure which may be suitable in the context of the present invention is a cap0 (methylation of the first nucleobase, e.g., m7GpppN), cap1 (additional methylation of the ribose of the adjacent nucleotide of m7GpppN), cap2 (additional methylation of the ribose of the 2nd nucleotide downstream of the m7GpppN), cap3 (additional methylation of the ribose of the 3rd nucleotide downstream of the m7GpppN), cap4 (additional methylation of the ribose of the 4th nucleotide downstream of the m7GpppN), ARCA (“anti-reverse cap analogue”), modified ARCA (e.g.
  • RNA e.g., mRNA
  • 5'-cap refers to a structure found on the 5'-end of an RNA, e.g., mRNA, and generally includes a guanosine nucleotide connected to an RNA, e.g., mRNA, via a 5'- to 5'-triphosphate linkage (also referred to as Gppp or G(5')ppp(5')).
  • a guanosine nucleoside included in a 5’ cap may be modified, for example, by methylation at one or more positions (e.g., at the 7-position) on a base (guanine), and/or by methylation at one or more positions of a ribose.
  • a guanosine nucleoside included in a 5’ cap comprises a 3’O methylation at a ribose (3’OMeG).
  • a guanosine nucleoside included in a 5’ cap comprises methylation at the 7-position of guanine (m7G).
  • a guanosine nucleoside included in a 5’ cap comprises methylation at the 7-position of guanine and a 3’ O methylation at a ribose (m7(3’OMeG)).
  • m7(3’OMeG) a ribose that is notation used in the above paragraph, e.g., “(m 2 7,3’-O )G” or “m7(3’OMeG)”, applies to other structures described herein.
  • providing an RNA with a 5'-cap disclosed herein may be achieved by in vitro transcription, in which a 5'-cap is co-transcriptionally expressed into an RNA strand, or may be attached to an RNA post-transcriptionally using capping enzymes.
  • co-transcriptional capping with a cap disclosed improves the capping efficiency of an RNA compared to co-transcriptional capping with an appropriate reference comparator.
  • improving capping efficiency can increase a translation efficiency and/or translation rate of an RNA, and/or increase expression of an encoded polypeptide.
  • alterations to polynucleotides generates a non-hydrolyzable cap structure which can, for example, prevent decapping and increase RNA half-life.
  • a utilized 5’ caps is a cap0, a cap1, or cap2 structure. See, e.g., Fig. 1 of Ramanathan A et al., and Fig.
  • an RNA described herein comprises a cap1 structure. In some embodiments, an RNA described herein comprises a cap2. [0553] In some embodiments, an RNA described herein comprises a cap0 structure. In some embodiments, a cap0 structure comprises a guanosine nucleoside methylated at the 7-position of guanine ((m 7 )G).
  • such a cap0 structure is connected to an RNA via a 5'- to 5'-triphosphate linkage and is also referred to herein as (m 7 )Gppp.
  • a cap0 structure comprises a guanosine nucleoside methylated at the 2’-position of the ribose of guanosine.
  • a cap0 structure comprises a guanosine nucleoside methylated at the 3’-position of the ribose of guanosine.
  • a guanosine nucleoside included in a 5’ cap comprises methylation at the 7-position of guanine and at the 2’-position of the ribose ((m 2 7,2’-O )G). In some embodiments, a guanosine nucleoside included in a 5’ cap comprises methylation at the 7- position of guanine and at the 2’-position of the ribose ((m 2 7,3’-O )G).
  • a cap1 structure comprises a guanosine nucleoside methylated at the 7- position of guanine ((m 7 )G) and optionally methylated at the 2’ or 3’ position pf the ribose, and a 2’O methylated first nucleotide in an RNA ((m 2’-O )N 1 ).
  • a cap1 structure comprises a guanosine nucleoside methylated at the 7-position of guanine ((m 7 )G) and the 3’ position of the ribose, and a 2’O methylated first nucleotide in an RNA 1 ).
  • a cap1 structure is connected to an RNA via a 5'- to 5'- triphosphate linkage and is also referred to herein as, e.g., ((m 7 )Gppp( 2'-O )N 1 ) or (m 2 7,3’-O )Gppp( 2'-O )N 1 ), wherein N 1 is as defined and described herein.
  • a cap1 structure comprises a second nucleotide, N 2 , which is at position 2 and is chosen from A, G, C, or U, e.g., (m 7 )Gppp( 2'-O )N 1 pN 2 or (m 2 7,3’-O )Gppp( 2'-O )N 1 pN 2 , wherein each of N 1 and N 2 is as defined and described herein.
  • a cap2 structure comprises a guanosine nucleoside methylated at the 7- position of guanine ((m 7 )G) and optionally methylated at the 2’ or 3’ position of the ribose, and a 2’O methylated first and second nucleotides in an RNA ((m 2’-O )N 1 p(m 2’-O )N 2 ).
  • a cap2 structure comprises a guanosine nucleoside methylated at the 7-position of guanine ((m 7 )G) and the 3’ position of the ribose, and a 2’O methylated first and second nucleotide in an RNA.
  • a cap2 structure is connected to an RNA via a 5'- to 5'-triphosphate linkage and is also referred to herein as, e.g., ((m 7 )Gppp( 2'-O )N 1 p( 2'-O )N 2 ) or (m 2 7,3’- O )Gppp( 2'-O )N 1 p( 2'-O )N 2 ), wherein each of N 1 and N 2 is as defined and described herein.
  • the 5’ cap is a dinucleotide cap structure.
  • the 5’ cap is a dinucleotide cap structure comprising N 1 , wherein N 1 is as defined and described herein.
  • the 5’ cap is a dinucleotide cap G*N 1 , wherein N 1 is as defined above and herein, and G* comprises a structure of formula (I): or a salt thereof, wherein each R 2 and R 3 is -OH or -OCH 3 ; and X is O or S.
  • R 2 is -OH.
  • R 2 is -OCH 3 .
  • R 3 is -OH.
  • R 3 is -OCH 3 .
  • R 2 is -OH and R 3 is -OH. In some embodiments, R 2 is -OH and R 3 is -CH 3 . In some embodiments, R 2 is -CH 3 and R 3 is -OH. In some embodiments, R 2 is -CH 3 and R 3 is - CH 3 . [0558] In some embodiments, X is O. In some embodiments, X is S.
  • the 5’ cap is a dinucleotide cap0 structure (e.g., (m 7 )GpppN 1 , (m 2 7,2’- O )GpppN 1 , (m 2 7,3’-O )GpppN 1 , (m 7 )GppSpN 1 , (m 2 7,2’-O )GppSpN 1 , or (m 2 7,3’-O )GppSpN 1 ), wherein N 1 is as defined and described herein.
  • N 1 is as defined and described herein.
  • the 5’ cap is a dinucleotide cap0 structure (e.g., (m 7 )GpppN 1 , (m 2 7,2’- O )GpppN 1 , (m 2 7,3’-O )GpppN 1 , (m 7 )GppSpN 1 , (m 2 7,2’-O )GppSpN 1 , or (m 2 7,3’-O )GppSpN 1 ), wherein N 1 is G.
  • the 5’ cap is a dinucleotide cap0 structure (e.g., (m 7 )GpppN 1 , (m 2 7,2’-O )GpppN 1 , (m 2 7,3’-O )GpppN 1 , (m 7 )GppSpN 1 , (m 2 7,2’-O )GppSpN 1 , or (m 2 7,3’-O )GppSpN 1 ), wherein N 1 is A, U, or C.
  • a dinucleotide cap0 structure e.g., (m 7 )GpppN 1 , (m 2 7,2’-O )GpppN 1 , (m 2 7,3’-O )GpppN 1 , wherein N 1 is A, U, or C.
  • the 5’ cap is a dinucleotide cap1 structure (e.g., (m 7 )Gppp(m 2’-O )N 1 , (m 2 7,2’-O )Gppp(m 2’-O )N 1 , (m 2 7,3’-O )Gppp(m 2’-O )N 1 , (m 7 )GppSp(m 2’-O )N 1 , (m 2 7,2’-O )GppSp(m 2’-O )N 1 , or (m 2 7,3’-O )GppSp(m 2’-O )N 1 ), wherein N 1 is as defined and described herein.
  • N 1 is as defined and described herein.
  • the 5’ cap is selected from the group consisting of (m 7 )GpppG (“Ecap0”), (m 7 )Gppp(m 2’-O )G (“Ecap1”), (m 2 7,3’-O )GpppG (“ARCA” or “D1”), and (m 2 7,2’-O )GppSpG (“beta-S-ARCA”).
  • the 5’ cap is (m 7 )GpppG (“Ecap0”), having a structure: or a salt thereof.
  • the 5’ cap is (m 7 )Gppp(m 2’-O )G (“Ecap1”), having a structure: or a salt thereof.
  • the 5’ cap is (m 2 7,3’-O )GpppG (“ARCA” or “D1”), having a structure: or a salt thereof.
  • the 5’ cap is (m2 7,2’-O )GppSpG (“beta-S-ARCA”), having a structure: or a salt thereof.
  • the 5’ cap is a trinucleotide cap structure.
  • the 5’ cap is a trinucleotide cap structure comprising N 1 pN 2 , wherein N 1 and N 2 are as defined and described herein.
  • the 5’ cap is a dinucleotide cap G*N 1 pN 2 , wherein N 1 and N 2 are as defined above and herein, and G* comprises a structure of formula (I): or a salt thereof, wherein R 2 , , .
  • the 5’ cap is a trinucleotide cap0 structure (e.g. (m 7 )GpppN 1 pN 2 , (m 2 7,2’- O )GpppN 1 pN 2 , or (m 2 7,3’-O )GpppN 1 pN 2 ), wherein N 1 and N 2 are as defined and described herein).
  • the 5’ cap is a trinucleotide cap1 structure (e.g., (m 7 )Gppp(m 2’-O )N 1 pN 2 , (m 2 7,2’-O )Gppp(m 2’-O )N 1 pN 2 , (m 2 7,3’-O )Gppp(m 2’-O )N 1 pN 2 ), wherein N 1 and N 2 are as defined and described herein.
  • the 5’ cap is a trinucleotide cap2 structure (e.g., (m 7 )Gppp(m 2’-O )N 1 p(m 2’-O )N 2 , (m 2 7,2’-O )Gppp(m 2’-O )N 1 p(m 2’-O )N 2 , (m 2 7,3’- O )Gppp(m 2’-O )N 1 p(m 2’-O )N 2 ), wherein N 1 and N 2 are as defined and described herein.
  • the 5’ cap is selected from the group consisting of (m 2 7,3’-O )Gppp(m 2’-O )ApG (“CleanCap AG”, “CC413”), (m 2 7,3’-O )Gppp(m 2’- O )GpG (“CleanCap GG”), (m 7 )Gppp(m 2’-O )ApG, (m 7 )Gppp(m 2’-O )G, (m 2 7,3’-O )Gppp(m 2 6,2’-O )ApG, and (m 7 )Gppp(m 2’- O )ApU.
  • the 5’ cap is (m 2 7,3’-O )Gppp(m 2’-O )ApG (“CleanCap AG”, “CC413”), having a structure: or a salt there [0566] In some embodiments, the 5’ cap is (m 2 7,3’-O )Gppp(m 2’-O )GpG (“CleanCap GG”), having a structure: or a salt thereo f. [0567] In some embodiments, the 5’ cap is (m 7 )Gppp(m 2’-O )ApG, having a structure:
  • the 5’ cap is (m 7 )Gppp(m 2’-O )GpG, having a structure: or a salt there .
  • the 5’ cap is (m 2 7,3’-O )Gppp(m 2 6,2’-O )ApG, having a structure:
  • the 5’ cap is (m 7 )Gppp(m 2’-O )ApU, having a structure: or a salt thereof.
  • the 5’ cap is a tetranucleotide cap structure.
  • the 5’ cap is a tetranucleotide cap structure comprising N 1 pN 2 pN 3 , wherein N 1 , N 2 , and N 3 are as defined and described herein.
  • the 5’ cap is a tetranucleotide cap G*N 1 pN 2 pN 3 , wherein N 1 , N 2 , and N 3 are as defined above and herein, and G* comprises a structure of formula (I): or a salt thereof, wherein R 2 , [0572]
  • the 5’ cap is a tetranucleotide cap0 structure (e.g.
  • the 5’ cap is a tetranucleotide Cap1 structure (e.g., (m 7 )Gppp(m 2’-O )N 1 pN 2 pN 3 , (m 2 7,2’- O )Gppp(m 2’-O )N 1 pN 2 pN 3 , (m 2 7,3’-O )Gppp(m 2’-O )N 1 pN 2 N 3 ), wherein N 1 , N 2 , and N 3 are as defined and described herein.
  • tetranucleotide Cap1 structure e.g., (m 7 )Gppp(m 2’-O )N 1 pN 2 pN 3 , (m 2 7,2’- O )Gppp(m 2’-O )N 1 pN 2 pN 3 , (m 2 7,3’-O )Gppp(m 2’-O )N 1 pN 2 N 3 ), wherein N 1
  • the 5’ cap is a tetranucleotide Cap2 structure (e.g., (m 7 )Gppp(m 2’-O )N 1 p(m 2’-O )N 2 pN 3 , (m 2 7,2’- O )Gppp(m 2’-O )N 1 p(m 2’-O )N 2 pN 3 , (m 2 7,3’-O )Gppp(m 2’-O )N 1 p(m 2’-O )N 2 pN 3 ), wherein N 1 , N 2 , and N 3 are as defined and described herein.
  • N 1 , N 2 , and N 3 are as defined and described herein.
  • the 5’ cap is selected from the group consisting of (m 2 7,3’-O )Gppp(m 2’- O )Ap(m 2’-O )GpG, (m 2 7,3’-O )Gppp(m 2’-O )Gp(m 2’-O )GpC, (m 7 )Gppp(m 2’-O )Ap(m 2’-O )UpA, and (m 7 )Gppp(m 2’-O )Ap(m 2’-O )GpG.
  • the 5’ cap is (m 2 7,3’-O )Gppp(m 2’-O )Ap(m 2’-O )GpG, having a structure: or a salt thereof.
  • the 5’ cap is (m 2 7,3’-O )Gppp(m 2’-O )Gp(m 2’-O )GpC, having a structure:
  • the 5’ cap is (m 7 )Gppp(m 2’-O )Ap(m 2’-O )UpA, having a structure: or a salt thereo f.
  • the 5’ cap is (m 7 )Gppp(m 2’-O )Ap(m 2’-O )GpG, having a structure:
  • a cap proximal sequence comprises a sequence adjacent to a 5’ cap.
  • a cap proximal sequence comprises nucleotides in positions +1, +2, +3, +4, and/or +5 of an RNA polynucleotide.
  • a cap structure comprises one or more polynucleotides of a cap proximal sequence.
  • a cap structure comprises an m 7 Guanosine cap and nucleotide +1 (N 1 ) of an RNA polynucleotide.
  • a cap structure comprises an m 7 Guanosine cap and nucleotide +2 (N 2 ) of an RNA polynucleotide. In some embodiments, a cap structure comprises an m 7 Guanosine cap and nucleotides +1 and +2 (N 1 and N 2 ) of an RNA polynucleotide. In some embodiments, a cap structure comprises an m 7 Guanosine cap and nucleotides +1, +2, and +3 (N1, N2, and N3) of an RNA polynucleotide.
  • one or more residues of a cap proximal sequence may be included in an RNA by virtue of having been included in a cap entity (e.g., a cap1 or cap2 structure, etc.); alternatively, in some embodiments, at least some of the residues in a cap proximal sequence may be enzymatically added (e.g., by a polymerase such as a T7 polymerase).
  • the 5’ cap is a dinucleotide cap structure, wherein the cap proximal sequence comprises N 1 of the 5’ cap, where N 1 is any nucleotide, e.g., A, C, G or U.
  • the 5’ cap is a trinucleotide cap structure (e.g., the trinucleotide cap structures described above and herein), wherein the cap proximal sequence comprises N 1 and N 2 of the 5’ cap, wherein N 1 and N 2 are independently any nucleotide, e.g., A, C, G or U.
  • the 5’ cap is a tetranucleotide cap structure (e.g., the trinucleotide cap structures described above and herein), wherein the cap proximal sequence comprises N 1 , N 2 , and N 3 of the 5’ cap, wherein N 1 , N 2 , and N 3 are any nucleotide, e.g., A, C, G or U.
  • a cap proximal sequence comprises N 1 of a the 5’ cap, and N 2 , N 3 , N 4 and N 5 , wherein N 1 to N 5 correspond to positions +1, +2, +3, +4, and/or +5 of an RNA polynucleotide.
  • a cap proximal sequence comprises N 1 and N 2 of a the 5’ cap, and N 3 , N 4 and N 5 , wherein N 1 to N 5 correspond to positions +1, +2, +3, +4, and/or +5 of an RNA polynucleotide.
  • a cap proximal sequence comprises N 1 , N 2 , and N 3 of a the 5’ cap, and N 4 and N 5 , wherein N 1 to N 5 correspond to positions +1, +2, +3, +4, and/or +5 of an RNA polynucleotide.
  • N 1 is A.
  • N 1 is C.
  • N 1 is G.
  • N 1 is U.
  • N 2 is A.
  • N 2 is C.
  • N 2 is G.
  • N 2 is U.
  • N 3 is A. In some embodiments, N 3 is C. In some embodiments, N 3 is G. In some embodiments, N 3 is U. In some embodiments, N 4 is A. In some embodiments, N 4 is C. In some embodiments, N 4 is G. In some embodiments, N 4 is U. In some embodiments, N 5 is A. In some embodiments, N 5 is C. In some embodiments, N 5 is G. In some embodiments, N 5 is U. It will be understood that each of the embodiments described above and herein (e.g., for N 1 through N 5 ) may be taken singly or in combination and/or may be combined with other embodiments of variables described above and herein (e.g., 5’ caps).
  • a cap proximal sequence comprises A 1 and G 2 of the Cap1 structure, and a sequence comprising: A 3 A 4 U 5 (SEQ ID NO: 150) at positions +3, +4 and +5 respectively of the polyribonucleotide.
  • a nucleic acid e.g., DNA, RNA
  • 5’-UTR may comprise a plurality of distinct sequence elements; in some embodiments, such plurality may be or comprise multiple copies of one or more particular sequence elements (e.g., as may be from a particular source or otherwise known as a functional or characteristic sequence element).
  • a 5’ UTR comprises multiple different sequence elements.
  • the term “untranslated region” or “UTR” is commonly used in the art to a region in a DNA molecule which is transcribed but is not translated into an amino acid sequence, or to the corresponding region in an RNA polynucleotide, such as an mRNA molecule.
  • An untranslated region (UTR) can be present 5' (upstream) of an open reading frame (5'-UTR) and/or 3' (downstream) of an open reading frame (3'-UTR).
  • the terms “five prime untranslated region” or “5' UTR” refer to a sequence of a polyribonucleotide between the 5' end of the polyribonucleotide (e.g., a transcription start site) and a start codon of a coding region of the polyribonucleotide.
  • “5' UTR” refers to a sequence of a polyribonucleotide that begins at the 5' end of the polyribonucleotide (e.g., a transcription start site) and ends one nucleotide (nt) before a start codon (usually AUG) of a coding region of the polyribonucleotide, e.g., in its natural context.
  • a 5' UTR comprises a Kozak sequence.
  • a 5'-UTR is downstream of the 5'-cap (if present), e.g., directly adjacent to the 5'-cap.
  • a 5’ UTR disclosed herein comprises a cap proximal sequence, e.g., as defined and described herein.
  • a cap proximal sequence comprises a sequence adjacent to a 5’ cap.
  • Example 5’ UTRs include a human alpha globin (hAg) 5’UTR or a portion thereof, a TEV 5’ UTR or a portion thereof, a HSP705’ UTR or a portion thereof, or a c-Jun 5’ UTR or a portion thereof.
  • an RNA disclosed herein comprises a hAg 5’ UTR or a portion thereof.
  • an RNA disclosed herein comprises a 5’ UTR having at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to a 5’ UTR with the sequence AGAATAAACTAGTATTCTTCTGGTCCCCACAGACTCAGAGAGAACCCGCCACC (SEQ ID NO: 151).
  • an RNA disclosed herein comprises a 5’ UTR having the sequence AGAATAAACTAGTATTCTTCTGGTCCCCACAGACTCAGAGAGAACCCGCCACC (SEQ ID NO: 151).
  • an RNA disclosed herein comprises a 5’ UTR having at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to a 5’ UTR with the sequence AACUAGUAUUCUUCUGGUCCCCACAGACUCAGAGAACCCGCCACC (SEQ ID NO: 152)(hAg- Kozak/5'UTR).
  • an RNA disclosed herein comprises a 5’ UTR having the sequence AACUAGUAUUCUUCUGGUCCCCACAGACUCAGAGAACCCGCCACC (SEQ ID NO: 152)(hAg-Kozak/5'UTR).
  • a polynucleotide e.g., DNA, RNA
  • a polyA sequence is situated downstream of a 3'-UTR, e.g., adjacent to a 3'-UTR.
  • poly(A) sequence or “poly-A tail” refers to an uninterrupted or interrupted sequence of adenylate residues which is typically located at the 3'-end of an RNA polynucleotide.
  • Poly(A) sequences are known to those of skill in the art and may follow the 3’-UTR in the RNAs described herein.
  • An uninterrupted poly(A) sequence is characterized by consecutive adenylate residues. In nature, an uninterrupted poly(A) sequence is typical.
  • polynucleotides disclosed herein comprise an uninterrupted Poly(A) sequence.
  • polynucleotides disclosed herein comprise interrupted Poly(A) sequence.
  • RNAs disclosed herein can have a poly(A) sequence attached to the free 3'-end of the RNA by a template-independent RNA polymerase after transcription or a poly(A) sequence encoded by DNA and transcribed by a template-dependent RNA polymerase.
  • a poly(A) sequence (SEQ ID NO: 829) of about 120 A nucleotides has a beneficial influence on the levels of RNA in transfected eukaryotic cells, as well as on the levels of protein that is translated from an open reading frame that is present upstream (5’) of the poly(A) sequence (Holtkamp et al., 2006, Blood, vol. 108, pp. 4009-4017, which is herein incorporated by reference).
  • a poly(A) sequence in accordance with the present disclosure is not limited to a particular length; in some embodiments, a poly(A) sequence is any length.
  • a poly(A) sequence comprises, essentially consists of, or consists of at least 20, at least 30, at least 40, at least 80, or at least 100 and up to 500, up to 400, up to 300, up to 200, or up to 150 A nucleotides, and, in particular, about 120 A nucleotides.
  • nucleotides in the poly(A) sequence typically at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% by number of nucleotides in the poly(A) sequence are A nucleotides, but permits that remaining nucleotides are nucleotides other than A nucleotides, such as U nucleotides (uridylate), G nucleotides (guanylate), or C nucleotides (cytidylate).
  • a poly(A) sequence is attached during RNA transcription, e.g., during preparation of in vitro transcribed RNA, based on a DNA template comprising repeated dT nucleotides (deoxythymidylate) in the strand complementary to the coding strand.
  • the DNA sequence encoding a poly(A) sequence (coding strand) is referred to as poly(A) cassette.
  • the poly(A) cassette present in the coding strand of DNA essentially consists of dA nucleotides, but is interrupted by a random sequence of the four nucleotides (dA, dC, dG, and dT). Such random sequence may be 5 to 50, 10 to 30, or 10 to 20 nucleotides in length.
  • a cassette is disclosed in WO 2016/005324 A1, hereby incorporated by reference. Any poly(A) cassette disclosed in WO 2016/005324 A1, which is incorporated herein by reference in its entirety, may be used in accordance with the present disclosure.
  • a poly(A) cassette that essentially consists of dA nucleotides, but is interrupted by a random sequence having an equal distribution of the four nucleotides (dA, dC, dG, dT) and having a length of e.g., 5 to 50 nucleotides shows, on DNA level, constant propagation of plasmid DNA in E. coli and is still associated, on RNA level, with the beneficial properties with respect to supporting RNA stability and translational efficiency is encompassed.
  • the poly(A) sequence contained in an RNA polynucleotide described herein essentially consists of A nucleotides, but is interrupted by a random sequence of the four nucleotides (A, C, G, U).
  • Such random sequence may be 5 to 50, 10 to 30, or 10 to 20 nucleotides in length.
  • no nucleotides other than A nucleotides flank a poly(A) sequence at its 3'- end, i.e., the poly(A) sequence is not masked or followed at its 3'-end by a nucleotide other than A.
  • the poly(A) sequence (SEQ ID NO: 829) may comprise at least 20, at least 30, at least 40, at least 80, or at least 100 and up to 500, up to 400, up to 300, up to 200, or up to 150 nucleotides.
  • the poly(A) sequence may essentially consist of at least 20, at least 30, at least 40, at least 80, or at least 100 and up to 500, up to 400, up to 300, up to 200, or up to 150 nucleotides. In some embodiments, the poly(A) sequence (SEQ ID NO: 829) may consist of at least 20, at least 30, at least 40, at least 80, or at least 100 and up to 500, up to 400, up to 300, up to 200, or up to 150 nucleotides. In some embodiments, the poly(A) sequence (SEQ ID NO: 829) comprises at least 100 nucleotides.
  • the poly(A) sequence (SEQ ID NO: 829) comprises about 150 nucleotides. In some embodiments, the poly(A) sequence comprises about 120 nucleotides.
  • a poly A tail comprises a specific number of Adenosines, such as about 50 or more, about 60 or more, about 70 or more, about 80 or more, about 90 or more, about 100 or more, about 120, or about 150 or about 200. In some embodiments a poly A tail of a string construct may comprise 200 A residues or less. In some embodiments, a poly A tail of a string construct may comprise about 200 A residues. In some embodiments, a poly A tail of a string construct may comprise 180 A residues or less.
  • RNA comprises a poly(A) sequence comprising the nucleotide sequence of AAAAAAAAAAAAAAAAAAAAAAAAAAGCATATGACTAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAA (SEQ ID NO: 153).
  • a poly(A) tail comprises a plurality of A residues interrupted by a linker.
  • a linker comprises the nucleotide sequence GCATATGAC (SEQ ID NO: 154).
  • RNA comprises a poly(A) sequence comprising the nucleotide sequence of AAAAAAAAAAAAAAAAAAAAAAAAAAAAGCAUAUGACUAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAA (SEQ ID NO: 155), or a nucleotide sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the nucleotide sequence of AAAAAAAAAAAAAAAAAAA
  • a poly(A) tail comprises a plurality of A residues interrupted by a linker.
  • a linker comprises the nucleotide sequence GCAUAUGAC (SEQ ID NO: 156). 5.
  • 3' UTR [0601]
  • an RNA utilized in accordance with the present disclosure comprises a 3'- UTR.
  • the terms “three prime untranslated region,” “3' untranslated region,” or “3' UTR” refer to a sequence of an RNA molecule that begins following a stop codon of a coding region of an open reading frame sequence.
  • the 3' UTR begins immediately after a stop codon of a coding region of an open reading frame sequence, e.g., in its natural context. In other embodiments, the 3' UTR does not begin immediately after stop codon of the coding region of an open reading frame sequence, e.g., in its natural context.
  • the term “3'- UTR” does preferably not include the poly(A) sequence. Thus, the 3'-UTR is upstream of the poly(A) sequence (if present), e.g., directly adjacent to the poly(A) sequence.
  • an RNA disclosed herein comprises a 3’ UTR comprising an F element d/ I l I b di 3’ UTR i l h i i i i e.
  • element sequence is a 3’-UTR of amino-terminal enhancer of split (AES).
  • an RNA disclosed herein comprises a 3’ UTR having at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to a 3’ UTR with the sequence of CTGGTACTGCATGCACGCAATGCTAGCTGCCCCTTTCCCGTCCTGGGTACCCCGAGTCTCCCCCGACCTCGGGTCCCAGGTATG CTCCCACCTCCACCTGCCCCACTCACCACCTCTGCTAGTTCCAGACACCTCCCAAGCACGCAGCAATGCAGCTCAAAACGCTTAG CCTAGCCACACCCCCACGGGAAACAGCAGTGATTAACCTTTAGCAATAAACGAAAGTTTAACTAAGCTATACTAACCCCAGGGTT GGTCAATTTCGTGCCAGCCACACC (SEQ ID NO: 157).
  • an RNA disclosed herein comprises a 3’ UTR with the sequence of CTGGTACTGCATGCACGCAATGCTAGCTGCCCCTTTCCCGTCCTGGGTACCCCGAGTCTCCCCCGACCTCGGGTCCCAGGTATG CTCCCACCTCCACCTGCCCCACTCACCACCTCTGCTAGTTCCAGACACCTCCCAAGCACGCAGCAATGCAGCTCAAAACGCTTAG CCTAGCCACACCCCCGGGAAACAGCAGTGATTAACCTTTAGCAATAAACGAAAGTTTAACTAAGCTATACTAACCCCAGGGTT GGTCAATTTCGTGCCAGCCACACC (SEQ ID NO: 157).
  • an RNA disclosed herein comprises a 3’ UTR having at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to a 3’ UTR with the sequence of CUGGUACUGCAUGCACGCAAUGCUAGCUGCCCCUUUCCCGUCCUGGGUACCCCGAGUCUCCCCCGACCUCGGGUCCCAGGUA UGCUCCCACCUCCACCUGCCCCACUCACCACCUCUGCUAGUUCCAGACACCUCCCAAGCACGCAGCAAUGCAGCUCAAAACGCU UAGCCUAGCCACACCCCCACGGGAAACAGCAGUGAUUAACCUUUAGCAAUAAACGAAAGUUUAACUAAGCUAUACUAACCCCA GGGUUGGUCAAUUUCGUGCCAGCCACACC (SEQ ID NO: 158).
  • an RNA disclosed herein comprises a 3’ UTR with the sequence of CUGGUACUGCAUGCACGCAAUGCUAGCUGCCCCUUUCCCGUCCUGGGUACCCCGAGUCUCCCCCGACCUCGGGUCCCAGGUA UGCUCCCACCUCCACCUGCCCCACUCACCACCUCUGCUAGUUCCAGACACCUCCCAAGCACGCAGCAAUGCAGCUCAAAACGCU UAGCCUAGCCACACCCCCACGGGAAACAGCAGUGAUUAACCUUUAGCAAUAAACGAAAGUUUAACUAAGCUAUACUAACCCCA GGGUUGGUCAAUUUCGUGCCAGCCACACC (SEQ ID NO: 158).
  • a 3’UTR is an FI element as described in WO2017/060314, which is herein incorporated by reference in its entirety.
  • RNA Formats At least three distinct formats useful for RNA compositions (e.g., pharmaceutical compositions) have been developed, namely non-modified uridine containing RNA (uRNA), nucleoside-modified mRNA (modRNA), and self-amplifying RNA (saRNA). Each of these platforms displays unique features. In general, in all three formats, RNA is capped, contains open reading frames (ORFs) flanked by untranslated regions (UTR), and have a polyA-tail at the 3' end.
  • ORFs open reading frames flanked by untranslated regions
  • an ORF of an uRNA and modRNA vectors encode an antibody agent or portion thereof.
  • An saRNA has multiple ORFs.
  • the RNA described herein may have modified nucleosides.
  • the RNA comprises a modified nucleoside in place of at least one (e.g., every) uridine.
  • uracil describes one of the nucleobases that can occur in the nucleic acid of RNA.
  • the structure of uracil is: .
  • the structure of uridine is: .
  • UTP uridine 5’-triphosphate
  • Pseudo-UTP Pseudo owing structure: .
  • “Pseudouridine” s o e exa pe o a o e uceos e at is an isomer of uridine, where the uracil is attached to the pentose ring via a carbon-carbon bond instead of a nitrogen-carbon glycosidic bond.
  • Another exemplary modified nucleoside is N1-methyl-pseudouridine (m1 ⁇ ), which has the structure: .
  • N1-methyl-pseudo-UTP has the following structure: .
  • Another exempl m5U which has the structure: .
  • the modified nucleoside is a modified uridine.
  • RNA comprises a modified nucleoside in place of at least one uridine. In some embodiments, RNA comprises a modified nucleoside in place of each uridine.
  • the modified nucleoside is independently selected from pseudouridine ( ⁇ ), N1-methyl-pseudouridine (m1 ⁇ ), and 5-methyl-uridine (m5U).
  • the modified nucleoside comprises pseudouridine ( ⁇ ).
  • the modified nucleoside comprises N1-methyl-pseudouridine (m1 ⁇ ).
  • the modified nucleoside comprises 5-methyl-uridine (m5U).
  • RNA may comprise more than one type of modified nucleoside, and the modified nucleosides are independently selected from pseudouridine ( ⁇ ), N1-methyl-pseudouridine (m1 ⁇ ), and 5-methyl-uridine (m5U).
  • the modified nucleosides comprise pseudouridine ( ⁇ ) and N1-methyl-pseudouridine (m1 ⁇ ). In some embodiments, the modified nucleosides comprise pseudouridine ( ⁇ ) and 5-methyl-uridine (m5U). In some embodiments, the modified nucleosides comprise N1-methyl-pseudouridine (m1 ⁇ ) and 5-methyl-uridine (m5U). In some embodiments, the modified nucleosides comprise pseudouridine ( ⁇ ), N1-methyl-pseudouridine (m1 ⁇ ), and 5- methyl-uridine (m5U).
  • the modified nucleoside replacing one or more, e.g., all, uridine in the RNA may be any one or more of 3-methyl-uridine (m3U), 5-methoxy-uridine (mo5U), 5-aza-uridine, 6-aza-uridine, 2-thio- 5-aza-uridine, 2-thio-uridine (s2U), 4-thio-uridine (s4U), 4-thio-pseudouridine, 2-thio-pseudouridine, 5-hydroxy- uridine (ho5U), 5-aminoallyl-uridine, 5-halo-uridine (e.g., 5-iodo-uridine or 5-bromo-uridine), uridine 5-oxyacetic acid (cmo5U), uridine 5-oxyacetic acid methyl ester (mcmo5U), 5-carboxymethyl-uridine (cm5U), 1-carboxymethyl- pseudouridine, 5-carboxyhydroxymethyl-uridine (chm
  • the RNA comprises other modified nucleosides or comprises further modified nucleosides, e.g., modified cytidine.
  • modified cytidine in the RNA 5-methylcytidine is substituted partially or completely, preferably completely, for cytidine.
  • the RNA comprises 5- methylcytidine and one or more selected from pseudouridine ( ⁇ ), N1-methyl-pseudouridine (m1 ⁇ ), and 5-methyl- uridine (m5U).
  • the RNA comprises 5-methylcytidine and N1-methyl-pseudouridine (m1 ⁇ ).
  • the RNA comprises 5-methylcytidine in place of each cytidine and N1-methyl-pseudouridine (m1 ⁇ ) in place of each uridine.
  • the RNA is “replicon RNA” or simply a “replicon,” in particular “self-replicating RNA” or “self-amplifying RNA.”
  • the replicon or self-replicating RNA is derived from or comprises elements derived from a single-stranded (ss) RNA virus, in particular a positive-stranded ssRNA virus, such as an alphavirus. Alphaviruses are typical representatives of positive- stranded RNA viruses.
  • Alphaviruses replicate in the cytoplasm of infected cells (for review of the alphaviral life cycle see Jose et al., Future Microbiol., 2009, vol. 4, pp. 837–856, which is incorporated herein by reference in its entirety).
  • the total genome length of many alphaviruses typically ranges between 11,000 and 12,000 nucleotides, and the genomic RNA typically has a 5’-cap, and a 3’ poly(A) tail.
  • the genome of alphaviruses encodes non-structural proteins (involved in transcription, modification and replication of viral RNA and in protein modification) and structural proteins (forming the virus particle). There are typically two open reading frames (ORFs) in the genome.
  • the four non-structural proteins are typically encoded together by a first ORF beginning near the 5′ terminus of the genome, while alphavirus structural proteins are encoded together by a second ORF which is found downstream of the first ORF and extends near the 3’ terminus of the genome.
  • first ORF is larger than the second ORF, the ratio being roughly 2:1.
  • RNA RNA molecule that resembles eukaryotic messenger RNA
  • mRNA messenger RNA
  • the (+) stranded genomic RNA directly acts like a messenger RNA for the translation of the open reading frame encoding the non-structural poly- protein (nsP1234).
  • Alphavirus-derived vectors have been proposed for delivery of foreign genetic information into target cells or target organisms.
  • a first ORF encodes an alphavirus-derived RNA-dependent RNA polymerase (replicase), which upon translation mediates self-amplification of the RNA.
  • a second ORF encoding alphaviral structural proteins is replaced by an open reading frame encoding an HSV-2 construct described herein.
  • Alphavirus-based trans-replication systems rely on alphavirus nucleotide sequence elements on two separate nucleic acid molecules: one nucleic acid molecule encodes a viral replicase, and the other nucleic acid molecule is capable of being replicated by said replicase in trans (hence the designation trans-replication system).
  • Trans-replication requires the presence of both these nucleic acid molecules in a given host cell.
  • the nucleic acid molecule capable of being replicated by the replicase in trans must comprise certain alphaviral sequence elements to allow recognition and RNA synthesis by the alphaviral replicase.
  • a non-modified uridine platform may include, for example, one or more of intrinsic adjuvant effect, as well as good tolerability and safety.
  • modified uridine (e.g., pseudouridine) platform may include reduced adjuvant effect, blunted immune innate immune sensor activating capacity and thus good tolerability and safety.
  • self-amplifying platform may include, for example, long duration of protein expression, good tolerability and safety, higher likelihood for efficacy with very low composition (e.g., immunogenic composition, e.g., vaccine) dose.
  • very low composition e.g., immunogenic composition, e.g., vaccine
  • the present disclosure provides particular RNA constructs optimized, for example, for improved manufacturability, encapsulation, expression level (and/or timing), etc. Certain components are discussed below, and certain preferred embodiments are exemplified herein. C.
  • Codon Optimization and GC Enrichment refers to alteration of codons in a coding region of a nucleic acid molecule (e.g., a polyribonucleotide) to reflect the typical codon usage of a host organism (e.g., a subject receiving a nucleic acid molecule (e.g., a polyribonucleotide)) without preferably altering the amino acid sequence encoded by the nucleic acid molecule.
  • coding regions are codon-optimized for optimal expression in a subject to be treated using the RNA molecules described herein.
  • codon-optimization may be performed such that codons for which frequently occurring tRNAs are available are inserted in place of “rare codons.”
  • codon- optimization may include increasing guanosine/cytosine (G/C) content of a coding region of RNA described herein as compared to the G/C content of the corresponding coding sequence of a wild type RNA, wherein the amino acid sequence encoded by the RNA is preferably not modified compared to the amino acid sequence.
  • G/C guanosine/cytosine
  • a coding sequence (also referred to as a “coding region”) is codon optimized for expression in the subject to whom a composition (e.g., a pharmaceutical composition) is to be administered (e.g., a human).
  • a composition e.g., a pharmaceutical composition
  • sequences in such a polynucleotide may differ from wild type sequences encoding the relevant antigen or portion or epitope thereof, even when the amino acid sequence of the antigen or portion or epitope thereof is wild type.
  • strategies for codon optimization for expression in a relevant subject e.g., a human
  • a relevant subject e.g., a human
  • strategies for codon optimization for expression in a particular cell or tissue e.g., a relevant subject
  • a relevant subject e.g., a human
  • Various species exhibit particular bias for certain codons of a particular amino acid.
  • codon bias differences in codon usage between organisms
  • codon bias often correlates with the efficiency of translation of messenger RNA (mRNA), which is in turn believed to be dependent on, among other things, the properties of the codons being translated and the availability of particular transfer RNA (tRNA) molecules.
  • mRNA messenger RNA
  • tRNA transfer RNA
  • the predominance of selected tRNAs in a cell may generally be a reflection of the codons used most frequently in peptide synthesis.
  • genes may be tailored for optimal gene expression in a given organism based on codon optimization.
  • Codon usage tables are available, for example, at the "Codon Usage Database” available at www.kazusa.orjp/codon/ and these tables may be adapted in a number of ways.
  • Computer algorithms for codon optimizing a particular sequence for expression in a particular subject or its cells are also available, such as Gene Forge (Aptagen; Jacobus, PA), are also available.
  • a polynucleotide (e.g., a polyribonucleotide) of the present disclosure is codon optimized, wherein the codons in the polynucleotide (e.g., the polyribonucleotide) are adapted to human codon usage (herein referred to as “human codon optimized polynucleotide”).
  • a portion of a polyribonucleotide is codon optimized (e.g., a portion of or the portion encoding a glycoprotein or a portion of or the portion encoding a secretory signal).
  • the entire polyribonucleotide is codon optimized.
  • Codons encoding the same amino acid occur at different frequencies in a subject, e.g., a human. Accordingly, in some embodiments, the coding sequence of a polynucleotide of the present disclosure is modified such that the frequency of the codons encoding the same amino acid corresponds to the naturally occurring frequency of that codon according to the human codon usage, e.g., as shown in Table 20.
  • the wild type coding sequence is preferably adapted in a way that the codon “GCC” is used with a frequency of 0.40, the codon “GCT” is used with a frequency of 0.28, the codon “GCA” is used with a frequency of 0.22 and the codon “GCG” is used with 30 a frequency of 0.10 etc. (see Table 20). Accordingly, in some embodiments, such a procedure ⁇ as exemplified for Ala) is applied for each amino acid encoded by the coding sequence of a polynucleotide to obtain sequences adapted to human codon usage. Table 20: Human codon usage with frequencies indicated for each amino acid.
  • a coding sequence may be optimized using a multiparametric optimization strategy.
  • optimization parameters may include parameters that influence protein expression, which can be, for example, impacted on a transcription level, an RNA level, and/or a translational level.
  • exemplary optimization parameters include, but are not limited to transcription-level parameters (including, e.g., GC content, consensus splice sites, cryptic splice sites, SD sequences, TATA boxes, termination signals, artificial recombination sites, and combinations thereof); RNA-level parameters (including, e.g., RNA instability motifs, ribosomal entry sites, repetitive sequences, and combinations thereof); translation-level parameters (including, e.g., codon usage, premature poly(A) sites, ribosomal entry sites, secondary structures, and combinations thereof); or combinations thereof.
  • a coding sequence may be optimized by a GeneOptimizer algorithm as described in Fath et al.
  • a coding sequence may be optimized by Eurofins’ adaption and optimization algorithm “GENEius” as described in Eurofins’ Application Notes: Eurofins’ adaption and optimization software “GENEius” in comparison to other optimization algorithms, the entire content of which is incorporated by reference for the purposes described herein.
  • a coding sequence utilized in accordance with the present disclosure has G/C content that is increased compared to a coding sequence for an HSV gC, gD, and/or gE (or portion thereof) construct described herein.
  • guanosine/cytidine (G/C) content of a coding region is modified relative to a comparable coding sequence for an HSV gC, gD, and/or gE (or portion thereof) construct described herein, but the amino acid sequence encoded by the polyribonucleotide not modified.
  • G/C guanosine/cytidine
  • GC enrichment may improve translation of a payload sequence.
  • sequences having an increased G (guanosine)/C (cytidine) content are more stable than sequences having an increased A (adenosine)/U (uridine) content.
  • codons which contain A and/or U nucleosides can be modified by substituting these codons by other codons, which code for the same amino acids but contain no A and/or U or contain a lower content of A and/or U nucleosides.
  • G/C content of a coding region of a polyribonucleotide described herein is increased by at least 1%, at least 2%, at least 3%, at least 4%, at least 5%, at least 6%, or even more compared to the G/C content of the coding region prior to codon optimization, e.g., of the wild type RNA.
  • G/C content of a coding region of a polyribonucleotide described herein is decreased by at least 1%, at least 2%, at least 3%, at least 4%, at least 5%, at least 6%, or even more compared to the G/C content of the coding region prior to codon optimization, e.g., of the wild type RNA.
  • G/C content of a coding region of a polyribonucleotide provided herein is decreased by at least 1%, at least 2%, at least 3%, at least 4%, at least 5%, at least 6%, or even more compared to the G/C content of the coding region prior to codon optimization, e.g., of the wild type RNA.
  • G/C content of a coding region of a polyribonucleotide provided herein comprises at the most 65%, such as at the most 64%, such as at the most 63%, such as at the most 62%, such as at the most 61%, such as at the most 60% G/C content compared to the G/C content of the coding region prior to codon optimization, e.g., of the wild type RNA.
  • C content of a coding region of a polyribonucleotide provided herein is decreased by at least 1%, at least 2%, at least 3%, at least 4%, at least 5%, at least 6%, or even more compared to the C content of the coding region prior to codon optimization, e.g., of the wild type RNA.
  • C content of a coding region of a polyribonucleotide provided herein comprises at the most 40%, such as at the most 39%, such as at the most 38%, such as at the most 37%, such as at the most 36%, such as at the most 35% C content compared to the C content of the coding region prior to codon optimization, e.g., of the wild type RNA.
  • stability and translation efficiency of a polyribonucleotide may incorporate one or more elements established to contribute to stability and/or translation efficiency of the polyribonucleotide; exemplary such elements are described, for example, in PCT/EP2006/009448 incorporated herein by reference.
  • a polyribonucleotide may be modified within the coding region, i.e., the sequence encoding the expressed peptide or protein, without altering the sequence of the expressed peptide or protein, for example so as to increase the GC- content to increase RNA stability and/or to perform a codon optimization and, thus, enhance translation in cells.
  • a polyribonucleotide provided herein encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 65.
  • an RNA construct comprises a 5’ cap, a 5’UTR, a polyribonucleotide that encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 65, a 3’ UTR, and a polyA tail.
  • a polyribonucleotide provided herein encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 70.
  • an RNA construct comprises a 5’ cap, a 5’UTR, a polyribonucleotide that encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 70, a 3’ UTR, and a polyA tail.
  • a polyribonucleotide provided herein encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 73.
  • an RNA construct comprises a 5’ cap, a 5’UTR, a polyribonucleotide that encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 73, a 3’ UTR, and a polyA tail.
  • a polyribonucleotide provided herein encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 67.
  • an RNA construct comprises a 5’ cap, a 5’UTR, a polyribonucleotide that encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 67, a 3’ UTR, and a polyA tail.
  • a polyribonucleotide provided herein encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 68.
  • an RNA construct comprises a 5’ cap, a 5’UTR, a polyribonucleotide that encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 68, a 3’ UTR, and a polyA tail.
  • a polyribonucleotide provided herein encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 75.
  • an RNA construct comprises a 5’ cap, a 5’UTR, a polyribonucleotide that encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 75, a 3’ UTR, and a polyA tail.
  • a polyribonucleotide provided herein encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 131.
  • an RNA construct comprises a 5’ cap, a 5’UTR, a polyribonucleotide that encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 131, a 3’ UTR, and a polyA tail.
  • a polyribonucleotide provided herein encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 132.
  • an RNA construct comprises a 5’ cap, a 5’UTR, a polyribonucleotide that encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 132, a 3’ UTR, and a polyA tail.
  • a polyribonucleotide provided herein encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 159.
  • an RNA construct comprises a 5’ cap, a 5’UTR, a polyribonucleotide that encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 159, a 3’ UTR, and a polyA tail.
  • a polyribonucleotide provided herein encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 160.
  • an RNA construct comprises a 5’ cap, a 5’UTR, a polyribonucleotide that encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 160, a 3’ UTR, and a polyA tail.
  • a polyribonucleotide provided herein encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 161.
  • an RNA construct comprises a 5’ cap, a 5’UTR, a polyribonucleotide that encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 161, a 3’ UTR, and a polyA tail.
  • a polyribonucleotide provided herein encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 162.
  • an RNA construct comprises a 5’ cap, a 5’UTR, a polyribonucleotide that encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 162, a 3’ UTR, and a polyA tail.
  • a polyribonucleotide provided herein encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 163.
  • an RNA construct comprises a 5’ cap, a 5’UTR, a polyribonucleotide that encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 163, a 3’ UTR, and a polyA tail.
  • a polyribonucleotide provided herein encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 164.
  • an RNA construct comprises a 5’ cap, a 5’UTR, a polyribonucleotide that encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 164, a 3’ UTR, and a polyA tail.
  • a polyribonucleotide provided herein encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 327.
  • an RNA construct comprises a 5’ cap, a 5’UTR, a polyribonucleotide that encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 327, a 3’ UTR, and a polyA tail.
  • a polyribonucleotide provided herein encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 328.
  • an RNA construct comprises a 5’ cap, a 5’UTR, a polyribonucleotide that encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 328, a 3’ UTR, and a polyA tail.
  • RNA Delivery Technologies Provided polyribonucleotides may be delivered for therapeutic applications described herein using any appropriate methods known in the art, including, e.g., delivery as naked RNAs, or delivery mediated by viral and/or non-viral vectors, polymer-based vectors, lipid compositions, nanoparticles (e.g., lipid nanoparticles, polymeric nanoparticles, lipid-polymer hybrid nanoparticles, etc.), and/or peptide-based vectors. See, e.g., Wadhwa et al.
  • one or more polyribonucleotides can be formulated with lipid nanoparticles for delivery (e.g., administration).
  • lipid nanoparticles can be designed to protect polyribonucleotides from extracellular RNases and/or engineered for systemic delivery of the RNA to target cells.
  • lipid nanoparticles may be particularly useful to deliver polyribonucleotides when polyribonucleotides are intravenously or intramuscularly administered to a subject.
  • A. Lipid Compositions 1. Lipids and Lipid-Like Materials [0657] The terms "lipid” and “lipid-like material” are broadly defined herein as molecules which comprise one or more hydrophobic moieties or groups and optionally also one or more hydrophilic moieties or groups. Molecules comprising hydrophobic moieties and hydrophilic moieties are also frequently denoted as amphiphiles. Lipids are usually poorly soluble in water. In an aqueous environment, the amphiphilic nature allows the molecules to self-assemble into organized structures and different phases.
  • One of those phases consists of lipid bilayers, as they are present in vesicles, multilamellar/unilamellar liposomes, or membranes in an aqueous environment.
  • Hydrophobicity can be conferred by the inclusion of a polar groups that include, but are not limited to, long-chain saturated and unsaturated aliphatic hydrocarbon groups and such groups substituted by one or more aromatic, cycloaliphatic, or heterocyclic group(s).
  • the hydrophilic groups may comprise polar and/or charged groups and include carbohydrates, phosphate, carboxylic, sulfate, amino, sulfhydryl, nitro, hydroxyl, and other like groups.
  • an amphiphilic compound has a polar head attached to a long hydrophobic tail.
  • the polar portion is soluble in water, while the non-polar portion is insoluble in water.
  • the polar portion may have either a formal positive charge, or a formal negative charge.
  • the polar portion may have both a formal positive and a negative charge, and be a zwitterion or inner salt.
  • the amphiphilic compound can be, but is not limited to, one or a plurality of natural or non-natural lipids and lipid-like compounds.
  • a "lipid-like material” is a substance that is structurally and/or functionally related to a lipid but may not be considered a lipid in a strict sense.
  • the term includes compounds that are able to form amphiphilic layers as they are present in vesicles, multilamellar/unilamellar liposomes, or membranes in an aqueous environment and includes surfactants, or synthesized compounds with both hydrophilic and hydrophobic moieties.
  • the term refers to molecules, which comprise hydrophilic and hydrophobic moieties with different structural organization, which may or may not be similar to that of lipids.
  • amphiphilic compounds that may be included in an amphiphilic layer include, but are not limited to, phospholipids, aminolipids and sphingolipids.
  • lipids may be divided into eight categories: fatty acids, glycerolipids, glycerophospholipids, sphingolipids, saccharolipids, polyketides (derived from condensation of ketoacyl subunits), sterols and prenol lipids (derived from condensation of isoprene subunits).
  • lipid is sometimes used as a synonym for fats, fats are a subgroup of lipids called triglycerides.
  • Lipids also encompass molecules such as fatty acids and their derivatives (including tri-, di-, monoglycerides, and phospholipids), as well as sterol- containing metabolites such as cholesterol.
  • Fatty acids are a diverse group of molecules made of a hydrocarbon chain that terminates with a carboxylic acid group; this arrangement confers the molecule with a polar, hydrophilic end, and a nonpolar, hydrophobic end that is insoluble in water.
  • the carbon chain typically between four and 24 carbons long, may be saturated or unsaturated, and may be attached to functional groups containing oxygen, halogens, nitrogen, and sulfur.
  • a fatty acid contains a double bond, there is the possibility of either a cis or trans geometric isomerism, which significantly affects the molecule's configuration. Cis-double bonds cause the fatty acid chain to bend, an effect that is compounded with more double bonds in the chain.
  • Other major lipid classes in the fatty acid category are the fatty esters and fatty amides.
  • Glycerolipids are composed of mono-, di-, and tri-substituted glycerols, the best-known being the fatty acid triesters of glycerol, called triglycerides. The word "triacylglycerol" is sometimes used synonymously with "triglyceride”.
  • glycosylglycerols which are characterized by the presence of one or more sugar residues attached to glycerol via a glycosidic linkage.
  • Glycerophospholipids are amphipathic molecules (containing both hydrophobic and hydrophilic regions) that contain a glycerol core linked to two fatty acid-derived "tails" by ester linkages and to one "head” group by a phosphate ester linkage.
  • glycerophospholipids usually referred to as phospholipids (though sphingomyelins are also classified as phospholipids) are phosphatidylcholine (also known as PC, GPCho or lecithin), phosphatidylethanolamine (PE or GPEtn) and phosphatidylserine (PS or GPSer).
  • Sphingolipids are members of a complex family of compounds that share a common structural feature, a sphingoid base backbone. The major sphingoid base in mammals is commonly referred to as sphingosine.
  • Ceramides are a major subclass of sphingoid base derivatives with an amide-linked fatty acid.
  • the fatty acids are typically saturated or mono-unsaturated with chain lengths from 16 to 26 carbon atoms.
  • the major phosphosphingolipids of mammals are sphingomyelins (ceramide phosphocholines), whereas insects contain mainly ceramide phosphoethanolamines and fungi have phytoceramide phosphoinositols and mannose-containing headgroups.
  • the glycosphingolipids are a diverse family of molecules composed of one or more sugar residues linked via a glycosidic bond to the sphingoid base.
  • glycosphingolipids such as cerebrosides and gangliosides.
  • Sterols such as cholesterol and its derivatives, or tocopherol and its derivatives, are important components of membrane lipids, along with the glycerophospholipids and sphingomyelins.
  • Saccharolipids are compounds in which fatty acids are linked directly to a sugar backbone, forming structures that are compatible with membrane bilayers. In the saccharolipids, a monosaccharide substitutes for the glycerol backbone present in glycerolipids and glycerophospholipids.
  • the most familiar saccharolipids are the acylated glucosamine precursors of the Lipid A component of the lipopolysaccharides in Gram-negative bacteria.
  • Typical lipid A molecules are disaccharides of glucosamine, which are derivatized with as many as seven fatty-acyl chains.
  • the minimal lipopolysaccharide required for growth in E. coli is Kdo2-Lipid A, a hexa-acylated disaccharide of glucosamine that is glycosylated with two 3-deoxy-D-manno-octulosonic acid (Kdo) residues.
  • Polyketides are synthesized by polymerization of acetyl and propionyl subunits by classic enzymes as well as iterative and multimodular enzymes that share mechanistic features with the fatty acid synthases. They comprise a large number of secondary metabolites and natural products from animal, plant, bacterial, fungal and marine sources, and have great structural diversity. Many polyketides are cyclic molecules whose backbones are often further modified by glycosylation, methylation, hydroxylation, oxidation, or other processes. [0669] Lipids and lipid-like materials may be cationic, anionic or neutral. Neutral lipids or lipid-like materials exist in an uncharged or neutral zwitterionic form at a selected pH.
  • suitable lipids or lipid-like materials for use in the present disclosure include those described in WO2020/128031 and US20200163878, the entire contents of each of which are incorporated herein by reference for the purposes described herein. 2.
  • Cationic or cationically ionizable lipids or lipid-like materials [0671] In some embodiments cationic or cationically ionizable lipids or lipid-like materials contemplated for use herein include any cationic or cationically ionizable lipids or lipid-like materials which are able to electrostatically bind nucleic acid.
  • cationic or cationically ionizable lipids or lipid-like materials contemplated for use herein can be associated with nucleic acid, e.g., by forming complexes with the nucleic acid or forming vesicles in which the nucleic acid is enclosed or encapsulated.
  • Cationic lipids or lipid-like materials are characterized in that they have a net positive charge (e.g., at a relevant pH). Cationic lipids or lipid-like materials bind negatively charged nucleic acid by electrostatic interaction.
  • cationic lipids possess a lipophilic moiety, such as a sterol, an acyl chain, a diacyl or more acyl chains, and the head group of the lipid typically carries the positive charge.
  • a cationic lipid or lipid-like material has a net positive charge only at certain pH, in particular acidic pH, while it has preferably no net positive charge, preferably has no charge, i.e., it is neutral, at a different, preferably higher pH such as physiological pH. This ionizable behavior is thought to enhance efficacy through helping with endosomal escape and reducing toxicity as compared with particles that remain cationic at physiological pH.
  • a cationic or cationically ionizable lipid or lipid-like material comprises a head group which includes at least one nitrogen atom (N) which is positive charged or capable of being protonated.
  • cationic lipids include, but are not limited to 1,2-dioleoyl-3-trimethylammonium propane (DOTAP); N,N-dimethyl-2,3-dioleyloxypropylamine (DODMA), 1,2-di-O-octadecenyl-3-trimethylammonium propane (DOTMA), 3-(N—(N′,N′-dimethylaminoethane)-carbamoyl)cholesterol (DC-Chol), dimethyldioctadecylammonium (DDAB); 1,2-dioleoyl-3-dimethylammonium-propane (DODAP); 1,2-diacyloxy-3- dimethylammonium propanes; 1,
  • Suitable cationic lipids for use in the present disclosure include those described in WO2020/128031 and US20200163878, the entire contents of each of which are incorporated herein by reference for the purposes described herein.
  • Further suitable cationic lipids for use in the present disclosure include those described in WO2010/053572 (including Cl 2-200 described at paragraph [00225]) and WO2012/170930, both of which are incorporated herein by reference for the purposes described herein.
  • Additional suitable cationic lipids for use in the present disclosure include HGT4003, HGT5000, HGTS001, HGT5001, HGT5002 (see US20150140070A1, which is incorporated herein by reference in its entirety).
  • formulations that are useful for pharmaceutical compositions can comprise at least one cationic lipid.
  • Representative cationic lipids include, but are not limited to, 1 ,2-dilinoleyoxy-3-(dimethylamino)acetoxypropane (DLin-DAC), 1 ,2-dilinoleyoxy-3morpholinopropane (DLin-MA), 1,2-dilinoleoyl-3-dimethylaminopropane (DLinDAP), 1 ,2-dilinoleylthio-3-dimethylaminopropane (DLin-S-DMA), 1 -linoleoyl-2-linoleyloxy-3dimethylaminopropane (DLin-2- DMAP), 1 ,2-dilinoleyloxy-3-trimethylaminopropane chloride salt (DLin-TMA.CI), 1 ,2-dilinoleyloxy-3-trimethylaminopropane chloride salt (DLin-TMA.CI), 1
  • amino or cationic lipids useful in accordance with the present disclosure have at least one protonatable or deprotonatable group, such that the lipid is positively charged at a pH at or below physiological pH (e.g. pH 7.4), and neutral at a second pH, preferably at or above physiological pH.
  • physiological pH e.g. pH 7.4
  • second pH preferably at or above physiological pH.
  • a protonatable lipid has a pKa of the protonatable group in the range of about 4 to about 11, e.g., a pKa of about 5 to about 7.
  • a cationic lipid may comprise from about 10 mol % to about 100 mol %, about 20 mol % to about 100 mol %, about 30 mol % to about 100 mol %, about 40 mol % to about 100 mol %, or about 50 mol % to about 100 mol % of total lipid present in a lipid composition utilized in accordance with the present disclosure. 3.
  • formulations utilized in accordance with the present disclosure may comprise lipids or lipid-like materials other than cationic or cationically ionizable lipids or lipid-like materials, i.e., non- cationic lipids or lipid-like materials (including non-cationically ionizable lipids or lipid-like materials).
  • anionic and neutral lipids or lipid-like materials are referred to herein as non-cationic lipids or lipid-like materials.
  • optimizing a formulation of nucleic acid particles by addition of other hydrophobic moieties, such as cholesterol and lipids, in addition to an ionizable/cationic lipid or lipid-like material may, for example, enhance particle stability and efficacy of nucleic acid delivery.
  • a lipid or lipid-like material may be incorporated which may or may not affect the overall charge of particles.
  • such lipid or lipid-like material is a non-cationic lipid or lipid-like material.
  • a non-cationic lipid may comprise, e.g., one or more anionic lipids and/or neutral lipids.
  • a formulation comprises one of the following neutral lipid components: (1) a phospholipid, (2) cholesterol or a derivative thereof; or (3) a mixture of a phospholipid and cholesterol or a derivative thereof.
  • cholesterol derivatives include, but are not limited to, cholestanol, cholestanone, cholestenone, coprostanol, cholesteryl-2'-hydroxyethyl ether, cholesteryl-4'- hydroxybutyl ether, tocopherol and derivatives thereof, and mixtures thereof.
  • Specific example phospholipids that can be used include, but are not limited to, phosphatidylcholines, phosphatidylethanolamines, phosphatidylglycerols, phosphatidic acids, phosphatidylserines or sphingomyelin.
  • Such phospholipids include in particular diacylphosphatidylcholines, such as distearoylphosphatidylcholine (DSPC), dioleoylphosphatidylcholine (DOPC), dimyristoylphosphatidylcholine (DMPC), dipentadecanoylphosphatidylcholine, dilauroylphosphatidylcholine, dipalmitoylphosphatidylcholine (DPPC), diarachidoylphosphatidylcholine (DAPC), dibehenoylphosphatidylcholine (DBPC), ditricosanoylphosphatidylcholine (DTPC), dilignoceroylphatidylcholine (DLPC), palmitoyloleoyl-phosphatidylcholine (POPC), 1,2-di-O-octadecenyl-sn- glycero-3-phosphocholine (18:0 Diether PC), 1-oleoyl
  • a formulation utilized in accordance with the present disclosure includes DSPC or DSPC and cholesterol.
  • formulations utilized in accordance with the present disclosure include both a cationic lipid and an additional (non-cationic) lipid.
  • formulations herein include a polymer conjugated lipid such as a pegylated lipid. "Pegylated lipids" comprise both a lipid portion and a polyethylene glycol portion. Pegylated lipids are known in the art.
  • the amount of (total) cationic lipid compared to the amount of other lipid(s) in formulation may affect important characteristics, such as charge, particle size, stability, tissue selectivity, and bioactivity of the nucleic acid.
  • the molar ratio of the at least one cationic lipid to the at least one additional lipid is from about 10:0 to about 1:9, about 4:1 to about 1:2, or about 3:1 to about 1:1.
  • a non-cationic lipid in particular a neutral lipid, (e.g., one or more phospholipids and/or cholesterol) may comprise from about 0 mol % to about 90 mol %, from about 0 mol % to about 80 mol %, from about 0 mol % to about 70 mol %, from about 0 mol % to about 60 mol %, or from about 0 mol % to about 50 mol %, of the total lipid present in a formulation. 4.
  • Lipoplex Particles In certain embodiments of the present disclosure, the RNA described herein may be present in RNA lipoplex particles.
  • RNA lipoplex particle contains lipid, in particular cationic lipid, and RNA. Electrostatic interactions between positively charged liposomes and negatively charged RNA results in complexation and spontaneous formation of RNA lipoplex particles. Positively charged liposomes may be generally synthesized using a cationic lipid, such as DOTMA, and additional lipids, such as DOPE. In one embodiment, a RNA lipoplex particle is a nanoparticle. [0692] In certain embodiments, RNA lipoplex particles include both a cationic lipid and an additional lipid. In some embodiments, a cationic lipid is DOTMA and the additional lipid is DOPE.
  • the molar ratio of the at least one cationic lipid to the at least one additional lipid is from about 10:0 to about 1:9, about 4:1 to about 1:2, or about 3:1 to about 1:1. In specific embodiments, the molar ratio may be about 3:1, about 2.75:1, about 2.5:1, about 2.25:1, about 2:1, about 1.75:1, about 1.5:1, about 1.25:1, or about 1:1. In some embodiments, the molar ratio of the at least one cationic lipid to the at least one additional lipid is about 2:1.
  • RNA lipoplex particles have an average diameter that in one embodiment ranges from about 200 nm to about 1000 nm, from about 200 nm to about 800 nm, from about 250 to about 700 nm, from about 400 to about 600 nm, from about 300 nm to about 500 nm, or from about 350 nm to about 400 nm.
  • the RNA lipoplex particles have an average diameter of about 200 nm, about 225 nm, about 250 nm, about 275 nm, about 300 nm, about 325 nm, about 350 nm, about 375 nm, about 400 nm, about 425 nm, about 450 nm, about 475 nm, about 500 nm, about 525 nm, about 550 nm, about 575 nm, about 600 nm, about 625 nm, about 650 nm, about 700 nm, about 725 nm, about 750 nm, about 775 nm, about 800 nm, about 825 nm, about 850 nm, about 875 nm, about 900 nm, about 925 nm, about 950 nm, about 975 nm, or about 1000 nm.
  • RNA lipoplex particles have an average diameter that ranges from about 250 nm to about 700 nm. In another embodiment, the RNA lipoplex particles have an average diameter that ranges from about 300 nm to about 500 nm. In some embodiments, RNA lipoplex particles have an average diameter of about 400 nm.
  • RNA lipoplex particles and compositions comprising RNA lipoplex particles described herein are useful for delivery of RNA to a target tissue after parenteral administration, in particular after intravenous administration.
  • the RNA lipoplex particles may be prepared using liposomes that may be obtained by injecting a solution of the lipids in ethanol into water or a suitable aqueous phase.
  • the aqueous phase has an acidic pH. In one embodiment, the aqueous phase comprises acetic acid, e.g., in an amount of about 5 mM.
  • Liposomes may be used for preparing RNA lipoplex particles by mixing the liposomes with RNA. In one embodiment, the liposomes and RNA lipoplex particles comprise at least one cationic lipid and at least one additional lipid. In one embodiment, the at least one cationic lipid comprises 1,2-di-O-octadecenyl-3-trimethylammonium propane (DOTMA) and/or 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP).
  • DOTMA 1,2-di-O-octadecenyl-3-trimethylammonium propane
  • DOTAP 1,2-dioleoyl-3-trimethylammonium-propane
  • the at least one additional lipid comprises 1,2-di-(9Z-octadecenoyl)-sn-glycero-3-phosphoethanolamine (DOPE), cholesterol (Chol) and/or 1,2- dioleoyl-sn-glycero-3-phosphocholine (DOPC).
  • the at least one cationic lipid comprises 1,2-di-O- octadecenyl-3-trimethylammonium propane (DOTMA) and the at least one additional lipid comprises 1,2-di-(9Z- octadecenoyl)-sn-glycero-3-phosphoethanolamine (DOPE).
  • DOPE 1,2-di-(9Z- octadecenoyl)-sn-glycero-3-phosphoethanolamine
  • the liposomes and RNA lipoplex particles comprise 1,2-di-O-octadecenyl-3-trimethylammonium propane (DOTMA) and 1,2-di-(9Z-octadecenoyl)-sn- glycero-3-phosphoethanolamine (DOPE).
  • DOTMA 1,2-di-O-octadecenyl-3-trimethylammonium propane
  • DOPE 1,2-di-(9Z-octadecenoyl)-sn- glycero-3-phosphoethanolamine
  • Spleen targeting RNA lipoplex particles are described in WO 2013/143683, herein incorporated by reference. It has been found that RNA lipoplex particles having a net negative charge may be used to preferentially target spleen tissue or spleen cells such as antigen-presenting cells, in particular dendritic cells.
  • RNA lipoplex particles of the disclosure may be used for expressing RNA in the spleen.
  • no or essentially no RNA accumulation and/or RNA expression in the lung and/or liver occurs.
  • RNA accumulation and/or RNA expression in antigen presenting cells such as professional antigen presenting cells in the spleen occurs.
  • RNA lipoplex particles of the disclosure may be used for expressing RNA in such antigen presenting cells.
  • the antigen presenting cells are dendritic cells and/or macrophages.
  • LNPs Lipid Nanoparticles
  • nucleic acid such as RNA described herein is administered in the form of lipid nanoparticles (LNPs).
  • LNPs may comprise any lipid capable of forming a particle to which the one or more nucleic acid molecules are attached, or in which the one or more nucleic acid molecules are encapsulated.
  • an LNP comprises one or more cationic lipids, and one or more stabilizing lipids. Stabilizing lipids include neutral lipids and pegylated lipids.
  • an LNP comprises a cationic lipid, a neutral lipid, a sterol, a polymer conjugated lipid; and an RNA, encapsulated within or associated with the lipid nanoparticle.
  • a neutral lipid is selected from the group consisting of DSPC, DPPC, DMPC, DOPC, POPC, DOPE, DOPG, DPPG, POPE, DPPE, DMPE, DSPE, and SM.
  • the neutral lipid is selected from the group consisting of DSPC, DPPC, DMPC, DOPC, POPC, DOPE and SM.
  • the neutral lipid is DSPC.
  • a sterol is cholesterol.
  • a polymer conjugated lipid is a pegylated lipid.
  • a pegylated lipid has the following structure: or a pharmaceutically acceptable salt, tautomer or stereoisomer thereof, wherein: R 12 and R 13 are each independently a straight or branched, saturated or unsaturated alkyl chain containing from 10 to 30 carbon atoms, wherein the alkyl chain is optionally interrupted by one or more ester bonds; and w has a mean value ranging from 30 to 60.
  • R 12 and R 13 are each independently straight, saturated alkyl chains containing from 12 to 16 carbon atoms.
  • w has a mean value ranging from 40 to 55. In some embodiments, the average w is about 45. In some embodiments, R 12 and R 13 are each independently a straight, saturated alkyl chain containing about 14 carbon atoms, and w has a mean value of about 45.
  • the lipid has one of the following structures (IIIA) or (IIIB): ( ) ( ) wherein: A is a 3 to 8-membered cycloalkyl or cycloalkylene ring; R 6 is, at each occurrence, independently H, OH or C 1 -C 24 alkyl; and n is an integer ranging from 1 to 15. [0706] In some of the foregoing embodiments of Formula (III), the lipid has structure (IIIA), and in other embodiments, the lipid has structure (IIIB).
  • the lipid has one of the following structures (IIIC) or (IIID): wherein y and z are each independently integers ranging from 1 to 12.
  • the lipid has one of the following structures (IIIE) or (IIIF): .
  • the lipid has one of the following structures (IIIG), (IIIH), (IIII), or (IIIJ): ; .
  • n is an integer ranging from 2 to 12, for example from 2 to 8 or from 2 to 4.
  • n is 3, 4, 5 or 6.
  • n is 3.
  • n is 4.
  • n is 5.
  • n is 6.
  • y and z are each independently an integer ranging from 2 to 10. For example, in some embodiments, y and z are each independently an integer ranging from 4 to 9 or from 4 to 6.
  • R 6 is H. In other of the foregoing embodiments, R 6 is C 1 -C 24 alkyl. In other embodiments, R 6 is OH.
  • G 3 is unsubstituted. In other embodiments, G3 is substituted. In various different embodiments, G 3 is linear C 1 -C 24 alkylene or linear C 1 -C 24 alkenylene.
  • R 1 or R 2 is C 6 -C 24 alkenyl.
  • R 1 and R 2 each, independently have the following structure: , wherein: R 7a and R 7b are, at each occurrence, independently H or C 1 -C 12 alkyl; and a is an integer from 2 to 12, and wherein R 7a , R 7b and a are each selected such that R 1 and R 2 each independently comprise from 6 to 20 carbon atoms.
  • a is an integer ranging from 5 to 9 or from 8 to 12.
  • at least one occurrence of R 7a is H.
  • R 7a is H at each occurrence.
  • at least one occurrence of R 7b is C 1 -C 8 alkyl.
  • C 1 -C 8 alkyl is methyl, ethyl, n-propyl, iso-propyl, n-butyl, iso-butyl, tert-butyl, n-hexyl or n-octyl.
  • R 4 is methyl or ethyl.
  • the cationic lipid of Formula (III) has one of the structures set forth in in Table 21 below.
  • Table 21 Example Compounds of Formula (III). No. Structure No. Structure No. Structure No. Structure No. Structure No. Structure
  • a cationic lipid has one of the structures set forth in Table 22 below.
  • Table 22 Example Cationic Lipid Structures No. Structure No. Structure H O O N [0721] In so e e o e s, a co p ses a cao c p a s a o za e lipid-like material (lipidoid).
  • a cationic lipid has the following structure: [0722] , p p v v g .g., ameter) of about 30 nm to about 150 nm, about 40 nm to about 150 nm, about 50 nm to about 150 nm, about 60 nm to about 130 nm, about 70 nm to about 110 nm, about 70 nm to about 100 nm, about 70 to about 90 nm, or about 70 nm to about 80 nm.
  • lipid nanoparticles in accordance with the present disclosure can have an average size (e.g., mean diameter) of about 50 nm to about 100 nm.
  • lipid nanoparticles may have an average size (e.g., mean diameter) of about 50 nm to about 150 nm. In some embodiments, lipid nanoparticles may have an average size (e.g., mean diameter) of about 60 nm to about 120 nm.
  • lipid nanoparticles in accordance with the present disclosure can have an average size (e.g., mean diameter) of about 30 nm, 35 nm, 40 nm, 45 nm, 50 nm, 55 nm, 60 nm, 65 nm, 70 nm, 75 nm, 80 nm, 85 nm, 90 nm, 95 nm, 100 nm, 105 nm, 110 nm, 115 nm, 120 nm, 125 nm, 130 nm, 135 nm, 140 nm, 145 nm, or 150 nm.
  • average size e.g., mean diameter
  • average diameter refers to the mean hydrodynamic diameter of particles as measured by dynamic laser light scattering (DLS) with data analysis using the so-called cumulant algorithm, which provides as results the so-called Z-average with the dimension of a length, and the polydispersity index (PI), which is dimensionless (Koppel, D., J. Chem. Phys.57, 1972, pp 4814-4820, ISO 13321, which is herein incorporated by reference).
  • average diameter “mean diameter,” “diameter,” or “size” for particles is used synonymously with this value of the Z-average.
  • lipid nanoparticles described herein may exhibit a polydispersity index less than about 0.5, less than about 0.4, less than about 0.3, or about 0.2 or less.
  • lipid nanoparticles can exhibit a polydispersity index in a range of about 0.1 to about 0.3 or about 0.2 to about 0.3.
  • the “polydispersity index” is preferably calculated based on dynamic light scattering measurements by the so-called cumulant analysis as mentioned in the definition of the “average diameter.” Under certain prerequisites, it can be taken as a measure of the size distribution of an ensemble of ribonucleic acid nanoparticles (e.g., ribonucleic acid nanoparticles).
  • Lipid nanoparticles described herein can be characterized by an “N/P ratio,” which is the molar ratio of cationic (nitrogen) groups (the “N” in N/P) in the cationic polymer to the anionic (phosphate) groups (the “P” in N/P) in RNA.
  • N/P ratio is the molar ratio of cationic (nitrogen) groups (the “N” in N/P) in the cationic polymer to the anionic (phosphate) groups (the “P” in N/P) in RNA.
  • N/P ratio is the molar ratio of cationic (nitrogen) groups (the “N” in N/P) in the cationic polymer to the anionic (phosphate) groups (the “P” in N/P) in RNA.
  • N + cationic form
  • Use of a single number in an N/P ratio e.g., an N/P ratio of about 5 is intended to refer to that number over 1, e.g., an N/P
  • a lipid nanoparticle described herein has an N/P ratio greater than or equal to 5. In some embodiments, a lipid nanoparticle described herein has an N/P ratio that is about 5, 6, 7, 8, 9, or 10. In some embodiments, an N/P ratio for a lipid nanoparticle described herein is from about 10 to about 50. In some embodiments, an N/P ratio for a lipid nanoparticle described herein is from about 10 to about 70. In some embodiments, an N/P ratio for a lipid nanoparticle described herein is from about 10 to about 120. B.
  • Lipids and lipid nanoparticles comprising nucleic acids and their method of preparation are known in the art, including, e.g., as described in U.S. Patent Nos. 8,569,256, 5,965,542 and U.S. Patent Publication Nos.
  • cationic lipids, neutral lipids (e.g., DSPC, and/or cholesterol) and polymer-conjugated lipids can be solubilized in ethanol at a pre-determined molar ratio (e.g., ones described herein).
  • lipid nanoparticles are prepared at a total lipid to polyribonucleotides weight ratio of approximately 10: 1 to 30: 1. In some embodiments, such polyribonucleotides can be diluted to 0.2 mg/mL in acetate buffer.
  • a colloidal lipid dispersion comprising polyribonucleotides can be formed as follows: an ethanol solution comprising lipids, such as cationic lipids, neutral lipids, and polymer-conjugated lipids, is injected into an aqueous solution comprising polyribonucleotides (e.g., ones described herein).
  • lipid and polyribonucleotide solutions can be mixed at room temperature by pumping each solution at controlled flow rates into a mixing unit, for example, using piston pumps.
  • the flow rates of a lipid solution and a RNA solution into a mixing unit are maintained at a ratio of 1:3.
  • RNA-encapsulated lipid nanoparticles can be processed by one or more of concentration adjustment, buffer exchange, formulation, and/or filtration.
  • RNA-encapsulated lipid nanoparticles can be processed through filtration.
  • compositions e.g., pharmaceutical compositions comprising one or more polyribonucleotides described herein.
  • an excipient is approved for use in humans and for veterinary use. In some embodiments, an excipient is approved by the United States Food and Drug Administration. In some embodiments, an excipient is pharmaceutical grade. In some embodiments, an excipient meets the standards of the United States Pharmacopoeia (USP), the European Pharmacopoeia (EP), the British Pharmacopoeia, and/or the International Pharmacopoeia.
  • USP United States Pharmacopoeia
  • EP European Pharmacopoeia
  • British Pharmacopoeia the British Pharmacopoeia
  • International Pharmacopoeia International Pharmacopoeia
  • compositions include, but are not limited to, inert diluents, dispersing and/or granulating agents, surface active agents and/or emulsifiers, disintegrating agents, binding agents, preservatives, buffering agents, lubricating agents, and/or oils. Such excipients may optionally be included in pharmaceutical formulations. Excipients such as cocoa butter and suppository waxes, coloring agents, coating agents, sweetening, flavoring, and/or perfuming agents can be present in the composition, according to the judgment of the formulator.
  • compositions provided herein may be formulated with one or more pharmaceutically acceptable carriers or diluents as well as any other known adjuvants and excipients in accordance with conventional techniques such as those disclosed in Remington: The Science and Practice of Pharmacy 21st ed., Lippincott Williams & Wilkins, 2005 (incorporated herein by reference).
  • Pharmaceutical compositions described herein can be administered by appropriate methods known in the art.
  • compositions described herein are formulated for parenteral administration, which includes modes of administration other than enteral and topical administration, usually by injection, and includes, without limitation, intravenous, intramuscular, intraarterial, intradermal, subcutaneous, subcuticular, or intraarticular injection and infusion.
  • parenteral administration which includes modes of administration other than enteral and topical administration, usually by injection, and includes, without limitation, intravenous, intramuscular, intraarterial, intradermal, subcutaneous, subcuticular, or intraarticular injection and infusion.
  • pharmaceutical compositions described herein are formulated for intravenous, intramuscular, or subcutaneous administration.
  • compositions described herein are formulated for intramuscular administration.
  • pharmaceutical compositions described herein are formulated for intravenous administration.
  • pharmaceutically acceptable excipients that may be useful for intravenous administration include sterile aqueous solutions or dispersions and sterile powders for preparation of sterile injectable solutions or dispersions.
  • Therapeutic compositions typically must be sterile and stable under the conditions of manufacture and storage. The composition can be formulated as a solution, microemulsion, lipid nanoparticles, or other ordered structure suitable to high drug concentration.
  • the carrier can be a solvent or dispersion medium containing, for example, water, ethanol, polyol (for example, glycerol, propylene glycol, and liquid polyethylene glycol, and the like), and suitable mixtures thereof. Proper fluidity can be maintained, for example, by the use of surfactants. In many cases, it will be preferable to include isotonic agents, for example, sugars, polyalcohols such as mannitol, sorbitol, or sodium chloride in the composition. In some embodiments, prolonged absorption of the injectable compositions can be brought about by including in the composition an agent that delays absorption, for example, monostearate salts and gelatin.
  • an agent that delays absorption for example, monostearate salts and gelatin.
  • Sterile injectable solutions can be prepared by incorporating the active compound in the required amount in an appropriate solvent with one or a combination of ingredients enumerated above, as required, followed by sterilization and/or microfiltration.
  • pharmaceutical compositions can be prepared as described herein and/or methods known in the art.
  • a pharmaceutical composition includes ALC-0315; ALC-0159; DSPC; Cholesterol; Sucrose; NaCl; KCl; Na 2 HPO 4 ; KH 2 PO 4 ; Water for injection.
  • normal saline is used as diluent.
  • compositions may also contain adjuvants such as preservatives, wetting agents, emulsifying agents and dispersing agents. Prevention of the presence of microorganisms may be ensured both by sterilization procedures, and by the inclusion of various antibacterial and antifungal agents, for example, paraben, chlorobutanol, phenol sorbic acid, and the like. It may also be desirable to include isotonic agents, such as sugars, sodium chloride, and the like into pharmaceutical compositions described herein. In addition, prolonged absorption of the injectable pharmaceutical form may be brought about by the inclusion of agents which delay absorption such as aluminum monostearate and gelatin.
  • adjuvants such as preservatives, wetting agents, emulsifying agents and dispersing agents. Prevention of the presence of microorganisms may be ensured both by sterilization procedures, and by the inclusion of various antibacterial and antifungal agents, for example, paraben, chlorobutanol, phenol sorbic acid, and the like. It may also be desirable to
  • Formulations of pharmaceutical compositions described herein may be prepared by any method known or hereafter developed in the art of pharmacology. In general, such preparatory methods include the step of bringing active ingredient(s) into association with a diluent or another excipient and/or one or more other accessory ingredients, and then, if necessary and/or desirable, shaping and/or packaging the product into a desired single- or multi-dose unit.
  • a pharmaceutical composition in accordance with the present disclosure may be prepared, packaged, and/or sold in bulk, as a single unit dose, and/or as a plurality of single unit doses.
  • a “unit dose” is discrete amount of the pharmaceutical composition comprising a predetermined amount of at least one RNA product produced using a system and/or method described herein.
  • Relative amounts of polyribonucleotides encapsulated in lipid nanoparticles, a pharmaceutically acceptable excipient, and/or any additional ingredients in a pharmaceutical composition can vary, depending upon the subject to be treated, target cells, diseases or disorders, and may also further depend upon the route by which the composition is to be administered.
  • pharmaceutical compositions described herein are formulated into pharmaceutically acceptable dosage forms by conventional methods known to those of skill in the art.
  • compositions described herein may be varied so as to obtain an amount of the active ingredient which is effective to achieve the desired therapeutic response for a particular patient, composition, and mode of administration, without being toxic to the patient.
  • active ingredients e.g., polyribonucleotides encapsulated in lipid nanoparticles
  • dosage levels of the active ingredients may be varied so as to obtain an amount of the active ingredient which is effective to achieve the desired therapeutic response for a particular patient, composition, and mode of administration, without being toxic to the patient.
  • the selected dosage level will depend upon a variety of pharmacokinetic factors including the activity of the particular compositions of the present disclosure employed, the route of administration, the time of administration, the rate of excretion of the particular compound being employed, the duration of the treatment, other drugs, compounds and/or materials used in combination with the particular compositions employed, the age, sex, weight, condition, general health and prior medical history of the patient being treated, and like factors well known in the medical arts. [0747] A physician having ordinary skill in the art can readily determine and prescribe the effective amount of the pharmaceutical composition required.
  • a physician could start doses of active ingredients (e.g., polyribonucleotides encapsulated in lipid nanoparticles) employed in the pharmaceutical composition at levels lower than that required in order to achieve the desired therapeutic effect and gradually increase the dosage until the desired effect is achieved.
  • active ingredients e.g., polyribonucleotides encapsulated in lipid nanoparticles
  • a pharmaceutical composition is formulated (e.g., but not limited to, for intravenous, intramuscular, or subcutaneous administration) to deliver a dose of about 5 mg RNA/kg.
  • a pharmaceutical composition described herein may further comprise one or more additives, for example, in some embodiments that may enhance stability of such a composition under certain conditions.
  • a pharmaceutical composition may further comprise a cryoprotectant (e.g., sucrose) and/or an aqueous buffered solution, which may in some embodiments include one or more salts, including, e.g., alkali metal salts or alkaline earth metal salts such as, e.g., sodium salts, potassium salts, and/or calcium salts.
  • a pharmaceutical composition provided herein is a preservative-free, sterile RNA-lipid nanoparticle dispersion in an aqueous buffer for intravenous or intramuscular administration.
  • compositions suitable for administration to humans are principally directed to pharmaceutical compositions that are suitable for administration to humans, it will be understood by the skilled artisan that such compositions are generally suitable for administration to animals of all sorts. Modification of pharmaceutical compositions suitable for administration to humans in order to render the compositions suitable for administration to various animals is well understood, and the ordinarily skilled veterinary pharmacologist can design and/or perform such modification with merely ordinary, if any, experimentation. A.
  • compositions comprising a plurality of polyribonucleotides, wherein the plurality of polyribonucleotides comprises a first set of polyribonucleotides that encode one or more glycoprotein (GP) polypeptides as described herein.
  • a combination comprises one polyribonucleotide that encodes a (GP) polypeptide.
  • a combination comprises two or more polyribonucleotides that encode one or more (GP) polypeptides.
  • a combination comprises three or more polyribonucleotides that encode three or more (GP) polypeptides.
  • a combination comprises four or more polyribonucleotides that encode four or more (GP) polypeptides. In some embodiments, a combination comprises three or more polyribonucleotides that encode three or more (GP) polypeptides and a polyribonucleotide that encodes a T-cell string polypeptide. [0753] Also provided herein are combinations that comprise two or more pharmaceutical compositions, wherein each pharmaceutical composition comprises one or more polyribonucleotides as described herein. Also provided herein are combinations that comprise three or more pharmaceutical compositions, wherein each pharmaceutical composition comprises one or more polyribonucleotides as described herein. [0754] Provided herein are combinations that comprise two or more RNA constructs as described herein.
  • a combination comprises three or more RNA constructs as described herein. In some embodiments, a combination comprises four or more RNA constructs as described herein. In some embodiments, a combination comprises five or more RNA constructs as described herein. [0755] Also provided herein are combinations that comprise two or more pharmaceutical compositions, wherein each pharmaceutical composition comprises one or more RNA constructs as described herein. Also provided herein are combinations that comprise three or more pharmaceutical compositions, wherein each pharmaceutical composition comprises one or more RNA constructs as described herein. Also provided herein are combinations that comprise four or more pharmaceutical compositions, wherein each pharmaceutical composition comprises one or more RNA constructs as described herein.
  • a combination comprises a plurality of polyribonucleotides comprising a first set of polyribonucleotides that comprises a polyribonucleotide encoding a GP polypeptide comprising an HSV-2 gC or antigenic portion thereof, and a polyribonucleotide encodes a GP polypeptide comprising an HSV-2 gD or antigenic portion thereof.
  • a combination comprises a plurality of polyribonucleotides comprising a first set of polyribonucleotides that comprises a polyribonucleotide encoding a GP polypeptide comprising an antigenic portion of HSV-2 gC, and a polyribonucleotide encoding a GP polypeptide comprising an antigenic portion of HSV-2 gD.
  • a GP polypeptide comprising an antigenic portion of HSV-2 gC comprises an amino acid sequence according to SEQ ID NO: 65.
  • a GP polypeptide comprising an antigenic portion of HSV-2 gD comprises an amino acid sequence according to SEQ ID NO: 70.
  • a combination comprises (i) a first pharmaceutical composition comprising a polyribonucleotide that encodes a GP polypeptide comprising an HSV-2 gC or antigenic portion thereof, and (ii) a second pharmaceutical composition comprising a polyribonucleotide that encodes a GP polypeptide comprising an HSV-2 gD antigen or antigenic portion thereof.
  • a combination comprises a plurality of polyribonucleotides comprising a first set of polyribonucleotides that comprises a polyribonucleotide encoding a GP polypeptide comprising an antigenic portion of HSV-2 gC, and a polyribonucleotide encoding a GP polypeptide comprising an antigenic portion of HSV-2 gE.
  • a GP polypeptide comprising an antigenic portion of HSV-2 gC comprises an amino acid sequence according to SEQ ID NO: 65.
  • a GP polypeptide comprising an antigenic portion of HSV-2 gE comprises an amino acid sequence according to SEQ ID NO: 73.
  • a combination comprises (i) a first pharmaceutical composition comprising a polyribonucleotide that encodes a GP polypeptide comprising an HSV-2 gC or antigenic portion thereof, and (ii) a second pharmaceutical composition comprising a polyribonucleotide that encodes a GP polypeptide comprising an HSV-2 gE antigen or antigenic portion thereof.
  • a combination comprises (i) a first pharmaceutical composition comprising a first polyribonucleotide that encodes a GP polypeptide comprising an antigenic portion of HSV-2 gC, and (ii) a second pharmaceutical composition comprising a polyribonucleotide that encodes a GP comprising an antigenic portion of HSV-2 gE.
  • a GP polypeptide comprising an antigenic portion of HSV-2 gC comprises an amino acid sequence according to SEQ ID NO: 65.
  • a GP polypeptide comprising an antigenic portion of HSV-2 gE comprises an amino acid sequence according to SEQ ID NO: 73.
  • a combination comprises a plurality of polyribonucleotides comprising a first set of polyribonucleotides that comprises a polyribonucleotide encoding a GP polypeptide comprising an HSV-2 gD or antigenic portion thereof, and a polyribonucleotide encodes a GP polypeptide comprising an HSV-2 gE or antigenic portion thereof.
  • a combination comprises a plurality of polyribonucleotides comprising a first set of polyribonucleotides that comprises a polyribonucleotide encoding a GP polypeptide comprising an antigenic portion of HSV-2 gD, and a polyribonucleotide encoding a GP polypeptide comprising an antigenic portion of HSV-2 gE.
  • a GP polypeptide comprising an antigenic portion of HSV-2 gD comprises an amino acid sequence according to SEQ ID NO: 70.
  • a GP polypeptide comprising an antigenic portion of HSV-2 gE comprises an amino acid sequence according to SEQ ID NO: 73.
  • a combination comprises (i) a first pharmaceutical composition comprising a polyribonucleotide that encodes a GP polypeptide comprising an HSV-2 gD or antigenic portion thereof, and (ii) a second pharmaceutical composition comprising a polyribonucleotide that encodes a GP polypeptide comprising an HSV-2 gE antigen or antigenic portion thereof.
  • a GP polypeptide that comprises an amino acid sequence according to SEQ ID NO: 65 is encoded by a ribonucleic acid sequence according to 106.
  • a GP polypeptide that comprises an amino acid sequence according to SEQ ID NO: 70 is encoded by a ribonucleic acid sequence according to 118.
  • a GP polypeptide that comprises an amino acid sequence according to SEQ ID NO: 73 is encoded by a ribonucleic acid sequence according to 123.
  • technologies of the present disclosure are used in the treatment and/or prophylactic of an HSV infection.
  • Prophylactic purposes of the present disclosure comprise pre-exposure prophylaxis and/or post-exposure prophylaxis.
  • technologies of the present disclosure are used in the treatment and/or prophylaxis of a disorder related to such an HSV (e.g., HSV-1 and/or HSV-2) infection.
  • a disordered related to such an HSV (e.g., HSV-1 and/or HSV-2) infection comprises, for example, a typical symptom and/or a complication of an HSV (e.g., HSV-1 and/or HSV-2) infection.
  • compositions may be useful to detect and/or characterize one or more features of an anti-HSV (e.g., anti-HSV-1 and/or anti-HSV-2) immune response (e.g., by detecting binding to a provided antigen by serum from an infected subject).
  • an anti-HSV e.g., anti-HSV-1 and/or anti-HSV-2
  • an immune response e.g., by detecting binding to a provided antigen by serum from an infected subject.
  • compositions e.g., that are or comprise one or more GP polypeptides and/or one or more T-cell string polypeptides
  • the present disclosure provides use of encoding nucleic acids (e.g., DNA or RNA) to produce encoded antigens and/or use of DNA constructs to produce RNA.
  • technologies of the present disclosure are utilized in a non-limited subject population; in some embodiments, technologies of the present disclosure are utilized in particular subject populations.
  • a subject population comprises an adult population.
  • an adult population comprises subjects between the ages of about 18 years and about 55 years of age (e.g., about 19, 20, 25, 30, 35, 40, 45, 50, 51, 52, 53, 54, or 55,).
  • a subject population comprises an elderly population.
  • an elderly population comprises subjects of, about 56 years of age, about 60 years of age, about 70 years of age, or older (e.g., about 60, 65, 70, 75, 80, 85, 90, 95, or 100 years of age).
  • a subject has a weight of at least about 50 kg.
  • a subject has a weight of at least about 51 kg (e.g., about 52, 53, 54, 55, 56,57, 58, 59, 60 kg).
  • a subject has a body mass index (BMI) in a range of about 17.5 kg/m 2 to about 37 kg/m 2 , such as about 18 kg/m 2 to about 36 kg/m 2 , such as about 18.5 kg/m 2 to about 35 kg/m 2 .
  • BMI body mass index
  • a subject has a BMI of at least 17 kg/m 2 , such as at least 17.5 kg/m 2 , such as at least 18 kg/m 2 , such as at least 18.5 kg/m 2 .
  • a subject has a BMI of at the most 40 kg/m 2 , such as at the most 39 kg/m 2 , such as at the most 38 kg/m 2 , such as at the most 37 kg/m 2 , such as at the most 36 kg/m 2 , such as at the most 35 kg/m 2 .
  • a subject population comprises a pediatric population. In some embodiments, a pediatric population comprises subjects approximately 18 years old or younger. In some such embodiments, a pediatric population comprises subjects between the ages of about 1 year and about 18 years (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, or 18 years of age).
  • a subject population comprises a newborn population.
  • a newborn population comprises subjects about 12 months or younger (e.g., 12, 11, 10, 9, 8, 7, 6, 5, 4, 3, 2, 1 months or younger).
  • subject populations to be treated with technologies described herein include infants (e.g., about 12 months or younger) whose mothers did not receive such technologies described herein during pregnancy.
  • subject populations to be treated with technologies described herein may include pregnant women; in some embodiments, infants whose mothers were treated with disclosed technologies during pregnancy (e.g., who received at least one dose, or alternatively only who received both doses), are not vaccinated during the first weeks, months, or even years (e.g., 1, 2, 3, 4, 5, 6, 7, 8 weeks or more, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24 months or more, or 1, 2, 3, 4, 5 years or more) post-birth.
  • infants whose mothers were treated with disclosed technologies during pregnancy e.g., who received at least one dose, or alternatively only who received both doses
  • are not vaccinated during the first weeks, months, or even years e.g., 1, 2, 3, 4, 5, 6, 7, 8 weeks or more, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24 months or more, or 1, 2, 3, 4, 5 years or more post-birth.
  • a subject population is or comprises children aged 6 weeks to up to 17 months of age.
  • a provided pharmaceutical composition e.g., immunogenic composition, e.g., vaccine
  • another pharmaceutical composition e.g., immunogenic composition, e.g., vaccine
  • therapeutic intervention e.g., to treat or prevent an HSV infection, or another disease, disorder, or condition.
  • a provided pharmaceutical composition may be administered with a protein vaccine, a DNA vaccine, an RNA vaccine, a cellular vaccine, a conjugate vaccine, etc.
  • one or more doses of a provided pharmaceutical composition may be administered together with (e.g., in a single visit) another composition (e.g., vaccine) or other therapy.
  • a provided pharmaceutical composition e.g., immunogenic composition, e.g., vaccine
  • HSV e.g., HSV-1 and/or HSV-2
  • a provided pharmaceutical composition e.g., immunogenic composition, e.g., vaccine
  • a subject has no prior history of known or suspected herpes simplex vaccination prior to administration of one or more doses of a composition as disclosed herein.
  • a subject does not have febrile illness prior to administration of one or more doses of a composition as disclosed herein. In some embodiments, a subject does not have febrile illness about 72 hours, about 48 hours, about 36 hours, about 24 hours, or about 12 hours prior to administration of one or more doses of a composition as disclosed herein. [0786] In some embodiments, a subject does not have an acute illness prior to administration of one or more doses of a composition as disclosed herein. In some embodiments, a subject does not have an acute illness about 72 hours, about 48 hours, about 36 hours, about 24 hours, or about 12 hours prior to administration of one or more doses of a composition as disclosed herein.
  • a subject has not received a vaccine 0 to 300 days, 0 to 290 days, 0 to 280 days, 0 to 270 days, 0 to 260 days, 0 to 250 days, 0 to 240 days, 0 to 230 days, 0 to 220 days, 0 to 210, 0 to 200 days, 0 to 190 days, 0 to 180 days, 0 to 170 days, 0 to 160 days, 0 to 150 days, 0 to 140 days, 0 to 130 days, 0 to 120 days, 0 to 110 days, 0 to 100 days, 0 to 90 days, 0 to 80 days, 0 to 70 days, 0 to 60 days, 0 to 50 days, 0 to 40 days, 0 to 35 days, 0 to 30 days, 0 to 29 days, 0 to 28 days before being administered one or more doses of a therapeutically effective amount of a composition as disclosed herein.
  • a subject has not received a vaccine about 7 days, about 14 days, about 21 days, about 28 days, about 35 days, about 42 days, about 49 days, about 56 days, about 60 days, about 70 days, about 80 days, about 90 days, about 100 days, about 125 days, about 150 days, about 175 days, about 190 days, about 200 days, about 210 days, or about 210 days before being administered one or more doses of a therapeutically effective amount of a composition as disclosed herein.
  • a vaccine is not a seasonal influenza vaccine or a medically indicated vaccine.
  • a subject does not receive a vaccine at least 2 weeks to 35 weeks, at least 3 weeks to 34 weeks, or at least 4 weeks to 33 weeks after administration of one or more doses of a therapeutically effective amount of a composition as disclosed herein.
  • a subject does not receive a vaccine at least about 2 weeks, at least about 4 weeks, at least about 6 weeks, at least about 8 weeks, at least about 10 weeks, at least about 12 weeks, at least about 14 weeks, at least about 16 weeks, at least about 18 weeks, at least about 20 weeks, at least about 22 weeks, at least about 24 weeks, at least about 26 weeks, at least about 28 weeks, or at least about 30 weeks after administration of one or more doses of a therapeutically effective amount of a composition as disclosed herein.
  • the vaccine is not a seasonal influenza vaccine or a medically indicated vaccine.
  • a subject has not received blood, plasma products, or immunoglobulins about 0 to 600 days, about 0 to 590 days, about 0 to 580 days, about 0 to 570 days, about 0 to 560 days, about 0 to 550 days, or about 0 to 545 days before administration of one or more doses of a therapeutically effective amount of a composition disclosed herein.
  • a subject has not received an allergy treatment 8 to 45 days, 12 to 40 days, 16 to 38 days, 21 to 35 days, 23 to 32 days, 25 to 30 days, or 26 to 29 days before administration of one or more doses of a therapeutically effective amount of a composition disclosed herein.
  • a subject has not received an allergy treatment about 14 days, about 16 days, about 18 days, about 20 days, about 22 days, about 24 days, about 26 days, about 28 days before administration of one or more doses of a therapeutically effective amount of a composition disclosed herein.
  • an allergy treatment comprises antigen injections.
  • a subject has not received an immunosuppressive medication 7 to 56 days, 14 to 56 days, 21 to 56 days, 28 to 56 days, 35 to 56 days, 42 to 56 days, 49 to 56 days, 7 to 49 days, 14 to 49 days, 21 to 49 days, 28 to 49 days, 35 to 49 days, 42 to 49 days, 7 to 42 days, 14 to 42 days, 21 to 42 days, 28 to 42 days, 35 to 42 days, 7 to 35 days, 14 to 35 days, 21 to 35 days, 28 to 35 days, 7 to 28 days, 14 to 28 days, 21 to 28 days, 7 to 21 days, 14 to 21 days, or 7 to 14 days before administration of one or more doses of a therapeutically effective amount of a composition disclosed herein.
  • a subject has not received an immunosuppressive medication about 7 days, about 14 days, about 21 days, about 28 days, about 35 days, about 42 days, about 49 days, or about 56 days before administration of one or more doses of a therapeutically effective amount of a composition disclosed herein.
  • a subject has not received an immunosuppressive medication about 28 days before administration of one or more doses of a therapeutically effective amount of a composition disclosed herein.
  • an immunosuppressive medication comprises a systemic corticosteroid or radiotherapy.
  • a systemic coriticosteroid is selected from, but not limited to, methylprednisolone, dexamethasone, hydrocortisone, prednisone, prednisolone, fluticasone, flumethasone, fluocinolone, budesonide, beclomethasone, ciclesonide, cortisone, triamcinolone, betamethasone, deflazacort, difluprednate, loteprednol, paramethasone, tixocortol, aldosterone, cloprednol, cortivazol, deoxycortone, desonide, desoximetasone, difluorocortolone, fluclorolone, fludrocortisone, flunisolide, fluocinonide, fluocortin butyl, fluorocortisone, fluorocortolone, fluorometholone, flurandrenolone, halcinon
  • a coriticosteroid is prednisone.
  • a subject has not received a prophylactic antipyretic and/or an analgesic medication 0 to 600 days, 0 to 550 days, 0 to 500 days, 0 to 500 days, 0 to 450 days, 0 to 400 days, 0 to 350 days, 0 to 300 days, 0 to 250 days, 0 to 200 days, 0 to 150 days, 0 to 150 days, 0 to 150 days, or 0 to 50 days before administration of one or more doses of a therapeutically effective amount of a composition disclosed herein.
  • a prophylactic antipyretic medication is selected from, but not limited to, acetaminophen, a non-steroidal anti-inflammatory drug (NSAID), salicylamide, salicyl salicylate, methyl salicylate, magnesium salicylate, fatelamine, ethenzamide, diflunisal, choline magnesium salicylate, benorylate/benorilatem and amoxiprin, acetylsalicylate, ceclofenac, acemetacin, alclofenac, bromfenac, diclofenac, etodolac, indomethacin, nabumetone, oxametacin, proglumetacin, sulindac, tolmetin, iminoprofen, benoxaprofen, carprofen, dexibuprofen, dexketoprofen, fenbufen, fenoprofen, flunoxaprofen,
  • NSAID non-ster
  • a prophylactic analgesic medication is selected from, but not limited to, acetaminophen, salicylamide, salicyl salicylate, methyl salicylate, magnesium salicylate, fatelamine, ethenzamide, diflunisal, choline magnesium salicylate, benorylate/benorilatem and amoxiprin, acetylsalicylate, ceclofenac, acemetacin, alclofenac, bromfenac, diclofenac, etodolac, indomethacin, nabumetone, oxametacin, proglumetacin, sulindac, tolmetin, iminoprofen, benoxaprofen, carprofen, dexibuprofen, dexketoprofen, fenbufen, fenoprofen, flunoxaprofen, flurbiprofen, ibuprof
  • technologies of the present disclosure may be administered to subjects according to a particular dosing regimen.
  • a dosing regimen may involve a single administration; in some embodiments, a dosing regimen may comprise one or more “booster” administrations after the initial administration.
  • initial and boost doses are the same amount; in some embodiments they differ.
  • two or more booster doses are administered.
  • a plurality of doses are administered at regular intervals. In some embodiments, periods of time between doses become longer.
  • one or more subsequent doses is administered if a particular clinical (e.g., reduction in neutralizing antibody levels) or situational (e.g., local development of a new strain) even arises or is detected.
  • administered pharmaceutical compositions e.g., immunogenic compositions, e.g., vaccines
  • RNA constructs that encode HSV-2 gC, gD and/or gE constructs are administered in RNA doses of from about 0.1 ⁇ g to about 300 ⁇ g, about 0.5 ⁇ g to about 200 ⁇ g, or about 1 ⁇ g to about 100 ⁇ g, such as about 1 ⁇ g, about 3 ⁇ g, about 10 ⁇ g, about 30 ⁇ g, about 50 ⁇ g, or about 100 ⁇ g.
  • a plurality of booster doses are administered within 6 months of the first dose, or within 12 months of the first dose.
  • 3 doses or fewer are required to achieve effective vaccination (e.g., greater than 60%, and in some embodiments greater than about 70%, about 75%, about 80%, about 85%, about 90% or more) reduction in risk of infection, or of serious disease.
  • not more than two doses are required.
  • a single dose is sufficient.
  • an RNA dose is about 60 ⁇ g or lower, 50 ⁇ g or lower, 40 ⁇ g or lower, 30 ⁇ g or lower, 20 ⁇ g or lower, 10 ⁇ g or lower, 5 ⁇ g or lower, 2.5 ⁇ g or lower, or 1 ⁇ g or lower. In some embodiments, an RNA dose is about 0.25 ⁇ g, at least 0.5 ⁇ g, at least 1 ⁇ g, at least 2 ⁇ g, at least 3 ⁇ g, at least 4 ⁇ g, at least 5 ⁇ g, at least 10 ⁇ g, at least 20 ⁇ g, at least 30 ⁇ g, or at least 40 ⁇ g.
  • an RNA dose is about 0.25 ⁇ g to 60 ⁇ g, 0.5 ⁇ g to 55 ⁇ g, 1 ⁇ g to 50 ⁇ g, 5 ⁇ g to 40 ⁇ g, or 10 ⁇ g to 30 ⁇ g may be administered per dose. In some embodiments, an RNA dose is about 30 ⁇ g. In some embodiments, at least two such doses are administered. For example, a second dose may be administered about 21 days following administration of the first dose. In some embodiments, a first booster dose is administered about one month after an initial dose. In some such embodiments, at least one further booster is administered at one-month interval(s).
  • polyribonucleotides can be produced by methods known in the art.
  • polyribonucleotides can be produced by in vitro transcription, for example, using a DNA template.
  • a plasmid DNA used as a template for in vitro transcription to generate a polyribonucleotide described herein is also within the scope of the present disclosure.
  • a DNA template is used for in vitro RNA synthesis in the presence of an appropriate RNA polymerase (e.g., a recombinant RNA-polymerase such as a T7 RNA-polymerase) with ribonucleotide triphosphates (e.g., ATP, CTP, GTP, UTP).
  • an appropriate RNA polymerase e.g., a recombinant RNA-polymerase such as a T7 RNA-polymerase
  • ribonucleotide triphosphates e.g., ATP, CTP, GTP, UTP.
  • polyribonucleotides e.g., ones described herein
  • pseudouridine ( ⁇ ), N1-methyl-pseudouridine (m1 ⁇ ), or 5-methyl-uridine (m5U) can be used to replace uridine triphosphate (UTP).
  • pseudouridine ( ⁇ ) can be used to replace uridine triphosphate (UTP).
  • N1-methyl-pseudouridine (m1 ⁇ ) can be used to replace uridine triphosphate (UTP).
  • 5-methyl-uridine (m5U) can be used to replace uridine triphosphate (UTP).
  • an RNA polymerase typically traverses at least a portion of a single-stranded DNA template in the 3' ⁇ 5' direction to produce a single-stranded complementary RNA in the 5' ⁇ 3' direction.
  • a polyribonucleotide comprises a polyA tail
  • a polyA tail may be encoded in a DNA template, e.g., by using an appropriately tailed PCR primer, or it can be added to a polyribonucleotide after in vitro transcription, e.g., by enzymatic treatment (e.g., using a poly(A) polymerase such as an E. coli Poly(A) polymerase).
  • enzymatic treatment e.g., using a poly(A) polymerase such as an E. coli Poly(A) polymerase.
  • Suitable poly(A) tails are described herein above.
  • a poly(A) tail comprises a nucleotide sequence of AAAAAAAAAAAAAAAAAAAAAAAAAAGCATATGACTAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAA (SEQ ID NO: 153).
  • a poly(A) tail comprises a plurality of A residues interrupted by a linker.
  • a linker comprises the nucleotide sequence GCATATGAC (SEQ ID NO: 154).
  • RNA e.g., mRNA
  • a 5' cap can also protect an RNA product from 5' exonuclease mediated degradation and thus increases half-life.
  • capping may be performed after in vitro transcription in the presence of a capping system (e.g., an enzyme-based capping system such as, e.g., capping enzymes of vaccinia virus).
  • a cap may be introduced during in vitro transcription, along with a plurality of ribonucleotide triphosphates such that a cap is incorporated into a polyribonucleotide during transcription (also known as co-transcriptional capping).
  • a GTP fed-batch procedure with multiple additions in the course of the reaction may be used to maintain a low concentration of GTP in order to effectively cap the RNA.
  • Suitable 5' cap are described herein above.
  • a 5' cap comprises m7(3'OMeG)(5')ppp(5')(2'OMeA)pG.
  • digestion can be achieved with the use of DNase I under appropriate conditions.
  • solution for example, in a buffer such as HEPES, a phosphate buffer solution, a citrate buffer solution, an acetate buffer solution; in some embodiments, such solution may be buffered to a pH within a range of, for example, about 6.5 to about 7.5; in some embodiments approximately 7.0.
  • production of polyribonucleotides may further include one or more of the following steps: purification, mixing, filtration, and/or filling.
  • polyribonucleotides can be purified (e.g., in some embodiments after in vitro transcription reaction), for example, to remove components utilized or formed in the course of the production, like, e.g., proteins, DNA portions, and/or or nucleotides.
  • Various nucleic acid purifications that are known in the art can be used in accordance with the present disclosure. Certain purification steps may be or include, for example, one or more of precipitation, column chromatography (including, e.g., but not limited to anionic, cationic, hydrophobic interaction chromatography (HIC)), solid substrate-based purification (e.g., magnetic bead-based purification).
  • polyribonucleotides may be purified using magnetic bead-based purification, which in some embodiments may be or comprise magnetic bead-based chromatography. In some embodiments, polyribonucleotides may be purified using hydrophobic interaction chromatography (HIC) and/or diafiltration. In some embodiments, polyribonucleotides may be purified using HIC followed by diafiltration.
  • HIC hydrophobic interaction chromatography
  • dsRNA may be obtained as side product during in vitro transcription. In some such embodiments, a second purification step may be performed to remove dsRNA contamination.
  • cellulose materials may be used to remove dsRNA contamination, for examples in some embodiments in a chromatographic format.
  • cellulose materials e.g., microcrystalline cellulose
  • cellulose materials may be used to purify polyribonucleotides according to methods described in WO 2017/182524, the entire content of which is incorporated herein by reference.
  • a batch of polyribonucleotides may be further processed by one or more steps of filtration and/or concentration.
  • polyribonucleotide(s) for example, after removal of dsRNA contamination, may be further subject to diafiltration (e.g., in some embodiments by tangential flow filtration), for example, to adjust the concentration of polyribonucleotides to a desirable RNA concentration and/or to exchange buffer to a drug substance buffer.
  • diafiltration e.g., in some embodiments by tangential flow filtration
  • polyribonucleotides may be processed through 0.2 ⁇ m filtration before they are filled into appropriate containers.
  • polyribonucleotides and compositions thereof may be manufactured in accordance with a process as described herein, or as otherwise known in the art.
  • polyribonucleotides and compositions thereof may be manufactured at a large scale.
  • a batch of polyribonucleotides can be manufactured at a scale of greater than 1 g, greater than 2 g, greater than 3 g, greater than 4 g, greater than 5 g, greater than 6 g, greater than 7 g, greater than 8 g, greater than 9 g, greater than 10 g, greater than 15 g, greater than 20 g, or higher.
  • RNA quality control may be performed and/or monitored at any time during production process of polyribonucleotides and/or compositions comprising the same.
  • RNA quality control parameters including one or more of RNA identity (e.g., sequence, length, and/or RNA natures), RNA integrity, RNA concentration, residual DNA template, and residual dsRNA, may be assessed and/or monitored after each or certain steps of a polyribonucleotide manufacturing process, e.g., after in vitro transcription, and/or each purification step.
  • the stability of polyribonucleotides can be assessed under various test storage conditions, for example, at room temperatures vs. fridge or sub-zero temperatures over a period of time (e.g., at least 3 months, at least 6 months, at least 9 months, at least 12 months, or longer).
  • polyribonucleotides e.g., ones described herein
  • compositions thereof may be stored stable at a fridge temperature (e.g., about 4qC to about 10qC) for at least 1 month or longer including, at least 2 months, at least 3 months, at least 4 months, at least 5 months, at least 6 months, at least 7 months, at least 8 months, at least 9 months, at least 10 months, at least 11 months, or at least 12 months or longer.
  • polyribonucleotides (e.g., ones described herein) and/or compositions thereof may be stored stable at a sub-zero temperature (e.g., -20qC or below) for at least 1 month or longer including, at least 2 months, at least 3 months, at least 4 months, at least 5 months, at least 6 months, at least 7 months, at least 8 months, at least 9 months, at least 10 months, at least 11 months, or at least 12 months or longer.
  • polyribonucleotides (e.g., ones described herein) and/or compositions thereof may be stored stable at room temperature (e.g., at about 25°C) for at least 1 month or longer.
  • one or more assessments may be utilized during manufacture, or other preparation or use of polyribonucleotides (e.g., as a release test).
  • one or more quality control parameters may be assessed to determine whether polyribonucleotides described herein meet or exceed acceptance criteria (e.g., for subsequent formulation and/or release for distribution).
  • quality control parameters may include, but are not limited to RNA integrity, RNA concentration, residual DNA template and/or residual dsRNA. Certain methods for assessing RNA quality are known in the art; for example, one of skill in the art will recognize that in some embodiments, one or more analytical tests can be used for RNA quality assessment.
  • a batch of polyribonucleotides may be assessed for one or more features as described herein to determine next action step(s). For example, a batch of polyribonucleotides can be designated for one or more further steps of manufacturing and/or formulation and/or distribution if RNA quality assessment indicates that such a batch of polyribonucleotides meet or exceed the relevant acceptance criteria. Otherwise, an alternative action can be taken (e.g., discarding the batch) if such a batch of polyribonucleotides does not meet or exceed the acceptance criteria.
  • a batch of polyribonucleotides that satisfy assessment results can be utilized for one or more further steps of manufacturing and/or formulation and/or distribution.
  • IX. DNA Constructs [0820] Among other things, the present disclosure provides DNA constructs, for example that may encode one or more antibody agents as described herein, or components thereof. In some embodiments, DNA constructs provided by and/or utilized in accordance with the present disclosure are comprised in a vector.
  • Non-limiting examples of a vector include plasmid vectors, cosmid vectors, phage vectors such as lambda phage, viral vectors such as retroviral, adenoviral or baculoviral vectors, or artificial chromosome vectors such as bacterial artificial chromosomes (BAC), yeast artificial chromosomes (YAC), or P1 artificial chromosomes (PAC).
  • a vector is an expression vector.
  • a vector is a cloning vector.
  • a vector is a nucleic acid construct that can receive or otherwise become linked to a nucleic acid element of interest (e.g., a construct that is or encodes a payload, or that imparts a particular functionality, etc.).
  • Expression vectors which may be plasmid or viral or other vectors, typically include an expressible sequence of interest (e.g., a coding sequence) that is functionally linked with one or more control elements (e.g., promoters, enhancers, transcription terminators, etc.). Typically, such control elements are selected for expression in a system of interest.
  • a system is ex vivo (e.g., an in vitro transcription system); in some embodiments, a system is in vivo (e.g., a bacterial, yeast, plant, insect, fish, vertebrate, mammalian cell or tissue, etc.).
  • Cloning vectors are generally used to modify, engineer, and/or duplicate (e.g., by replication in vivo, for example in a simple system such as bacteria or yeast, or in vitro, such as by amplification such as polymerase chain reaction or other amplification process).
  • a cloning vector may lack expression signals.
  • a vector may include replication elements such as primer binding site(s) and/or origin(s) of replication. In many embodiments, a vector may include insertion or modification sites such as restriction endonuclease recognition sites and/or guide RNA binding sites, etc.
  • a vector is a viral vector (e.g., an AAV vector). In some embodiments, a vector is a non-viral vector. In some embodiments, a vector is a plasmid.
  • a viral vector e.g., an AAV vector
  • a vector is a non-viral vector.
  • a vector is a plasmid.
  • polynucleotide(s) of the present disclosure are included in a DNA construct (e.g., a vector) amenable to transcription and/or translation.
  • an expression vector comprises a polynucleotide that encodes proteins and/or polypeptides of the present disclosure operatively linked to a sequence or sequences that control expression (e.g., promoters, start signals, stop signals, polyadenylation signals, activators, repressors, etc.).
  • a sequence or sequences that control expression are selected to achieve a desired level of expression.
  • more than one sequence that controls expression are utilized.
  • more than one sequence that controls expression are utilized to achieve a desired level of expression of a plurality of polynucleotides that encode a plurality of proteins and/or polypeptides.
  • a plurality of recombinant proteins and/or polypeptides are expressed from the same vector (e.g., a bi-cistronic vector, a tri-cistronic vector, multi-cistronic).
  • a plurality of polypeptides are expressed, each of which is expressed from a separate vector.
  • an expression vector comprising a polynucleotide of the present disclosure is used to produce an RNA and/or protein and/or polypeptide in a host cell.
  • a host cell may be in vitro (e.g., a cell line) – for example a cell or cell line (e.g., Human Embryonic Kidney (HEK cells), Chinese Hamster Ovary cells, etc.) suitable for producing polynucleotides of the present disclosure and proteins and/or polypeptides encoded by said polynucleotides.
  • HEK cells Human Embryonic Kidney
  • Chinese Hamster Ovary cells etc.
  • an expression vector is an RNA expression vector.
  • an RNA expression vector comprises a polynucleotide template used to produce a RNA in cell-free enzymatic mix.
  • an RNA expression vector comprising a polynucleotide template is enzymatically linearized prior to in vitro transcription.
  • a polynucleotide template is generated through PCR as a linear polynucleotide template.
  • a linearized polynucleotide is mixed with enzymes suitable for RNA synthesis, RNA capping and/or purification.
  • the resulting RNA is suitable for producing proteins encoded by the RNA.
  • a vector may be introduced into host cells using transfection.
  • transfection is completed, for example, using calcium phosphate transfection, lipofection, or polyethylenimine- mediated transfection.
  • a vector may be introduced into a host cell using transduction.
  • transformed host cells are cultured following introduction of a vector into a host cell to allow for expression of said recombinant polynucleotides.
  • a transformed host cells are cultured for at least 12 hours, 16 hours, 20 hours, 24 hours, 28 hours, 32 hours, 36 hours 40 hours, 44 hours, 48 hours, 52 hours, 56 hours, 60 hours, 64 hours, 68 hours, 72 hours or longer.
  • Transformed host cells are cultured in growth conditions (e.g., temperature, carbon-dioxide levels, growth medium) in accordance with the requirements of a host cell selected.
  • growth conditions e.g., temperature, carbon-dioxide levels, growth medium
  • a skilled artisan would recognize culture conditions for host cells selected are well known in the art.
  • X. Dosage Regimens [0833]
  • the present disclosure provides a method of treating or preventing herpes simplex virus (HSV) infection comprising administering to a subject in need thereof a therapeutically effective amount of a composition disclosed herein, in a treatment cycle comprising one or more doses (e.g., one dose, two doses, three doses, four doses, five doses, six doses, seven doses, eight doses, nine doses, or ten doses) of the composition.
  • doses e.g., one dose, two doses, three doses, four doses, five doses, six doses, seven doses, eight doses,
  • a treatment cycle comprises two or more doses (e.g., two doses, three doses, four doses, five doses, six doses, seven doses, eight doses, nine doses, or ten doses) of the composition.
  • a treatment cycle comprises two doses.
  • a first dose is a priming dose of a composition disclosed herein.
  • a second dose is a booster dose of a composition disclosed herein.
  • a subject is administered one or more doses of a composition disclosed herein prior to infection with HSV (e.g., HSV-1, HSV-2, or a combination thereof).
  • a subject is administered two or more doses of a composition disclosed herein prior to infection with HSV (e.g., HSV-1, HSV-2, or a combination thereof). In some embodiments, a subject is administered three or more doses of a composition disclosed herein prior to infection with HSV (e.g., HSV-1, HSV-2, or a combination thereof).
  • a second dose of the therapeutically effective amount of a composition disclosed herein is administered to a subject 1 day to 24 weeks, 3.5 days to 24 weeks, 1 week to 24 weeks, 2 weeks to 24 weeks, 4 weeks to 24 weeks, 6 weeks to 24 weeks, 8 weeks to 24 weeks, 10 weeks to 24 weeks, 12 weeks to 24 weeks, 16 weeks to 24 weeks, 20 weeks to 24 weeks, 1 day to 20 weeks, 3.5 days to 20 weeks, 1 week to 20 weeks, 2 weeks to 20 weeks, 4 weeks to 20 weeks, 6 weeks to 20 weeks, 8 weeks to 20 weeks, 10 weeks to 20 weeks, 12 weeks to 20 weeks, 16 weeks to 20 weeks, 1 day to 16 weeks, 3.5 days to 16 weeks, 1 week to 16 weeks, 2 weeks to 16 weeks, 4 weeks to 16 weeks, 6 weeks to 16 weeks, 8 weeks to 16 weeks, 10 weeks to 16 weeks, 12 weeks to 16 weeks, 1 day to 12 weeks, 3.5 days to 12 weeks, 1 week to 12 weeks, 2 weeks to 12 weeks, 4 weeks to 12 weeks, 6 weeks to 12 weeks, 8 weeks to 16 weeks, 10 weeks to 16 weeks
  • a second dose of the therapeutically effective amount of a composition disclosed herein is administered 1 week to 14 weeks after administration of the first dose of the therapeutically effective amount of the composition to the subject. In some embodiments, a second dose of the therapeutically effective amount of a composition disclosed herein is administered 4 weeks to 12 weeks after administration of the first dose of the therapeutically effective amount of the composition to the subject. In some embodiments, a second dose of the therapeutically effective amount of a composition disclosed herein is administered 6 weeks to 10 weeks after administration of the first dose of the therapeutically effective amount of the composition to the subject.
  • a second dose of the therapeutically effective amount of a composition disclosed herein is administered about 1 week, about 2 weeks, about 4 weeks, about 6 weeks, about 8 weeks, about 10 weeks, about 12 weeks, about 16 weeks, about 20 weeks, or about 24 weeks after administration of the first dose of the therapeutically effective amount of the composition to the subject. In some embodiments, a second dose of the therapeutically effective amount of a composition disclosed herein is administered about 8 weeks after administration of the first dose of the therapeutically effective amount of the composition to the subject.
  • a second dose of the therapeutically effective amount of a composition disclosed herein is administered about 1 day, about 7 days, about 14 days, about 28 days, about 35 days, about 40 days, about 45 days, about 50 days, about 51 days, about 52 days, about 53 days, about 54 days, about 55 days, about 56 days, about 57 days, about 58 days, about 59 days, or about 60 days after administration of the first dose of the therapeutically effective amount of the composition to the subject.
  • a third dose of the therapeutically effective amount of a composition disclosed herein is administered to a subject 1 day to 24 weeks, 3.5 days to 24 weeks, 1 week to 24 weeks, 2 weeks to 24 weeks, 4 weeks to 24 weeks, 6 weeks to 24 weeks, 8 weeks to 24 weeks, 10 weeks to 24 weeks, 12 weeks to 24 weeks, 16 weeks to 24 weeks, 20 weeks to 24 weeks, 1 day to 20 weeks, 3.5 days to 20 weeks, 1 week to 20 weeks, 2 weeks to 20 weeks, 4 weeks to 20 weeks, 6 weeks to 20 weeks, 8 weeks to 20 weeks, 10 weeks to 20 weeks, 12 weeks to 20 weeks, 16 weeks to 20 weeks, 1 day to 16 weeks, 3.5 days to 16 weeks, 1 week to 16 weeks, 2 weeks to 16 weeks, 4 weeks to 16 weeks, 6 weeks to 16 weeks, 8 weeks to 16 weeks, 10 weeks to 16 weeks, 12 weeks to 16 weeks, 1 day to 12 weeks, 3.5 days to 12 weeks, 1 week to 12 weeks, 2 weeks to 12 weeks, 4 weeks to 12 weeks, 6 weeks to 12 weeks, 8 weeks to 16 weeks, 10 weeks to 16 weeks
  • a third dose of the therapeutically effective amount of a composition disclosed herein is administered 4 weeks to 24 weeks after administration of the first dose of the therapeutically effective amount of the composition to the subject. In some embodiments, a third dose of the therapeutically effective amount of a composition disclosed herein is administered 12 weeks to 20 weeks after administration of the first dose of the therapeutically effective amount of the composition to the subject. In some embodiments, a third dose of the therapeutically effective amount of a composition disclosed herein is administered 14 weeks to 18 weeks after administration of the first dose of the therapeutically effective amount of the composition to the subject.
  • a third dose of the therapeutically effective amount of a composition disclosed herein is administered about 6 weeks, about 8 weeks, about 10 weeks, about 12 weeks, about 13 weeks, about 14 weeks, about 15 weeks, about 16 weeks, about 17 weeks, about 18 weeks, about 19 weeks, about 20 weeks, or about 24 weeks after administration of the first dose of the therapeutically effective amount of the composition to the subject.
  • a third dose of the therapeutically effective amount of a composition disclosed herein is administered about 8 weeks after administration of the first dose of the therapeutically effective amount of the composition to the subject.
  • a third dose of the therapeutically effective amount of a composition disclosed herein is administered about 10 days, about 20 days, about 30 days, about 40 days, about 50 days, about 60 days, about 70 days, about 80 days, about 90 days, about 100 days, about 105 days, about 110 days, about 111 days, about 112 days, about 113 days, about 114 days, about 115 days, about 116 days, or about 117 days after administration of the first dose of the therapeutically effective amount of the composition to the subject.
  • each of the one or more doses of a therapeutically effective amount of a composition disclosed herein is administered to a subject intramuscularly, subcutaneously, orally, or intranasally.
  • each of the one or more doses of a therapeutically effective amount of a composition disclosed herein is administered to a subject intramuscularly.
  • each of the one or more doses comprises 0.1 to 500 ⁇ g, 0.2 to 500 ⁇ g, 0.5 to 500 ⁇ g, 0.75 to 500 ⁇ g, 1 to 500 ⁇ g.
  • each of the one or more doses comprises 1 ⁇ g to 250 ⁇ g of one or more polyribonucleotides encoding one or more HSV glycoproteins or antigenic fragments thereof.
  • each of the one or more doses comprises 2 ⁇ g to 200 ⁇ g of one or more polyribonucleotides encoding one or more HSV glycoproteins or antigenic fragments thereof. In some embodiments, each of the one or more doses comprises 3 ⁇ g to 100 ⁇ g of one or more polyribonucleotides encoding one or more HSV glycoproteins or antigenic fragments thereof. In some embodiments, each of the one or more doses comprises 3 ⁇ g to 60 ⁇ g of one or more polyribonucleotides encoding one or more HSV glycoproteins or antigenic fragments thereof.
  • each of the one or more doses comprises 3 ⁇ g of one or more polyribonucleotides encoding one or more HSV glycoproteins or antigenic fragments thereof. In some embodiments, each of the one or more doses comprises 10 ⁇ g of one or more polyribonucleotides encoding one or more HSV glycoproteins or antigenic fragments thereof. In some embodiments, each of the one or more doses comprises 30 ⁇ g of one or more polyribonucleotides encoding one or more HSV glycoproteins or antigenic fragments thereof. In some embodiments, each of the one or more doses comprises 60 ⁇ g of one or more polyribonucleotides encoding one or more HSV glycoproteins or antigenic fragments thereof.
  • each of the one or more doses comprises 0.5 ⁇ g, 0.75 ⁇ g, 1.0 ⁇ g, 1.25 ⁇ g, 1.33 ⁇ g, 1.5 ⁇ g, 1.75 ⁇ g, 2.0 ⁇ g, 2.25 ⁇ g, 2.5 ⁇ g, 5 ⁇ g, 7.5 ⁇ g, 10 ⁇ g, 15 ⁇ g, 20 ⁇ g, or 25 ⁇ g of one or more polyribonucleotides encoding one or more HSV glycoproteins or antigenic fragments thereof.
  • each of the one or more doses comprises 1 ⁇ g of each of three polyribonucleotides, wherein each polyribonucleotide encodes a different HSV glycoprotein or antigenic fragment thereof.
  • each of the one or more doses comprises 3.33 ⁇ g of each of three polyribonucleotides, wherein each polyribonucleotide encodes a different HSV glycoprotein or antigenic fragment thereof.
  • each of the one or more doses comprises 10 ⁇ g of each of three polyribonucleotides, wherein each polyribonucleotide encodes a different HSV glycoprotein or antigenic fragment thereof.
  • each of the one or more doses comprises 20 ⁇ g of each of three polyribonucleotides, wherein each polyribonucleotide encodes a different HSV glycoprotein or antigenic fragment thereof.
  • the present disclosure provides a method of treating or preventing HSV infection comprising administering to a subject in need thereof one or more doses of a therapeutically effective amount of a composition as disclosed herein as part of a combination therapy.
  • the present disclosure provides a method of treating or preventing HSV infection comprising administering to a subject in need thereof one or more doses of a therapeutically effective amount of a composition as disclosed herein concomitantly with administration of an antipyretic medication.
  • an antipyretic medication is administered within 60 minutes, within 30 minutes, or within 15 minutes after administration of one or more doses of a therapeutically effective amount of a composition as disclosed herein.
  • an antipyretic medication is administered concurrently with administration of one or more doses of a therapeutically effective amount of a composition as disclosed herein.
  • an antipyretic medication is administered within 60 minutes, within 30 minutes, or within 15 minutes before administration of one of more doses of a therapeutically effective amount of a composition as disclosed herein.
  • the antipyretic comprises acetaminophen, a non-steroidal anti-inflammatory drug (NSAID), salicylamide, salicyl salicylate, methyl salicylate, magnesium salicylate, fatelamine, ethenzamide, diflunisal, choline magnesium salicylate, benorylate/benorilatem and amoxiprin, acetylsalicylate, ceclofenac, acemetacin, alclofenac, bromfenac, diclofenac, etodolac, indomethacin, nabumetone, oxametacin, proglumetacin, sulindac, tolmetin, iminoprofen, benoxaprofen, carprofen, dexi
  • NSAID non-steroidal anti-
  • the antipyretic is an NSAID. In certain embodiments, the antipyretic is acetaminophen.
  • the present disclosure provides a method of treating or preventing HSV (HSV-1, HSV-2, or a combination thereof) infection comprising administering to a subject in need thereof one or more doses of a therapeutically effective amount of a composition as disclosed herein concomitantly with administration of an analgesic medication. In some embodiments, an analgesic medication is administered within 60 minutes, within 30 minutes, or within 15 minutes after administration of one or more doses of a therapeutically effective amount of a composition as disclosed herein.
  • an analgesic medication is administered concurrently with administration of one or more doses of a therapeutically effective amount of a composition as disclosed herein. In some embodiments, an analgesic medication is administered within 60 minutes, within 30 minutes, or within 15 minutes before administration of one of more doses of a therapeutically effective amount of a composition as disclosed herein.
  • an analgesic comprises acetaminophen, salicylamide, salicyl salicylate, methyl salicylate, magnesium salicylate, fatelamine, ethenzamide, diflunisal, choline magnesium salicylate, benorylate/benorilatem and amoxiprin, acetylsalicylate, ceclofenac, acemetacin, alclofenac, bromfenac, diclofenac, etodolac, indomethacin, nabumetone, oxametacin, proglumetacin, sulindac, tolmetin, iminoprofen, benoxaprofen, carprofen, dexibuprofen, dexketoprofen, fenbufen, fenoprofen, flunoxaprofen, flurbiprofen, ibuprofen, ibuproxam, indoprof
  • the analgesic is acetaminophen.
  • the present disclosure provides a method of treating or preventing HSV (HSV-1, HSV-2, or a combination thereof) infection comprising administering to a subject in need thereof one or more doses of a therapeutically effective amount of a composition as disclosed herein concomitantly with administration of an antipyretic medication and an analgesic medication.
  • an antipyretic medication and analgesic medication is administered within 60 minutes, within 30 minutes, or within 15 minutes after administration of one or more doses of a therapeutically effective amount of a composition as disclosed herein.
  • an antipyretic medication and analgesic medication is administered concurrently with administration of one or more doses of a therapeutically effective amount of a composition as disclosed herein. In some embodiments, an antipyretic medication and analgesic medication is administered within 60 minutes, within 30 minutes, or within 15 minutes before administration of one of more doses of a therapeutically effective amount of a composition as disclosed herein. [0851] In some embodiments, an antipyretic medication and/or analgesic medication is acetaminophen.
  • acetaminophen is administered to the subject at a dose of 0.1 g/day to 20 g/day, 0.25 g/day to 20 g/day, 0.5 g/day to 20 g/day, 0.75 g/day to 20 g/day, 1.0 g/day to 20 g/day, 1.25 g/day to 20 g/day, 1.5 g/day to 20 g/day, 1.75 g/day to 20 g/day, 2.0 g/day to 20 g/day, 2.25 g/day to 201 g/day, 2.5 g/day to 20 g/day, 2.75 g/day to 20 g/day, 3.0 g/day to 20 g/day, 3.25 g/day to 20 g/day, 3.5 g/day to 20 g/day, 3.75 g/day to 20 g/day, 4.0 g/day to 20 g/day, 4.25 g/day to 20 g/day,
  • acetaminophen is administered to the subject at a dose of 0.5 g/day to 10 g/day. In some embodiments, acetaminophen is administered to the subject at a dose of 1 g/day to 5 g/day. [0852] In some embodiments, acetaminophen is administered to the subject at a dose of about 0.5 g/day, about 1 g/day, about, about 1.5 g/day, about 2 g/day, about 2.5 g/day, about 3 g/day, about 3.5 g/day, about 4 g/day, about 4.5 g/day, or about 5 g/day.
  • acetaminophen is administered to the subject at a dose of about 4 g/day.
  • the present disclosure provides a method of treating or preventing HSV (HSV-1, HSV-2, or a combination thereof) infection.
  • the present disclosure provides a method of treating or preventing HSV (HSV-1, HSV-2, or a combination thereof) infection comprising administering to a subject in need thereof one or more doses of a therapeutically effective amount of a composition as disclosed herein, wherein the subject is further administered a medically indicated vaccine at least 1 day, at least 2 days, at least 3 days, at least 4 days, at least 5 days, at least 6 days, at least 7 days, at least 8 days, at least 9 days, at least 10 days, at least 12 days, at least 14 days, at least 16 days, at least 17 days, at least 18 days, at least 19 days, or at least 20 days before administration of the therapeutically acceptable amount of the composition.
  • a medically indicated vaccine at least 1 day, at least 2 days, at least 3 days, at least 4 days, at least 5 days, at least 6 days, at least 7 days, at least 8 days, at least 9 days, at least 10 days, at least 12 days, at least 14 days, at least 16 days, at least 17 days, at least 18 days
  • the subject is further administered a medically indicated vaccine at least 14 days before the administration of the therapeutically acceptable amount of the composition.
  • the present disclosure provides a method of treating or preventing HSV (HSV-1, HSV-2, or a combination thereof) infection comprising administering to a subject in need thereof one or more doses of a therapeutically effective amount of a composition as disclosed herein, wherein the subject is further administered a medically indicated vaccine at least 1 day, at least 2 days, at least 3 days, at least 4 days, at least 5 days, at least 6 days, at least 7 days, at least 8 days, at least 9 days, at least 10 days, at least 12 days, at least 14 days, at least 16 days, at least 17 days, at least 18 days, at least 19 days, or at least 20 days after administration of the therapeutically acceptable amount of the composition.
  • a medically indicated vaccine includes, but is not limited to, a rabies vaccine, a tetanus vaccine, a hepatitis A vaccine, a hepatitis B vaccine, a measles mumps rubella (MMR) vaccine, a polio vaccine, a diphtheria vaccine, a varicella vaccine, a pertussis vaccine, a shingles vaccine, a pneumococcal vaccine, a human papillomavirus vaccine (HPV), a meningococcal vaccine, or a rotavirus vaccine.
  • MMR measles mumps rubella
  • HPV human papillomavirus vaccine
  • a medically indicated vaccine is a rabies vaccine or a tetanus vaccine.
  • XII. Monitoring Efficacy [0857]
  • the present disclosure provides a method of treating or preventing HSV (HSV-1, HSV-2, or a combination thereof) infection comprising administering to a subject in need thereof one or more doses of a therapeutically effective amount of a composition as disclosed herein, wherein the method further comprises collecting one or more samples from the subject after administration of the one or more doses of a therapeutically effective amount of the composition.
  • At least one, at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, at least ten, at least eleven, at least twelve, at least thirteen, at least fourteen, or at least fifteen samples are collected from the subject after administration of the one or more doses of a therapeutically effective amount of the composition.
  • the one or more samples collected from the subject is a blood volume draw.
  • a sample is collected from the subject about 120 minutes, about 90 minutes, about 60 minutes, about 45 minutes, about 30 minutes, about 20 minutes, about 15 minutes, about 10 minutes, about 7.5 minutes, about 5 minutes, about 4 minutes, about 3 minutes, about 2 minutes or about 1 minute before administration of a first dose of one or more doses of a therapeutically effective amount of the composition.
  • a sample is collected from the subject about 1 minute, about 2 minutes, about 3 minutes, about 4 minutes, about 5 minutes, about 7.5 minutes, about 10 minutes, about 15 minutes, about 20 minutes, about 30 minutes, about 45 minutes, about 60 minutes, about 90 minutes, or about 120 minutes after administration of a first dose of one or more doses of a therapeutically effective amount of the composition.
  • a sample is collected about 1 week after administration of a first dose of one or more doses of a therapeutically effective amount of the composition. In some embodiments, a sample is collected about 2 weeks after administration of a first dose of one or more doses of a therapeutically effective amount of the composition. In some embodiments, a sample is collected about 4 weeks after administration of a first dose of one or more doses of a therapeutically effective amount of the composition. In some embodiments, a sample is collected about 5 weeks after administration of a first dose of one or more doses of a therapeutically effective amount of the composition. In some embodiments, a sample is collected about 6 weeks after administration of a first dose of one or more doses of a therapeutically effective amount of the composition.
  • a sample is collected about 8 weeks after administration of a first dose of one or more doses of a therapeutically effective amount of the composition. In some embodiments, a sample is collected about 16 weeks after administration of a first dose of one or more doses of a therapeutically effective amount of the composition. In some embodiments, a sample is collected about 7 months after administration of a first dose of one or more doses of a therapeutically effective amount of the composition. In some embodiments, a sample is collected about 13 months after administration of a first dose of one or more doses of a therapeutically effective amount of the composition.
  • a sample is collected from the subject about 120 minutes, about 90 minutes, about 60 minutes, about 45 minutes, about 30 minutes, about 20 minutes, about 15 minutes, about 10 minutes, about 7.5 minutes, about 5 minutes, about 4 minutes, about 3 minutes, about 2 minutes or about 1 minute before administration of a second dose of two or more doses of a therapeutically effective amount of the composition.
  • a sample is collected from the subject about 1 minute, about 2 minutes, about 3 minutes, about 4 minutes, about 5 minutes, about 7.5 minutes, about 10 minutes, about 15 minutes, about 20 minutes, about 30 minutes, about 45 minutes, about 60 minutes, about 90 minutes, or about 120 minutes after administration of a second dose of two or more doses of a therapeutically effective amount of the composition.
  • a sample is collected about 1 week after administration of a second dose of two or more doses of a therapeutically effective amount of the composition. In some embodiments, a sample is collected about 2 weeks after administration of a second dose of two or more doses of a therapeutically effective amount of the composition. In some embodiments, a sample is collected about 1 month after administration of a second dose of two or more doses of a therapeutically effective amount of the composition. In some embodiments, a sample is collected about 3 months after administration of a second dose of two or more doses of a therapeutically effective amount of the composition. In some embodiments, a sample is collected about 6 months after administration of a second dose of two or more doses of a therapeutically effective amount of the composition.
  • methods of the present disclosure further comprise a step of measuring levels of HSV (HSV-1 and/or HSV-2) virus-specific neutralizing antibodies in one or more samples collected from the subject.
  • levels of HSV (HSV-1 and/or HSV-2) virus-specific neutralizing antibodies are measured using any one of a number of assays known to persons of ordinary skill in the art.
  • levels of HSV (HSV-1 and/or HSV-2) virus-specific neutralizing antibodies are measured using a plaque reduction neutralization test (PRNT).
  • PRNT plaque reduction neutralization test
  • levels of HSV (HSV-1 and/or HSV-2) virus-specific neutralizing antibodies are measured using a pseudo-viral neutralization test.
  • methods of the present disclosure further comprise a step of measuring levels of neutralizing antibodies in one or more samples collected from a subject, where the neutralizing antibodies are specific for one or more HSV (HSV-1 and/or HSV-2) antigens or antigenic fragments thereof encoded by one or more polyribonucleotides in a composition disclosed herein.
  • levels of neutralizing antibodies are measured using any one of a number of assays known to persons of ordinary skill in the art.
  • ELISA enzyme-linked immunosorbent assay
  • exemplary LNP formulations that are useful for compositions can comprise at least one ionizable aminolipid.
  • exemplary LNP formulations that are useful for compositions can further comprise a helper lipid, which in some embodiments may be or comprise a neutral helper lipid.
  • exemplary LNP formulations that are useful for compositions can further comprise a polymer-conjugated lipid, for example in some embodiments PEG-conjugated lipids.
  • exemplary LNP formulations that are useful for compositions (e.g., immunogenic compositions, e.g., vaccines) as described herein can comprise at least one ionizable aminolipid, at least one helper lipid (e.g., a neutral helper lipid, which in some embodiments may comprise a phospholipid, a steroid, or combinations thereof), and at least one polymer-conjugated lipid (e.g., PEG-conjugated lipid).
  • an exemplary LNP formulation may comprise an ionizable aminolipid, a phospholipid, a steroid, and a PEG-conjugated lipid.
  • an ionizable aminolipid may be present in an exemplary LNP formulation within a range of 45 to 55 mol percent, 40 to 50 mol percent, 41 to 49 mol percent, 41 to 48 mol percent, 42 to 48 mol percent, 43 to 48 mol percent, 44 to 48 mol percent of total lipids.
  • an ionizable aminolipid is or comprises ((4-hydroxybutyl)azanediyl)bis(hexane-6,1-diyl)bis(2-hexyldecanoate) (also known as 6- [N-6-(2-hexyldecanoyloxy)hexyl-N-(4-hydroxybutyl)amino]hexyl 2-hexyldecanoate).
  • an ionizable aminolipid is or comprises SM-102 (heptadecan-9-yl 8 ((2 hydroxyethyl)(6 oxo 6- (undecyloxy)hexyl)amino)octanoate) or an aminolipid as described in Sabnis et al. “ A Novel Amino Lipid Series for mRNA Delivery: Improved Endosomal Escape and Sustained Pharmacology and Safety in Non-human Primates” Mol. Ther. (2018) 26:1509-1519.
  • an ionizable aminolipid is or comprises an ionizable aminolipid as disclosed in US2020/0163878 or WO2018/078053, the entire contents of each of which are incorporated herein by reference for the purposes described herein.
  • a phospholipid may be present in an exemplary LNP formulation within a range of 5 to 15 mol percent, 7 to 13 mol percent, or 9 to 11 mol percent of total lipids.
  • a phospholipid is or comprises 1,2-Distearoyl-sn-glycero- 3-phosphocholine (DSPC).
  • a sterol may be present in an exemplary LNP formulation within a range of 30 to 50 mol percent, 35 to 45 mol percent or 38 to 43 mol percent of total lipids.
  • a sterol is or comprises cholesterol.
  • a polymer conjugated lipid e.g., PEG-conjugated lipid
  • PEG-conjugated lipid may be present in an exemplary LNP formulation within a range of 1 to 10 mol percent, 1 to 5 mol percent, or 1 to 2.5 mol percent of total lipids.
  • a PEG-conjugated lipid is or comprises 2-[(polyethylene glycol)-2000]-N,N- ditetradecylacetamide (also known as 2-[2-( ⁇ -methoxy (polyethyleneglycol2000) ethoxy]-N,N- ditetradecylacetamide).
  • a phospholipid is or comprises PEG2000-DMG (1- monomethoxypolyethyleneglycol-2,3- dimyristylglycerol with polyethylene glycol of average molecular weight 2000).
  • a PEG-conjugated lipid is or comprises a PEG-lipid as disclosed in US2020/0163878 or WO2018/078053, the entire contents of each of which are incorporated herein by reference for the purposes described herein.
  • an exemplary LNP formulation comprises (i) an ionizable aminolipid within a range of 45 to 55 mol percent of total lipids; (ii) a phospholipid within a range of 8 to 12 mol percent of total lipids; (iii) a steroid within a range of 35 to 45 mol percent of total lipids; and (iv) a polymer conjugated (e.g., PEG- conjugated polymer) within a range of 1 to 2 mol percent of total lipids; and RNA molecules as described herein that are encapsulated within or associated with the lipid nanoparticles.
  • a polymer conjugated e.g., PEG- conjugated polymer
  • an exemplary LNP formulation comprises (i) ionizable amino lipid within a range of 45 to 55 mol percent of total lipids; (ii) DSPC within a range of 5 to 15 mol percent of total lipids; (iii) cholesterol within a range of 35 to 45 mol percent of total lipids; and (iv) a PEG-conjugated lipid within a range of 1 to 2 mol percent of total lipids; and RNA molecules as described herein that are encapsulated within or associated with the lipid nanoparticles.
  • an exemplary LNP formulation comprises (i) an ionizable aminolipid within a range of 40 to 50 mol percent of total lipids; (ii) a phospholipid within a range of 5 to 15 mol percent of total lipids; (iii) a steroid within a range of 35 to 45 mol percent of total lipids; and (iv) a polymer conjugated (e.g., PEG- conjugated polymer) within a range of 1 to 10 mol percent of total lipids; and RNA molecules as described herein that are encapsulated within or associated with the lipid nanoparticles.
  • a polymer conjugated e.g., PEG- conjugated polymer
  • an ionizable aminolipid is or comprises ((4-hydroxybutyl)azanediyl)bis(hexane-6,1-diyl)bis(2-hexyldecanoate) (also known as 6- [N-6-(2-hexyldecanoyloxy)hexyl-N-(4-hydroxybutyl)amino]hexyl 2-hexyldecanoate).
  • a phospholipid is or comprises 1,2-Distearoyl-sn-glycero-3-phosphocholine (DSPC).
  • a steroid is or comprises cholesterol.

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Abstract

The present disclosure provides pharmaceutical compositions for delivery of HSV antigens (e.g., an HSV vaccine) and related technologies (e.g., components thereof and/or methods relating thereto).

Description

PHARMACEUTICAL COMPOSITIONS FOR DELIVERY OF HERPES SIMPLEX VIRUS ANTIGENS AND RELATED METHODS RELATED APPLICATIONS [0001] This application claims priority to and the benefit of U.S. Provisional Patent Application No. 63/517597, filed on August 3, 2023, and U.S. Provisional Patent Application No. 63/639567, filed April 26, 2024, the entire contents of which are hereby incorporated by reference in their entirety. SEQUENCE LISTING [0002] The instant application contains a Sequence Listing which has been submitted herewith and is hereby incorporated by reference in its entirety. Said .xml copy, created on August 2, 2024 is named 2013237-1141, and is 1,333,061 bytes in size. BACKGROUND [0003] Herpes simplex viruses (HSV), commonly referred to only as herpes, are categorized into two types: herpes simplex virus, type 1 (HSV-1, or oral herpes) and herpes simplex virus, type 2 (HSV-2, or genital herpes). According to the World Health Organization, an estimated 3.7 billion people under age 50 (67% of global population) have HSV-1 infection globally. HSV-1 prevalence is understood as being highest in Africa and lowest in the Americas. An estimated 491 million people aged 15-49 (13% of global population) worldwide have HSV-2 infection. More women are infected with HSV-2 than men, because sexual transmission of HSV is more efficient from men to women than from women to men. Prevalence of HSV-2 infection was estimated to be highest in Africa, followed by the Americas. Prevalence of HSV-2 was also shown to increase with age, though the highest numbers of people newly-infected have historically been in adolescents. Both HSV-1 and HSV-2 infections are lifelong. SUMMARY [0004] The present disclosure provides technologies (e.g., combinations, compositions, methods, etc.) for delivery of HSV polypeptides or antigenic portions thereof. [0005] The present disclosure provides pharmaceutical compositions (e.g., immunogenic compositions, e.g., vaccines) for delivering herpes simplex virus (HSV) antigen constructs to a subject (e.g., a patient) and related technologies (e.g., methods). In particular, the present disclosure provides HSV compositions (e.g., immunogenic compositions, e.g., vaccines) and related technologies (e.g., methods). [0006] The present disclosure also provides that HSV glycoprotein C (gC) or antigenic portions thereof, HSV glycoprotein D (gD) or antigenic portions thereof, glycoprotein E (gE) or antigenic portions thereof, or combinations thereof can be useful in preventing or treating HSV, e.g., HSV-1, HSV-2, or both, as further disclosed herein. [0007] The present disclosure provides, for example, combinations comprising a plurality of polyribonucleotides, wherein the plurality of polyribonucleotides comprises a first set of polyribonucleotides that encode one or more glycoprotein (GP) polypeptides. In some embodiments, a GP polypeptide comprises an HSV glycoprotein or antigenic portions thereof. In some embodiments, polyribonucleotides that encode one or more GP polypeptides provided herein encode one or more of HSV-2 gC, gD, and/or gE or antigenic portions thereof (e.g., in a construct). In some embodiments, such polyribonucleotides of a first set of polyribonucleotides can be part of an RNA construct. In some embodiments, a polyribonucleotide that encodes a GP polypeptide or RNA construct as described herein can be part of a composition (e.g., a pharmaceutical composition, e.g., an immunogenic composition, e.g., a vaccine). [0008] The present disclosure also provides a combination comprising a plurality of polyribonucleotides, wherein the plurality of polyribonucleotides comprises a first set of polyribonucleotides and a second set of polyribonucleotides. In some embodiments, a second set of polyribonucleotides encode one or more T-cell string polypeptides. In some embodiments, a T-cell string polypeptide comprises one or more HSV T-cell antigens or antigenic portions thereof. In some embodiments, such polyribonucleotides of a second set of polyribonucleotides can be part of an RNA construct. In some embodiments, a polyribonucleotide that encodes a T-cell string polypeptide or a corresponding RNA construct as described herein can be part of a composition (e.g., a pharmaceutical composition, e.g., an immunogenic composition, e.g., a vaccine). [0009] In some embodiments, technologies provided herein are directed against HSV. BRIEF DESCRIPTION OF THE DRAWING [0010] The Drawing included herein, which is composed of the following Figures, is for illustration purposes only and not for limitation. [0011] FIG.1 is a schematic of an HSV particle. [0012] FIG.2 is a schematic overview of the HSV life cycle. FIG.2 has been modified from Ibanez, F.J., et al., “Experimental Dissection of the Lytic Replication Cycles of Herpes Simplex Virus in vitro,” Front Microbiol. 2018; 9: 2406, which is incorporated herein by reference in its entirety. [0013] FIGS.3A-3F show HSV-2 gC, gD, or gE expression in HEK293T cells transfected with 0.2 μg/mL LNP-formulated trivalent nucleoside-modified RNA (modRNA) encoding gC, gD, and gE (drug product, DP). Expression of HSV-2 gC, gD and gE protein was detected by flow cytometry using primary monoclonal mouse antibodies detecting the respective antigen and a secondary fluorescent tagged anti-mouse antibody. Representative data from one experiment showing percentage of gC (FIG.3A), gD (FIG.3B), and gE (FIG.3C) protein-expressing cells and median fluorescence intensities (MFI) (FIG.3D, 3E, and 3F, respectively) of the total HEK293T population are depicted per antigen. Data shown are mean+SD of HEK293T transfections performed in triplicates. [0014] FIG.4 shows a schematic overview of a study in guinea pigs investigating a composition candidate against HSV-2. Guinea pigs were immunized IM on day 0 and day 28 with an HSV-2 composition candidate containing total HSV-2 gC/gD/gE RNA at a concentration of 3 μg, 15 μg, or PBS control, as outlined in Table 24. Twenty-eight days after the second immunization i.e., on day 56, animals were bled. On day 60, the guinea pigs were challenged with a lethal dose of 5 x 105 PFU of HSV-2 strain MS (25-fold LD50). d = day; DRG = dorsal root ganglia; HSV-2 = herpes simplex virus-2; gC2 = glycoprotein C from herpes simplex virus-2; gD2 = glycoprotein D from herpes simplex virus-2; gE2 = glycoprotein E from herpes simplex virus-2; IM = intramuscular; PFU = plaque forming unit; RNA-LNP315 = RNA lipid nanoparticles formulated with ALC-0315. [0015] FIGS.5A-5C show serum IgG antibody titers observed one month after a 2nd immunization in guinea pigs immunized with an HSV-2 composition candidate described herein. Serum antibody titers were determined by ELISA at day 56, 4 weeks after the second immunization with a composition (“trivalent vaccine”) comprising three polyribonucleotides encoding glycoprotein C (gC), glycoprotein D (gD) and glycoprotein E (gE), respectively. The dose level represents total RNA content of three RNAs encoding for the respective HSV-2 gC, HSV-2 gD and HSV-2 gE antigens in a 1:1:1 ratio. Geometric mean ± 95% CI and individual animal values are shown. P values were calculated by Kruskal-Wallis test. * = p value ≤ 0.05; ** = p value ≤ 0.01; **** = p value ≤ 0.0001; gC2 = glycoprotein C from herpes simplex virus-2; gD2 = glycoprotein D from herpes simplex virus-2; gE2 = glycoprotein E from herpes simplex virus-2; IgG = immunoglobulin G; RNA = ribonucleic acid, GMT = geometric mean; CI = confidence interval. As shown in FIG.5, administration of the HSV-2 composition candidate induced high IgG antibody titers against each of HSV-2 gC (FIG.5A), HSV-2 gD (FIG.5B) and HSV-2 gE (FIG.5C), with a 15 μg dose inducing higher titers for gC and gD antigens than a 3 μg dose. [0016] FIGS.6A-6C show vaginal IgG antibody titers in guinea pigs one month after a 2nd immunization with an HSV-2 modRNA composition described herein. Vaginal antibody titers were determined by ELISA at day 56, four weeks after the second immunization with a trivalent composition. The dose level represents total RNA content of three RNAs encoding for the respective HSV-2 gC, HSV-2 gD and HSV-2 gE antigens in a 1:1:1 ratio. Geometric mean ± 95% CI and individual animal values are shown. P values were calculated by Kruskal-Wallis test. * = p value ≤ 0.05; ** = p value ≤ 0.01; *** = p value ≤ 0.001; **** = p value ≤ 0.0001; gC2 = glycoprotein C from herpes simplex virus-2; gD2 = glycoprotein D from herpes simplex virus-2; gE2 = glycoprotein E from herpes simplex virus- 2; IgG = immunoglobulin G; RNA = ribonucleic acid; GMT = geometric mean; CI = confidence interval. As shown in FIG.6, high vaginal IgG titers were induced against each of HSV-2 gC (FIG.6A), HSV-2 gD (FIG.6B), and HSV-2 gE (FIG.6C), with a 15 μg dose inducing higher titers for gE antigen than a 3 μg dose. [0017] FIG.7 shows serum neutralizing antibody titers to HSV-2 in guinea pigs one month after a 2nd immunization with an HSV-2 modified RNA (modRNA) composition described herein. Neutralizing antibody titers were determined using a serum HSV-2 plaque reduction assay and defined as highest dilution of serum with 5% human complement that reduced the number of HSV-2 plaques by 50%. Samples were collected at day 56, 4 weeks after the second immunization. The dose level represents total RNA content of three RNAs encoding for the respective HSV-2 gC, HSV-2 gD and HSV-2 gE antigens in a 1:1:1 ratio. Geometric mean ± 95% CI and individual animal values are shown. P values were calculated by Mann-Whitney test. * = p-value ≤ 0.05; gC2 = glycoprotein C from herpes simplex virus-2; gD2 = glycoprotein D from herpes simplex virus-2; gE2 = glycoprotein E from herpes simplex virus-2; RNA = ribonucleic acid; GMT = geometric mean; CI = confidence interval. As shown in FIG.7, high neutralization titers were observed at both a 3 μg dose and a 15 μg dose, with a 15 μg dose inducing higher neutralizing antibody titers than a 3 μg dose. [0018] FIGS.8A-8C shows weight loss in guinea pigs administered an HSV-2 composition described herein, following HSV-2 viral challenge. Relative body weight changes in guinea pigs up to 14 days after viral challenge with a lethal intravaginal dose of HSV-2 at day 60, approximately one month after second immunization with 3 μg or 15 μg of a trivalent composition, or PBS. The dose level represents total RNA content of three RNAs encoding for the respective gC2, gD2 and gE2 antigens in a 1:1:1 ratio. PBS = phosphate buffered saline; gC2 = glycoprotein C from herpes simplex virus-2; gD2 = glycoprotein D from herpes simplex virus-2; gE2 = glycoprotein E from herpes simplex virus-2; RNA = ribonucleic acid. As shown in FIG.8, administration of a 3 μg dose of an HSV-2 composition (FIG.8B) decreased body weight loss relative to the PBS negative control (FIG.8A), and a 15 μg dose of an HSV-2 composition (FIG.8C) decreased body weight losses further still. [0019] FIG.9 shows survival of guinea pigs immunized with an HSV-2 composition described herein, up to day 48 after HSV-2 viral challenge. Probability of survival of guinea pigs up to 48 days after lethal intravaginal challenge with HSV-2 at day 60, approximately one month after second immunization with 3 μg or 15 μg of trivalent a composition, or PBS. The dose level represents total RNA content of three RNAs encoding for the respective HSV-2 gC, HSV-2 gD and HSV-2 gE antigens in a 1:1:1 ratio. P values were calculated by log-rank (Mantel-Cox) test. ** = p-value ≤ 0.01; PBS = phosphate buffered saline, gC2 = glycoprotein C from herpes simplex virus-2; gD2 = glycoprotein D from herpes simplex virus-2; gE2 = glycoprotein E from herpes simplex virus-2; HSV-2 = herpes simplex virus-2; RNA = ribonucleic acid. As shown in FIG.9, administration of a 3 μg dose of an HSV-2 composition significantly increased survival of the guinea pigs, and administration a 15 μg dose increased survival further still. [0020] FIGS.10A-10C show individual evaluation of genital disease in guinea pigs administered an RNA composition described herein, up to day 48 after challenge with a lethal intravaginal dose of HSV-2. Results at day 60, approximately one month after the second vaccination with an HSV-2 modified RNA (modRNA) composition are shown. FIG.10A shows the mean number of days with genital disease during this period and FIG.10B shows the mean severity of genital lesions of days with genital disease. Mean ± SEM and individual animal values are shown. FIG.10C shows the mean number of urinary retention days. The dose level represents total RNA content of three RNAs encoding for the respective HSV-2 gC, HSV-2 gD and HSV-2 gE antigens in a 1:1:1 ratio. P values were calculated by Mann-Whitney test. Black circles with red outlines are associated with animals that succumbed after viral challenge in the PBS group. * = p-value ≤ 0.05; HSV-2 = herpes simplex virus-2; SEM = standard error of the mean; PBS = phosphate buffered saline, gC2 = glycoprotein C from herpes simplex virus-2; gD2 = glycoprotein D from herpes simplex virus-2; gE2 = glycoprotein E from herpes simplex virus-2; RNA = ribonucleic acid. Because a majority of the animals in the PBS control group died within two weeks of viral challenge, disease score as measured by number of genital lesion days and/or severity of genital lesions was underrepresented in this group, and a statistical analysis of this group was not performed. As shown in FIG.10, administering an HSV-2 composition described herein reduces the number of days during which a genital lesion was observed, decreased the severity of lesions that were observed, and decreased urinary retention days. [0021] FIG.11 shows cumulative disease score in guinea pigs administered an RNA composition described herein, up to day 48 after challenge with a lethal intravaginal dose of HSV-2. Results at day 60 are shown, approximately one month after the second vaccination. The mean number of days with genital disease per group is shown over the course of 48 days. The dose level represents total RNA content of three RNAs encoding for the respective gC2, gD2 and gE2 antigens in a 1:1:1 ratio. No scoring for days after death was assigned to animals that succumbed to viral disease. HSV-2 = herpes simplex virus-2; PBS = phosphate buffered saline. gC2 = glycoprotein C from herpes simplex virus-2; gD2 = glycoprotein D from herpes simplex virus-2; gE2 = glycoprotein E from herpes simplex virus-2; RNA = ribonucleic acid. As shown in FIG.11, administration of 3 μg of an HSV-2 composition significantly decreased mean cumulative disease days, and administration 15 μg decreased mean cumulative disease days further still. [0022] FIGS.12A-12C show vaginal virus titers in guinea pigs administered an HSV-2 composition described herein, 2 and 4 days after viral challenge. Vaginal HSV-2 titers were determined by plaque assay 2 days (FIG.12A) and 4 days (FIG.12B) after a lethal intravaginal challenge with HSV-2. Results are plotted as means ± SEM and individual animal values. Mean days of genital shedding of HSV-2 DNA were analyzed by PCR and displayed in (FIG.12C). The dose level for the HSV-2 composition represents total RNA content of three RNAs in a 1:1:1 ratio encoding for the respective HSV-2 gC, HSV-2 gD and HSV-2 gE antigens. P values were calculated by Kruskal-Wallis test. SEM = standard error of the mean; gC2 = glycoprotein C from herpes simplex virus-2; gD2 = glycoprotein D from herpes simplex virus-2; gE2 = glycoprotein E from herpes simplex virus-2; HSV-2 = herpes simplex virus-2; DNA = deoxyribonucleic acid; RNA = ribonucleic acid. [0023] FIGS.13A-13B show DNA copy numbers in DRG and spinal cord of guinea pigs administered an HSV-2 composition disclosed herein, on day 48 after viral challenge. DRG and spinal cord HSV-2 DNA copy numbers in guinea pigs on day 48 following viral challenge with a lethal intravaginal dose of HSV-2 were analyzed by qPCR. HSV-2 genome copies in DRG (FIG.13A) and spinal cord (FIG.13B) relative to GAPDH expression at day 48 after viral challenge are shown for immunized animals. Mean ± SEM and individual animal values are shown. The dose level for the HSV-2 composition represents total RNA content of three RNAs in a 1:1:1 ratio encoding for the respective HSV-2 gC, HSV-2 gD and HSV-2 gE antigens. P values were calculated by Mann-Whitney test. DRG = dorsal root ganglia; SEM = standard error of the mean; gC2 = glycoprotein C from herpes simplex virus-2; gD2 = glycoprotein D from herpes simplex virus-2; gE2 = glycoprotein E from herpes simplex virus-2; HSV-2 = herpes simplex virus-2; DNA = deoxyribonucleic acid; RNA = ribonucleic acid. [0024] FIGS.14A-14I show transfection rates and expression levels in HEK293T cells transfected with RNA encoding HSV-2 gC (gC2), HSV-2 gD (gD2) and HSV-2 gE (gE2) antigens. Cells were transfected with 0.2 μg/mL modRNA encoding antigens using a commercial transfection reagent. Expression of gC2, gD2 and gE2 protein was detected by flow cytometry using primary monoclonal mouse antibodies detecting the respective antigen and a secondary fluorescent tagged anti-mouse antibody. Representative data from one experiment showing percentage of gC2 (FIG.14A), gD2 (FIG.14D) and gE2 (FIG.14G) protein-expressing cells and median fluorescence intensities (MFI) (FIG.14B, FIG.14E, and FIG.14H respectively) of the total HEK293T population are depicted per antigen. Data shown are mean+SD of HEK293T transfections performed in triplicates. Cumulative total HEK expression data from up to n=3 (FIG.14C), n=4 (FIG.14F) or n=7 (FIG.14I) experiments are shown in relation to the gC2 (1600, IL2 secretory signal and HSV-2 gC antigen (version 2)), gD2 (1601, HSV-2 gD secretory signal and HSV-2 gD antigen (version 2)) or gE2 (1602, IL2 secretory signal and HSV-2 gE antigen (version 2)) construct, respectively. 2138: HSV-2 gE secretory signal and HSV-2 gC antigen (version 4), 2140: HSV-1 gD secretory signal and HSV-2 gC antigen (version 4), 2141: HSV-1 gB secretory signal and HSV-2 gC antigen (version 4), 1876: IL2 secretory signal and HSV-2 gC antigen (version 3). 1874: HSV-2 gD secretory signal and HSV-2 gD antigen (version 1), 1877: HSV-2 gD secretory signal and HSV-2 gD antigen (version 3), 1659: HSV-2 gD secretory signal and HSV-2 gD antigen (version 2), 1660: HSV-2 gD secretory signal and HSV-2 gE antigen (version 2), 2143: HSV-1 gD secretory signal and HSV-2 gE antigen (version 4). [0025] FIGS.15A-15F show transfection rates and expression levels in HEK293T cells transfected with RNA encoding HSV-2 gC (gC2), HSV-2 gD (gD2) and HSV-2 gE (gE2) antigens. Cells were transfected with 0.2 μg/mL modRNA encoding antigens using a commercial transfection reagent. Expression of gC2, gD2 and gE2 protein was detected by flow cytometry using primary monoclonal mouse antibodies detecting the respective antigen and a secondary fluorescent tagged anti-mouse antibody. Representative data from one experiment showing percentage of gC2 (FIG.15A) and gE2 (FIG.15B) protein-expressing cells and median fluorescence intensities of gC2, gD2 and gE2 (MFI) (FIG.15C, FIG.15D, and FIG.15E respectively) of the total HEK293T population are depicted per antigen. Data shown are mean+SD of HEK293T transfections performed in triplicates. Cumulative total HEK expression data from up to n=7 (FIG.15F) experiments are shown in relation to the gC2 (1600, IL2 secretory signal and HSV-2 gC antigen (version 2)), gD2 (1601, HSV-2 gD secretory signal and HSV-2 gD antigen (version 2)) or gE2 (1602, IL2 secretory signal and HSV-2 gE antigen (version 2)) construct, respectively. The RNA constructs characterized in FIG.15 were found to produce similar or improved expression as compared to 1600 (IL2 secretory signal and HSV-2 gC antigen (version 2)), 1601 (HSV-2 gD secretory signal and HSV-2 gD antigen (version 2)) and 1602 (IL2 secretory signal and HSV-2 gE antigen (version 2)). 1597: IL2 secretory signal and HSV-2 gC antigen; 1598: HSV-2 gD secretory signal and HSV-2 gD antigen; 1599: IL2 secretory signal and HSV-2 gE antigen. [0026] FIG.16. depicts conformational changes in HSV glycoprotein B (gB). Cell entry of HSV is achieved via viral proteins that mediates fution with the host membrane by substantial structural rearrangements of viral proteins, including gB, from a metastable prefusion conformation to a stable postfusion conformation. [0027] FIG.17 depicts conservation scores determined for amino acids located at positions along an UL27 consensus sequence. The UL27 open reading frame encodes HSV gB. For this analysis, complete HSV-1 and HSV-2 genomes were downloaded from VIPR database, and HSV-1 strain 17 and HSV-2 strain HG52 were used as reference strains for HSV-1 and HSV-2 respectively. [0028] FIGS.18A-18D depict four HSV T-cell string polypeptide constructs, referred to as A) RNA construct 1 (Het 1), B) RNA construct 3 (Het 3), C) RNA construct 5 (Het 5), and D) RNA construct 7 (Het 7). RNA construct 1 (Het 1) includes RL2, RL2, RS1 and UL54 T cell antigenic fragments. RNA construct 3 (Het 3) includes UL29, UL39, UL49, and UL9 T cell antigenic fragments. RNA construct 5 (Het 5) includes UL30, UL40, UL5, and UL52 T cell antigenic fragments. RNA construct 7 (Het 7) includes UL1, UL19, UL21, UL27, UL46, UL47, UL25 and UL48 T cell antigenic fragments. [0029] FIGS.19A-19B depict HSV proteins split by kinetics (A) or all proteins together (B) in cells infected with HSV-2. [0030] FIG.20 shows flow diagrams of Part A and Part B of Example 34. The flow diagrams show an overview of the protocol for safety and immunigenicity of an HSV RNA-based composition as three IM administrations at Visits 1, 4, and 7 in healthy subjects. DL = dose level; HSV = Herpes simplex virus; IM = intramucular; P = placebo (isotonic NaCl solution); V = BNT163 composition. [0031] FIG.21 shows flow diagram of Part C of Example 34. The flow diagram shows an overview of the protocol for safety and immunogenicity of an HSV RNA-based composition as two IM administrations in subjects with a history of genital herpes. Syringe icons represent composition administration. Swab icons represent 28 day twice daily anogenital swabbing periods with daily symptom diary. Abbreviations: HSV = Herpes simplex virus; IM = intramuscular; Wk = week. [0032] FIG.22 shows a dose escalating schema for Part A. Abbreviations: d = day; DL = dose level. [0033] FIG.23 depicts four HSV antigen constructs referred to as A) RNA construct 1 (Het 1), B) RNA construct 4 (Het 4), C) RNA construct 6 (Het 6), and D) RNA construct 8 (Het 8). RNA construct 1 (Het 1) includes RL2, RL2, RS1 and UL54 T cell antigenic fragments. RNA construct 4 (Het 4) includes UL9, UL49, UL39, and UL29 T cell antigenic fragments. RNA construct 6 (Het 6) includes UL52, UL5.1, UL5.2, UL40, UL30.1, and T cell antigenic fragments. RNA construct 8 (Het 8) includes UL48, UL25, UL47, UL46, UL27.1, UL27.2, UL21, UL19, and UL1 T cell antigenic fragments. HLA-I peptides were detected by mass spectrometry. [0034] FIGS.24A-24B depict a graphic overview of all HLA-I epitopes A) and specific A*02:01 epitopes B) in response to RNA construct 1 (Het 1) (SEQ ID NO: 619). [0035] FIGS.25A-25B depict a graphic overview of all HLA-I epitopes A) and specific A*02:01 epitopes B) in response to RNA construct 4 (Het 4) (SEQ ID NO: 622). [0036] FIGS.26A-26B depict a graphic overview of all HLA-I epitopes A) and specific A*02:01 epitopes B) in response to RNA construct 6 (Het 6) (SEQ ID NO: 624). [0037] FIGS.27A-27B depict a graphic overview of all HLA-I epitopes A) and specific A*02:01 epitopes B) in response to RNA construct 8 (Het 8) (SEQ ID NO: 626). [0038] FIGS.28A-28B depict antigen specific T cell response in mice. A) data is compared to vehicle control; B) data is compared to DMSO. Mice were immunized with 1 ug of each of RNA construct 1 (Het 1), RNA construct 4 (Het 4), RNA construct 6 (Het 6), and RNA construct 8 (Het 8) (Group 5 in Table 23 Example 37). [0039] FIGS.29A-29B depict antigen specific T cell response in mice. A) data is compared to vehicle control B) data is compared to DMSO. Mice were immunized with 1 ug of each of RNA construct 1 (Het 1) and RNA construct 8 (Het 8) (Group 6 in Table 23 Example 37). [0040] FIGS.30A-30B depict antigen specific T cell response in mice. A) data is compared to vehicle control; B) data is compared to DMSO. Mice were immunized with 1 ug of each of RNA construct 1 (Het 1), RNA construct 4 (Het 4), and RNA construct 6 (Het 6) (Group 7 in Table 23 Example 37). [0041] FIG.31 depicts antigen specific T cell response in mice. Mice received saline and data is compared to DMSO. [0042] FIG.32 depicts antigen specific T cell response for each T-cell antigenic fragment. Fragments were classified as having a low curated score if the fragments resulted in <100 spots per 1x106 cells. Fragments were classified as having a moderate curated score if the fragments resulted in 100-400 spots per 1x106 cells. Fragment were classified as having a moderate curated score if the fragments resulted in >400 spots per 1x106 cells. [0043] FIGS.33A-33B depict T-cell response to the antigenic fragments. A) non-log data. B) log data. Mice were immunized with 1 ug of each of RNA construct 1 (Het 1), RNA construct 4 (Het 4), RNA construct 6 (Het 6), and RNA construct 8 (Het 8) (Group 5; total 4 ug, 1 ug of each RNA construct). [0044] FIGS.34A-34B depict T-cell response to the antigenic fragments. A) non-log data. B) log data. Mice were immunized with 1 ug of each of RNA construct 1 (Het 1), RNA construct 4 (Het 4), and RNA construct 8 (Het 8) (Group 7; total 3 ug, 1 ug of each RNA construct). [0045] FIGS.35A-35B depict T-cell response to the antigenic fragments. A) non-log data. B) log data. Mice were immunized with 1 ug of each of RNA construct 1 (Het 1), and RNA construct 8 (Het 8) (Group 6; total 2 ug, 1 ug of each RNA construct). [0046] FIGS.36A-36B depict A) polyfunctional CD4 T- cell response and B) polyfunctional CD8 T cell response in mice immunized with 1 ug of each of RNA construct 1 (Het 1), RNA construct 4 (Het 4), RNA construct 6 (Het 6), and RNA construct 8 (Het 8) (Group 5; total 4 ug, 1 ug of each RNA construct). [0047] FIGS.37A-37D depict A) UL54, B) UL29, C) UL40, and D) UL47-specific polyfunctional CD8 T- cell response. [0048] FIG.38 shows an exemplary study design. Immunized mice (Day 0 and Day 21) were injected with medroxyprogesterone (subcutaneous injection 2mg/mouse at Day 56) and challenged intravaginally with HSV-2 strain MS (5×105 PFU (25 LD50)) at Day 63 and monitored for survival, genital disease scoring and vaginal virus titers at days 2 and 4. [0049] FIG.39 depicts female A02 mice bodyweight 0-12 days post intravaginally challenge with HSV-2 strain MS (5×105 PFU (25 LD50)). [0050] FIG.40 depicts survival curve for female A02 mice 0-12 days post intravaginally challenge with HSV-2 strain MS (5×105 PFU (25 LD50)). [0051] FIGS.41A-41C depict survival curve of female A02 mice immunized with individual RNA constructs. Mice received A) RNA construct 1 (Het 1) or BNT163; B) RNA construct 4 (Het 4) or RNA construct 6 (Het 6); or C) RNA construct 8 (Het 8. BNT163 was used a control: BNT163 is a trivalent composition comprising RNA encoding a glycoprotein C (gC) antigenic fragment, a glycoprotein D (gD) antigenic fragment, and a glycoprotein E (gE) antigenic fragment. [0052] FIGS.42A-42C depict survival curve of female A02 mice immunized with a combination of RNA constructs. Mice received A) combination of RNA construct 1 (Het 1) an immediate early (IE) string and RNA constructs 4 (Het 4) and 6 (Het 6) both early strings; B) combination of RNA construct 4 (Het 4) an immediate early and RNA construct 8 (Het 8) a late string; or C) all four constructs: RNA construct 1 (Het 1), RNA construct 4 (Het 4), RNA construct 6 (Het 6), and RNA construct 8 (Het 8) (total 4 ug, 1 ug of each string). [0053] FIGS.43A-43B depict survival curves. A) survival curve using criteria including clinical readouts such as hindleg paralysis and B) survival curve using criteria excluding hindleg paralysis as a clinical readout for female A02 mice 0-12 days post intravaginally challenge with HSV-2 strain MS (1x106 PFU HSV-2 (500 LD50)) immunized with RNA construct according to Table 23. [0054] FIGS.44A-44B depict A) survival curve using criteria including clinical readouts such as hindleg paralysis and B) survival curve using criteria excluding hindleg paralysis as a clinical readout for female A02 mice 0- 12 days post intravaginally challenge with HSV-2 strain MS (1x106 PFU HSV-2 (500 LD50)) immunized with all four construct: RNA construct 1 (Het 1), RNA construct 4 (Het 4), RNA construct 6 (Het 6), and RNA construct 8 (Het 8) (total 4 ug, 1 ug of each RNA construct). [0055] FIGS.45A-45B depict cumulative survival days for A) individual RNA constructs; RNA construct 1 (Het 1), RNA construct 4 (Het 4), RNA construct 6 (Het 6), or RNA construct 8 (Het 8) and B) combination of constructs. BNT163 was used a control: BNT163 is a trivalent composition comprising RNA encoding a glycoprotein C (gC) antigenic fragment, a glycoprotein D (gD) antigenic fragment, and a glycoprotein E (gE) antigenic fragment. [0056] FIGS.46A-46B depict cumulative survival days for A) all construct and combination of constructs and B) constructs and combination of constructs showing best and worst cumulative survival. BNT163 was used a control: BNT163 is a trivalent composition comprising RNA encoding a glycoprotein C (gC) antigenic fragment, a glycoprotein D (gD) antigenic fragment, and a glycoprotein E (gE) antigenic fragment. [0057] FIGS.47A-47D depict vaginal titers using A+B) plaque assay or C+D) qPCR at A+C) 2 days after challenge or B+D) 4 days after challenge. [0058] FIG.48 depicts ten HSV antigen constructs: RNA construct 15 (Het 15); RNA construct 16 (Het 16); RNA construct 17 (Het 17); RNA construct 18 (Het 18); RNA construct 19 (Het 19); RNA construct 20 (Het 20); RNA construct 21 (Het 21); RNA construct 22 (Het 22); RNA construct 23 (Het 23); RNA construct 24 (Het 4). RNA construct 15 and RNA construct 16 are immediate early and late constructs and include RL2.1, UL54, UL47, UL46 and UL21 T cell antigenic fragments. RNA construct 17 and RNA construct 18 are early constructs and include UL29, UL39, UL9, and UL5.1, UL40, and UL30.1 T cell antigenic fragments. RNA construct 19 and RNA construct 20 are immediate early and early constructs and include UL2.1, UL54, UL9, UL39, and UL5.1 T cell antigenic fragments. RNA construct 21 and RNA construct 22 are late and early constructs and include UL47, UL46, UL21, UL5.2, UL40, UL30.1, and UL29 T cell antigenic fragments. RNA construct 23 and RNA construct 24 include UL2.1, U54, UL5.2., UL40, UL47, and UL46 cell antigenic fragments. [0059] FIGS.49A-49E depict protein expression of the RNA constructs shown in FIG.80 by pair with proteasome inhibitor. [0060] FIGS.50A-50E depict protein expression of the RNA constructs shown in FIG.80 by pair without proteasome inhibitor. [0061] FIGS.51A-51B depict antigen specific T cell response in A02 mice under A) loose saturation (<220 spots per 1x106 cells) and B) under stringent saturation (<100 spots per 1x106 cells). Mice were immunized with 2 ug of RNA construct 15 (Het 15) and 2 ug of RNA construct 17 (Het 17) (Group 1 in Table 24 Example 39). [0062] FIGS.52A-52B depict antigen specific T cell response in A02 mice under A) loose saturation (<220 spots per 1x106 cells) and B) under stringent saturation (<100 spots per 1x106 cells). Mice were immunized with 2 ug of RNA construct 20 (Het 20) and 2 ug of RNA construct 22 (Het 22) (Group 2 in Table 24 Example 39). [0063] FIGS.53A-53B depict antigen specific T cell response in A02 mice under A) loose saturation (<220 spots per 1x106 cells) and B) under stringent saturation (<100 spots per 1x106 cells). Mice were immunized with 4 ug of RNA construct 23 (Het 23) and 2 ug of RNA construct 22 (Het 22) (Group 3 in Table 24 Example 39). [0064] FIG.54 depicts antigen specific T cell response in Balb/c mice immunized with 2 ug of RNA construct 15 (Het 15) and 2 ug of RNA construct 17 (Het 17) (Group 1 in Table 24 Example 39). [0065] FIG.55 depicts antigen specific T cell response in Balb/c mice immunized with 2 ug of RNA construct 20 (Het 20) and 2 ug of RNA construct 22 (Het 22) (Group 2 in Table 24 Example 39). [0066] FIG.56 depicts antigen specific T cell response in Balb/c mice immunized with 4 ug of RNA construct 23 (Het 23) (Group 3 in Table 24 Example 39). [0067] FIGS.57A-57C depict CD4 and CD8 T-cell responses in mice immunized with 2 ug of RNA construct 15 (Het 15) and 2 ug of RNA construct 17 (Het 17) (Group 1 in Table 24 Example 39). A) total response; B) CD4 depleted and shows CD8 response; and C) CD8 depleted and shows CD4 response. [0068] FIGS.58A-58B depict CD4 and CD8 T-cell responses in mice immunized with 2 ug of RNA construct 15 (Het 15) and 2 ug of RNA construct 17 (Het 17) (Group 1 in Table 24 Example 39). A) CD8 vs. CD4 response; and B) CD8 to CD4 ratio vs. total responses. [0069] FIGS.59A-59C depict CD4 and CD8 T-cell response in mice immunized with 4 ug of RNA construct 23 (Het 23) (Group 3 in Table 24 Example 39). A) total response; B) CD4 depleted and shows CD8 response; and C) CD8 depleted and shows CD4 response. [0070] FIGS.60A-60B depict CD4 and CD8 T-cell response in mice immunized with 4 ug of RNA construct 23 (Het 23) (Group 3 in Table 24 Example 39). A) CD8 vs. CD4 response; and B) CD8 to CD4 ratio vs. total responses. [0071] FIG.61 Immunized mice (Day 0 and Day 21) were injected with medroxyprogesterone (subcutaneous injection 2mg/mouse at Day 46) and challenged intravaginally with HSV-2 strain MS (5x103 PFU HSV- 2 (~10xLD50)) at Day 51 and monitored for survival, genital disease scoring, weight and vaginal virus titers at 6-h, days 2, 4 and 7. [0072] FIGS.62A-62E depict female mice bodyweight 0-16 days post intravaginally challenge with HSV- 2 strain MS (5x103 PFU HSV-2 (~10xLD50)). Mice were administered with A) PBS (Group 1 in Table 25 Example 40); B) 4 ug of RNA construct 23 (Het 23) (Group 2 in Table 25 Example 40); C) with 2 ug of RNA construct 15 (Het 15) and 2 ug of RNA construct 17 (Het 17) (Group 3 in Table 25 Example 40); D) a trivalent (BNT163) composition comprising RNA encoding a glycoprotein C (gC) antigenic fragment, a glycoprotein D (gD) antigenic fragment, and a glycoprotein E (gE) antigenic fragment; and E) combined graph. [0073] FIG.63 depicts survival for female mice 0-15 days post intravaginally challenge with HSV-2 strain MS (5x103 PFU HSV-2 (~10xLD50)). [0074] FIGS.64A-64B depict effect of RNA construct immunization on vaginal disease following HSV-2 infection. [0075] FIG.65 depicts vaginal HSV-2 replication kinetics over 7 days. [0076] FIG.66 depicts cumulative survival for female mice 0-15 days post intravaginally challenge with HSV-2 strain MS (5x103 PFU HSV-2 (~10xLD50)). [0077] FIGS.67A-67B depict the effect of RNA construct immunizations on HSV-2 replication in mice vaginal cavity. A) day 2 post challenge; and B) day 4 post challenge. TCS-23 = RNA construct 23 (Het 23); TCS- 15+17 = RNA constructs 15+17 (Het 15+ 17); and BNT163 = a trivalent composition comprising RNA encoding a glycoprotein C (gC) antigenic fragment, a glycoprotein D (gD) antigenic fragment, and a glycoprotein E (gE) antigenic fragment. [0078] FIGS.68A-68B depict the effect of RNA construct immunizations on HSV-2 replication in mice vaginal cavity. A) day 6-h post challenge; and B) day 7 post challenge. TCS-23 = RNA construct 23 (Het 23); TCS- 15+17 = RNA constructs 15+17 (Het 15+ 17); and BNT163 = a trivalent composition comprising RNA encoding a glycoprotein C (gC) antigenic fragment, a glycoprotein D (gD) antigenic fragment, and a glycoprotein E (gE) antigenic fragment. [0079] FIG.69 depicts female mice bodyweight 0-16 days post intravaginally challenge with HSV-2 strain MS (5x103 PFU HSV-2 (~10xLD50)). BNT163 = a trivalent composition comprising RNA encoding a glycoprotein C (gC) antigenic fragment, a glycoprotein D (gD) antigenic fragment, and a glycoprotein E (gE) antigenic fragment, . TCS-23 = RNA construct 23 (Het 23); TCS 15+17 = RNA constructs 15+17 (Het 15+ 17). [0080] FIG.70 depicts survival for female mice 0-15 days post intravaginally challenge with HSV-2 strain MS (5x103 PFU HSV-2 (~10xLD50)). [0081] FIGS.71A-71B depict effect of RNA construct immunization on vaginal disease following HSV-2 infection. [0082] FIGS.72A-72D depict the effect of RNA construct immunizations on HSV-2 replication kinetics in mice vaginal cavity. A) Day 6-h post challenge; and B) Day 2 post challenge. C) Day 4 post challenge. D) Day 7 post challenge. BNT163 = a trivalent composition comprising RNA encoding a glycoprotein C (gC) antigenic fragment, a glycoprotein D (gD) antigenic fragment, and a glycoprotein E (gE) antigenic fragment;TCS-23 = RNA construct 23 (Het 23); and TCS 15+17 = RNA constructs 15+17 (Het 15+ 17). [0083] FIGS.73A-73F depict expression levels in HEK293T cells transfected with RNA encoding HSV-2 gC (gC2), gD (gD2) and gE (gE2) antigens. [0084] FIGS.74A-74F depict expression levels in HEK293T cells transfected with RNA encoding HSV-2 gC (gC2) antigens. [0085] FIGS.75A-75B depict expression levels in HEK293T cells transfected with RNA encoding HSV-2 gD (gD2) antigens. [0086] FIGS.76A-76D depict expression levels in HEK293T cells transfected with RNA encoding HSV-2 gE (gE2) antigens. [0087] FIGS.77A-77C depict secretion levels of HEK293T cells transfected with RNA encoding HSV-2 encoding HSV-2 gC (gC2), gD (gD2) and gE (gE2) antigens. [0088] FIG.78 shows cumulative recurrent genital lesion days per group. Previously infected guinea pigs were immunized on days 35 and 65 post-infection with nucleoside-modified RNA encapsulated in lipid nanoparticle 315 and expressing an exemplary immunogenic fragment of HSV-2 gB (gB2) (30 μg) or PBS (control). Animals were scored daily Monday to Friday for recurrent genital lesions from 1 day after the first immunization until the end of the study on day 116. From the time of the second immunization, major differences appeared comparing the gB2 group with the PBS group. [0089] FIG.79 shows days with recurrent genital lesions for each animal starting 1 day after the second immunization. The same animals as in FIG.78 are shown here. P values were calculated by the two-tailed Mann Whitney test and demonstrate highly significant differences comparing the gB2 group with PBS (**, P<0.01). [0090] FIG.80 shows cytokine production by CD4+ T cell in response to compositions comprising HSV-2 glycoproteins. Mice were immunized twice as described herein with 10 μg of an exemplary immunogenic fragment of HSV-2 gB (gB2) RNA-LNP. Splenocytes from these mice were stimulated with a gB2 overlapping peptide pool. CD4+ cytokine-producing T cells were analyzed by flow cytometry. [0091] FIG.81 shows cytokine production by CD8+ T cell in response to compositions comprising HSV-2 glycoproteins. Mice were immunized twice as described herein with 10 μg of an exemplary immunogenic fragment of HSV-2 gB (gB2) mRNA-LNP. Splenocytes from these mice were stimulated with a gB2 overlapping peptide pool. CD8+ cytokine-producing T cells were analyzed by flow cytometry. [0092] FIG.82 depicts expression levels in HEK293T cells transfected with RNA encoding HSV-2 gB (gB2) antigens. [0093] FIGS.83A-83B show recurrent genital lesion days per group. Guinea pigs were immunized twice on days 25 and 65 post-infection with nucleoside modified RNAs encapsulated in a lipid nanoparticle and expressing an exemplary HSV-2 gE (gE2) immunogenic fragment and an exemplary HSV-2 gI (gI2) immunogenic fragment (15ug each), or PBS (control). Animals were scored daily Monday to Friday for recurrent genital lesions from 1 day after the first immunization until the end of the study on day 116. From the time of the second immunization, significant differences appeared comparing guinea pigs immunized with modified RNAs encoding exemplary gE2 immunogenic fragment and an exemplary gI2 immunogenic fragment, with the PBS group. FIG.83A shows the cumulative recurrent genital lesion days per group. FIG.83B shows days with recurrent genital lesions for each animal starting 1 day after the second immunization. P values were calculated by the two-tailed Mann Whitney test and demonstrate highly significant differences comparing gE2/gI2 with PBS (**, P<0.01; ns, P value not significant). [0094] FIGS.84A-84D show T cell responses to gE2 stimulation in mice immunized with gE2/gI2 bivalent RNA composition in CD4+ T cells (FIG.84A) and in CD8+ T cells (FIG.84B); and T cell responses to gI2 stimulation in mice immunized with E2/gI2 bivalent RNA composition in CD4+ T cells (FIG.84C) and in CD8+ T cells (FIG.84D). [0095] FIG.85 shows antibody responses to gE2/gI2 in mice immunized with gE2 RNA composition. [0096] FIGS.86A-86C show antibody responses to gE2/gI2 (FIG.86A), gE2 (FIG.86B), and gI2 (FIG. 86C) in mice immunized with gE2/gI2 bivalent RNA composition. [0097] FIG.87 shows a scheme of the protocol of the mice model experiments described in Example 51 [0098] FIGS.88A-88C show survival (FIG.88A), weight loss (FIG.88B), and disease score (FIG.88C) in a mouse model of HSV-2 in mice immunized with gE2/gI2 bivalent RNA [0099] FIGS.89A-89C show HSV virus titer 2 days (FIG.89A) and 4 days (FIG.89B) after infection in mice immunized with gE2/gI2 bivalent; as well as HSV-2 DNA copy number in DRG 28 days after infection (FIG. 89C). [0100] FIGS.91A-91D show the prophylactic effect of BNT163, gB, gE2/gI2, or a combination of them on survival (FIG.91A), disease severity (FIG.91B), genital lesions (FIG.91C), and urinary retention (FIG.91D) in guinea pigs infected with HSV-2. [0101] FIGS.92A-92B show the therapeutic effect of BNT163, gB and gE2/gI2 on recurrent lesions in a first (FIG.92A) and the combined results of two experiments (FIG.92B) experiments. CERTAIN DEFINITIONS [0102] In general, terminology used herein is in accordance with its understood meaning in the art, unless clearly indicated otherwise. Explicit definitions of certain terms are provided below; meanings of these and other terms in particular instances throughout this specification will be clear to those skilled in the art from context. [0103] In order that the present invention may be more readily understood, certain terms are first defined below. Additional definitions for the following terms and other terms are set forth throughout the specification. [0104] About: The term “about”, when used herein in reference to a value, refers to a value that is similar, in context to the referenced value. In general, those skilled in the art, familiar with the context, will appreciate the relevant degree of variance encompassed by “about” in that context. For example, in some embodiments, the term “about” may encompass a range of values that within 25%, 20%, 19%, 18%, 17%, 16%, 15%, 14%, 13%, 12%, 11%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, or less of the referred value. [0105] Agent: As used herein, the term “agent”, may refer to a physical entity or phenomenon. In some embodiments, an agent may be characterized by a particular feature and/or effect. In some embodiments, an agent may be a compound, molecule, or entity of any chemical class including, for example, a small molecule, polypeptide, nucleic acid, saccharide, lipid, metal, or a combination or complex thereof. In some embodiments, the term “agent” may refer to a compound, molecule, or entity that comprises a polymer. In some embodiments, the term may refer to a compound or entity that comprises one or more polymeric moieties. In some embodiments, the term “agent” may refer to a compound, molecule, or entity that is substantially free of a particular polymer or polymeric moiety. In some embodiments, the term may refer to a compound, molecule, or entity that lacks or is substantially free of any polymer or polymeric moiety. [0106] Amino acid: In its broadest sense, as used herein, the term “amino acid” refers to a compound and/or substance that can be, is, or has been incorporated into a polypeptide chain, e.g., through formation of one or more peptide bonds. In some embodiments, an amino acid has the general structure H2N–C(H)(R)–COOH. In some embodiments, an amino acid is a naturally-occurring amino acid. In some embodiments, an amino acid is a non-natural amino acid; in some embodiments, an amino acid is a D-amino acid; in some embodiments, an amino acid is an L-amino acid. “Standard amino acid” refers to any of the twenty standard L-amino acids commonly found in naturally occurring peptides. “Nonstandard amino acid” refers to any amino acid, other than the standard amino acids, regardless of whether it is prepared synthetically or obtained from a natural source. In some embodiments, an amino acid, including a carboxy- and/or amino-terminal amino acid in a polypeptide, can contain a structural modification as compared with the general structure above. For example, in some embodiments, an amino acid may be modified by methylation, amidation, acetylation, pegylation, glycosylation, phosphorylation, and/or substitution (e.g., of the amino group, the carboxylic acid group, one or more protons, and/or the hydroxyl group) as compared with the general structure. In some embodiments, such modification may, for example, alter the circulating half-life of a polypeptide containing the modified amino acid as compared with one containing an otherwise identical unmodified amino acid. In some embodiments, such modification does not significantly alter a relevant activity of a polypeptide containing the modified amino acid, as compared with one containing an otherwise identical unmodified amino acid. As will be clear from context, in some embodiments, the term “amino acid” may be used to refer to a free amino acid; in some embodiments it may be used to refer to an amino acid residue of a polypeptide. [0107] Antibody agent: As used herein, the term “antibody agent” refers to an agent that specifically binds to a particular antigen. In some embodiments, the term encompasses a polypeptide or polypeptide complex that includes immunoglobulin structural elements sufficient to confer specific binding. For example, in some embodiments, an antibody agent is or comprises a polypeptide whose amino acid sequence includes one or more structural elements recognized by those skilled in the art as a complementarity determining region (CDR); in some embodiments an antibody agent is or comprises a polypeptide whose amino acid sequence includes at least one CDR (e.g., at least one heavy chain CDR and/or at least one light chain CDR) that is substantially identical to one found in a reference antibody. In some embodiments an included CDR is substantially identical to a reference CDR in that it is either identical in sequence or contains between 1-5 amino acid substitutions as compared with the reference CDR. In some embodiments an included CDR is substantially identical to a reference CDR in that it shows at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with the reference CDR. In some embodiments an included CDR is substantially identical to a reference CDR in that it shows at least 96%, 96%, 97%, 98%, 99%, or 100% sequence identity with the reference CDR. In some embodiments an included CDR is substantially identical to a reference CDR in that at least one amino acid within the included CDR is deleted, added, or substituted as compared with the reference CDR but the included CDR has an amino acid sequence that is otherwise identical with that of the reference CDR. In some embodiments an included CDR is substantially identical to a reference CDR in that 1-5 amino acids within the included CDR are deleted, added, or substituted as compared with the reference CDR but the included CDR has an amino acid sequence that is otherwise identical to the reference CDR. In some embodiments an included CDR is substantially identical to a reference CDR in that at least one amino acid within the included CDR is substituted as compared with the reference CDR but the included CDR has an amino acid sequence that is otherwise identical with that of the reference CDR. In some embodiments an included CDR is substantially identical to a reference CDR in that 1-5 amino acids within the included CDR are deleted, added, or substituted as compared with the reference CDR but the included CDR has an amino acid sequence that is otherwise identical to the reference CDR. In some embodiments, an antibody agent is or comprises a polypeptide whose amino acid sequence includes structural elements recognized by those skilled in the art as an immunoglobulin variable domain. In some embodiments, an antibody agent in or comprises a polypeptide whose amino acid sequence includes structural elements recognized by those skilled in the art to correspond to CDRs1, 2, and 3 of an antibody variable domain; in some such embodiments, an antibody agent in or comprises a polypeptide or set of polypeptides whose amino acid sequence(s) together include structural elements recognized by those skilled in the art to correspond to both heavy chain and light chain variable region CDRs, e.g., heavy chain CDRs 1, 2, and/or 3 and light chain CDRs 1, 2, and/or 3. In some embodiments, an antibody agent is a polypeptide protein having a binding domain which is homologous or largely homologous to an immunoglobulin-binding domain. In some embodiments, an antibody agent may be or comprise a polyclonal antibody preparation. In some embodiments, an antibody agent may be or comprise a monoclonal antibody preparation. In some embodiments, an antibody agent may include one or more constant region sequences that are characteristic of a particular organism, such as a camel, human, mouse, primate, rabbit, rat; in many embodiments, an antibody agent may include one or more constant region sequences that are characteristic of a human. In some embodiments, an antibody agent may include one or more sequence elements that would be recognized by one skilled in the art as a humanized sequence, a primatized sequence, a chimeric sequence, etc. In some embodiments, an antibody agent may be a canonical antibody (e.g., may comprise two heavy chains and two light chains). In some embodiments, an antibody agent may be in a format selected from, but not limited to, intact IgA, IgG, IgE or IgM antibodies; bi- or multi- specific antibodies (e.g., Zybodies®, etc); antibody fragments such as Fab fragments, Fab’ fragments, F(ab’)2 fragments, Fd’ fragments, Fd fragments, and isolated CDRs or sets thereof; single chain Fvs; polypeptide-Fc fusions; single domain antibodies (e.g., shark single domain antibodies such as IgNAR or fragments thereof); cameloid antibodies; masked antibodies (e.g., Probodies®); Small Modular ImmunoPharmaceuticals (“SMIPsTM”); single chain or Tandem diabodies (TandAb®); VHHs; Anticalins®; Nanobodies® minibodies; BiTE®s; ankyrin repeat proteins or DARPINs®; Avimers®; DARTs; TCR-like antibodies;, Adnectins®; Affilins®; Trans-bodies®; Affibodies®; TrimerX®; MicroProteins; Fynomers®, Centyrins®; and KALBITOR®s. In some embodiments, an antibody may lack a covalent modification (e.g., attachment of a glycan) that it would have if produced naturally. In some embodiments, an antibody may contain a covalent modification (e.g., attachment of a glycan, a payload (e.g., a detectable moiety, a therapeutic moiety, a catalytic moiety, etc.), or other pendant group (e.g., poly-ethylene glycol, etc.)). [0108] Antigen: Those skilled in the art, reading the present specification, will appreciate that the term “antigen” refers to a molecule that is recognized by the immune system, e.g., in some embodiments the adaptive immune system, such that it elicits an antigen-specific immune response. In some embodiments, an antigen-specific immune response may be or comprise generation of antibodies and/or antigen-specific T cells. In some embodiments, an antigen is a peptide or polypeptide that comprises at least one epitope against which an immune response can be generated. In one embodiment, an antigen is presented by cells of the immune system such as antigen presenting cells like dendritic cells or macrophages. In one embodiment, an antigen or a processed product thereof such as a T-cell epitope is bound by a T- or B-cell receptor, or by an immunoglobulin molecule such as an antibody. Accordingly, an antigen or a processed product thereof may react specifically with antibodies or T lymphocytes (T cells). In one embodiment, an antigen is a parasitic antigen. In accordance with the present disclosure, in some embodiments, an antigen may be delivered by RNA molecules as described herein. In some embodiments, a peptide or polypeptide antigen can be 2-100 amino acids, including for example, 5 amino acids, 10 amino acids, 15 amino acids, 20 amino acids, 25 amino acids, 30 amino acids, 35 amino acids, 40 amino acids, 45 amino acids, or 50 amino acids in length. In some embodiments, a peptide or polypeptide antigen can be greater than 50 amino acids. In some embodiments, a peptide or polypeptide antigen can be greater than 100 amino acids. In some embodiments, an antigen is recognized by an immune effector cell. In some embodiments, an antigen if recognized by an immune effector cell is able to induce in the presence of appropriate co-stimulatory signals, stimulation, priming and/or expansion of the immune effector cell carrying an antigen receptor recognizing the antigen. In the context of the embodiments of the present disclosure, in some embodiments, an antigen can be presented or present on the surface of a cell, e.g., an antigen presenting cell. In one embodiment, an antigen is presented by a diseased cell such as a virus-infected cell. In one embodiment, an antigen receptor is a TCR which binds to an epitope of an antigen presented in the context of MHC. In one embodiment, binding of a TCR when expressed by T cells and/or present on T cells to an antigen presented by cells such as antigen presenting cells results in stimulation, priming and/or expansion of said T cells. In one embodiment, binding of a TCR when expressed by T cells and/or present on T cells to an antigen presented on diseased cells results in cytolysis and/or apoptosis of the diseased cells, wherein said T cells preferably release cytotoxic factors, e.g., perforins and granzymes. [0109] Associated: Two events or entities are “associated” with one another, as that term is used herein, if the presence, level, degree, type and/or form of one is correlated with that of the other. For example, a particular entity (e.g., poypeptide, genetic signature, metabolite, microbe, etc.) is considered to be associated with a particular disease, disorder, or condition, if its presence, level and/or form correlates with incidence of, susceptibility to, severity of, stage of, etc. the disease, disorder, or condition (e.g., across a relevant population). In some embodiments, two or more entities are physically “associated” with one another if they interact, directly or indirectly, so that they are and/or remain in physical proximity with one another. In some embodiments, two or more entities that are physically associated with one another are covalently linked to one another; in some embodiments, two or more entities that are physically associated with one another are not covalently linked to one another but are non- covalently associated, for example by means of hydrogen bonds, van der Waals interaction, hydrophobic interactions, magnetism, and combinations thereof. [0110] Binding: Those skilled in the art, reading the present specification, will appreciate that the term “binding” typically refers to a non-covalent association between or among entities or moieties. In some embodiments, binding data are expressed in terms of “IC50”. As is understood in the art, IC50 is the concentration of an assessed agent in a binding assay at which 50% inhibition of binding of reference agent known to bind the relevant binding partner is observed. In some embodiments, assays are run under conditions in which the assays are run (e.g., limiting binding target and reference concentrations), these values approximate KD values. Assays for determining binding are well known in the art and are described in detail, for example, in PCT publications WO 94/20127 and WO 94/03205, and other publications such Sidney et al., Current Protocols in Immunology 18.3.1 (1998); Sidney, et al., J. Immunol. 154:247 (1995); and Sette, et al., Mol. Immunol. 31:813 (1994). Alternatively, binding can be expressed relative to binding by a reference standard peptide. For example, can be based on its IC50, relative to the IC50 of a reference standard peptide. Binding can also be determined using other assay systems including those using: live cells (e.g., Ceppellini et al., Nature 339:392 (1989); Christnick et al., Nature 352:67 (1991); Busch et al., Int. Immunol.2:443 (1990); Hill et al., J. Immunol.147:189 (1991); del Guercio et al., J. Immunol. 154:685 (1995)), cell free systems using detergent lysates (e.g., Cerundolo et al., J. Immunol 21:2069 (1991)), immobilized purified MHC (e.g., Hill et al., J. Immunol. 152, 2890 (1994); Marshall et al., J. Immunol. 152:4946 (1994)), ELISA systems (e.g., Reay et al., EMBO J. 11:2829 (1992)), surface plasmon resonance (e.g., Khilko et al., J. Biol. Chem. 268:15425 (1993)); high flux soluble phase assays (Hammer et al., J. Exp. Med. 180:2353 (1994)), and measurement of class I MHC stabilization or assembly (e.g., Ljunggren et al., Nature 346:476 (1990); Schumacher et al., Cell 62:563 (1990); Townsend et al., Cell 62:285 (1990); Parker et al., J. Immunol. 149:1896 (1992)). [0111] Cap: As used herein, the term “cap” refers to a structure comprising or essentially consisting of a nucleoside-5 '-triphosphate that is typically joined to a 5'-end of an uncapped RNA (e.g., an uncapped RNA having a 5'- diphosphate). ome embodiments, a cap is or comprises a guanine nucleotide. In some embodiments, a cap is or comprises a naturally-occurring RNA 5’ cap, including, e.g., but not limited to a 7- methylguanosine cap, which has a structure designated as “m7G.” In some embodiments, a cap is or comprises a synthetic cap analog that resembles an RNA cap structure and possesses the ability to stabilize RNA if attached thereto, including, e.g., but not limited to anti-reverse cap analogs (ARCAs) known in the art). Those skilled in the art will appreciate that methods for joining a cap to a 5’ end of an RNA are known in the art. For example, in some embodiments, a capped RNA may be obtained by in vitro capping of RNA that has a 5' triphosphate group or RNA that has a 5' diphosphate group with a capping enzyme system (including, e.g., but not limited to vaccinia capping enzyme system or Saccharomyces cerevisiae capping enzyme system). Alternatively, a capped RNA can be obtained by in vitro transcription (IVT) of a single- stranded DNA template in the presence of a dinucleotide or trinucleotide cap analog. [0112] Cell-mediated immunity: “Cell-mediated immunity,” “cellular immunity,” “cellular immune response,” or similar terms are meant to include a cellular response directed to cells characterized by expression of an antigen, in particular characterized by presentation of an antigen with class I or class II MHC. A cellular response relates to immune effector cells, in particular to T cells or T lymphocytes which act as either “helpers” or “killers.” The helper T cells (also termed CD4+ T cells or CD4 T cells) play a central role by regulating the immune response and the killer cells (also termed cytotoxic T cells, cytolytic T cells, CD8+ T cells, CD8 T cells, or CTLs) kill diseased cells such as virus-infected cells, preventing the production of more diseased cells. [0113] Co-administration: As used herein, the term “co-administration” refers to use of a pharmaceutical composition (e.g., immunogenic composition, e.g., vaccine) described herein and an additional therapeutic agent. The combined use of a pharmaceutical composition (e.g., immunogenic composition, e.g., vaccine) described herein and an additional therapeutic agent may be performed concurrently or separately (e.g., sequentially in any order). In some embodiments, a pharmaceutical composition (e.g., immunogenic composition, e.g., vaccine) described herein and an additional therapeutic agent may be combined in one pharmaceutically- acceptable carrier, or they may be placed in separate carriers and delivered to a target cell or administered to a subject at different times. Each of these situations is contemplated as falling within the meaning of “co- administration” or “combination,” provided that a pharmaceutical composition (e.g., immunogenic composition, e.g., vaccine) described herein and an additional therapeutic agent are delivered or administered sufficiently close in time that there is at least some temporal overlap in biological effect(s) generated by each on a target cell or a subject being treated. [0114] Codon-optimized: As used herein, the term “codon-optimized” refers to alteration of codons in a coding region of a nucleic acid molecule to reflect the typical codon usage of a host organism without preferably altering the amino acid sequence encoded by the nucleic acid molecule. Within the context of the present disclosure, in some embodiments coding regions are codon-optimized for optimal expression in a subject to be treated using the RNA molecules described herein. In some embodiments, codon-optimization may be performed such that codons for which frequently occurring tRNAs are available are inserted in place of “rare codons.” In some embodiments, codon- optimization may include increasing guanosine/cytosine (G/C) content of a coding region of RNA described herein as compared to the G/C content of the corresponding coding sequence of a wild type RNA, wherein the amino acid sequence encoded by the RNA is preferably not modified compared to the amino acid sequence. [0115] Combination therapy: As used herein, the term “combination therapy” refers to those situations in which a subject is simultaneously exposed to two or more therapeutic regimens (e.g., two or more therapeutic agents). In some embodiments, the two or more regimens may be administered simultaneously; in some embodiments, such regimens may be administered sequentially (e.g., all “doses” of a first regimen are administered prior to administration of any doses of a second regimen); in some embodiments, such agents are administered in overlapping dosing regimens. In some embodiments, “administration” of combination therapy may involve administration of one or more agent(s) or modality(ies) to a subject receiving the other agent(s) or modality(ies) in the combination. For clarity, combination therapy does not require that individual agents be administered together in a single composition (or even necessarily at the same time), although in some embodiments, two or more agents, or active moieties thereof, may be administered together in a combination composition. [0116] Comparable: As used herein, the term “comparable” refers to two or more agents, entities, situations, sets of conditions, etc., that may not be identical to one another but that are sufficiently similar to permit comparison there between so that one skilled in the art will appreciate that conclusions may reasonably be drawn based on differences or similarities observed. In some embodiments, comparable sets of conditions, circumstances, individuals, or populations are characterized by a plurality of substantially identical features and one or a small number of varied features. Those of ordinary skill in the art will understand, in context, what degree of identity is required in any given circumstance for two or more such agents, entities, situations, sets of conditions, etc to be considered comparable. For example, those of ordinary skill in the art will appreciate that sets of circumstances, individuals, or populations are comparable to one another when characterized by a sufficient number and type of substantially identical features to warrant a reasonable conclusion that differences in results obtained or phenomena observed under or with different sets of circumstances, individuals, or populations are caused by or indicative of the variation in those features that are varied. [0117] Corresponding to: As used herein, the term “corresponding to” refers to a relationship between two or more entities. For example, the term “corresponding to” may be used to designate the position/identity of a structural element in a compound or composition relative to another compound or composition (e.g., to an appropriate reference compound or composition). For example, in some embodiments, a monomeric residue in a polymer (e.g., an amino acid residue in a polypeptide or a nucleic acid residue in a polynucleotide) may be identified as “corresponding to” a residue in an appropriate reference polymer. For example, those of ordinary skill will appreciate that, for purposes of simplicity, residues in a polypeptide are often designated using a canonical numbering system based on a reference related polypeptide, so that an amino acid “corresponding to” a residue at position 190, for example, need not actually be the 190th amino acid in a particular amino acid chain but rather corresponds to the residue found at 190 in the reference polypeptide; those of ordinary skill in the art readily appreciate how to identify “corresponding” amino acids. For example, those skilled in the art will be aware of various sequence alignment strategies, including software programs such as, for example, BLAST, CS-BLAST, CUSASW++, DIAMOND, FASTA, GGSEARCH/GLSEARCH, Genoogle, HMMER, HHpred/HHsearch, IDF, Infernal, KLAST, USEARCH, parasail, PSI-BLAST, PSI-Search, ScalaBLAST, Sequilab, SAM, SSEARCH, SWAPHI, SWAPHI-LS, SWIMM, or SWIPE that can be utilized, for example, to identify “corresponding” residues in polypeptides and/or nucleic acids in accordance with the present disclosure. Those of skill in the art will also appreciate that, in some instances, the term “corresponding to” may be used to describe an event or entity that shares a relevant similarity with another event or entity (e.g., an appropriate reference event or entity). To give but one example, a gene or protein in one organism may be described as “corresponding to” a gene or protein from another organism in order to indicate, in some embodiments, that it plays an analogous role or performs an analogous function and/or that it shows a particular degree of sequence identity or homology, or shares a particular characteristic sequence element. [0118] Derived: In the context of an amino acid sequence (peptide or polypeptide) “derived from” a designated amino acid sequence (peptide or polypeptide), it refers to a structural analogue of a designated amino acid sequence. In some embodiments, an amino acid sequence which is derived from a particular amino acid sequence has an amino acid sequence that is identical, essentially identical or homologous to that particular sequence or a fragment thereof. Amino acid sequences derived from a particular amino acid sequence may be variants of that particular sequence or a fragment thereof. For example, it will be understood by one of ordinary skill in the art that the antigens suitable for use herein may be altered such that they vary in sequence from the naturally occurring or native sequences from which they were derived, while retaining the desirable activity of the native sequences. [0119] Designed: As used herein, the term “designed” refers to an agent (i) whose structure is or was selected by the hand of man; (ii) that is produced by a process requiring the hand of man; and/or (iii) that is distinct from natural substances and other known agents. [0120] Dosing regimen: Those skilled in the art will appreciate that the term “dosing regimen” may be used to refer to a set of unit doses (typically more than one) that are administered individually to a subject, typically separated by periods of time. In some embodiments, a given therapeutic agent has a recommended dosing regimen, which may involve one or more doses. In some embodiments, a dosing regimen comprises a plurality of doses each of which is separated in time from other doses. In some embodiments, individual doses are separated from one another by a time period of the same length; in some embodiments, a dosing regimen comprises a plurality of doses and at least two different time periods separating individual doses. In some embodiments, all doses within a dosing regimen are of the same unit dose amount. In some embodiments, different doses within a dosing regimen are of different amounts. In some embodiments, a dosing regimen comprises a first dose in a first dose amount, followed by one or more additional doses in a second dose amount different from the first dose amount. In some embodiments, a dosing regimen comprises a first dose in a first dose amount, followed by one or more additional doses in a second dose amount same as the first dose amount. In some embodiments, a dosing regimen is correlated with a desired or beneficial outcome when administered across a relevant population (i.e., is a therapeutic dosing regimen). [0121] Encode: As used herein, the term “encode” or “encoding” refers to sequence information of a first molecule that guides production of a second molecule having a defined sequence of nucleotides (e.g., mRNA) or a defined sequence of amino acids. For example, a DNA molecule can encode an RNA molecule (e.g., by a transcription process that includes a DNA-dependent RNA polymerase enzyme). An RNA molecule can encode a polypeptide (e.g., by a translation process). Thus, a gene, a cDNA, or an RNA molecule (e.g., an mRNA) encodes a polypeptide if transcription and translation of RNA corresponding to that gene produces the polypeptide in a cell or other biological system. In some embodiments, a coding region of an RNA molecule encoding a target antigen refers to a coding strand, the nucleotide sequence of which is identical to the RNA sequence of such a target antigen. In some embodiments, a coding region of an RNA molecule encoding a target antigen refers to a non-coding strand of such a target antigen, which may be used as a template for transcription of a gene or cDNA. [0122] Engineered: In general, the term “engineered” refers to the aspect of having been manipulated by the hand of man. For example, a polynucleotide is considered to be “engineered” when two or more sequences that are not linked together in that order in nature are manipulated by the hand of man to be directly linked to one another in the engineered polynucleotide and/or when a particular residue in a polynucleotide is non-naturally occurring and/or is caused through action of the hand of man to be linked with an entity or moiety with which it is not linked in nature. [0123] Epitope: As used herein, the term “epitope” refers to a moiety that is specifically recognized by an immunoglobulin (e.g., antibody or receptor) binding component. For example, an epitope may be recognized by a T cell, a B cell, or an antibody. In some embodiments, an epitope is comprised of a plurality of chemical atoms or groups on an antigen. In some embodiments, such chemical atoms or groups are surface-exposed when the antigen adopts a relevant three-dimensional conformation. In some embodiments, such chemical atoms or groups are physically near to each other in space when the antigen adopts such a conformation. In some embodiments, at least some such chemical atoms are groups are physically separated from one another when the antigen adopts an alternative conformation (e.g., is linearized). Accordingly, in some embodiments, an epitope of an antigen may include a continuous or discontinuous portion of the antigen. In some embodiments, an epitope is or comprises a T cell epitope. In some embodiments, an epitope may have a length of about 5 to about 30 amino acids, or about 10 to about 25 amino acids, or about 5 to about 15 amino acids, or about 5 to 12 amino acids, or about 6 to about 9 amino acids. [0124] Expression: As used herein, the term “expression” of a nucleic acid sequence refers to the generation of a gene product from the nucleic acid sequence. In some embodiments, a gene product can be a transcript. In some embodiments, a gene product can be a polypeptide. In some embodiments, expression of a nucleic acid sequence involves one or more of the following: (1) production of an RNA template from a DNA sequence (e.g., by transcription); (2) processing of an RNA transcript (e.g., by splicing, editing, etc.); (3) translation of an RNA into a polypeptide or protein; and/or (4) post-translational modification of a polypeptide or protein. [0125] Five prime untranslated region: As used herein, the terms “five prime untranslated region” or “5' UTR” refer to a sequence of an RNA molecule between a transcription start site and a start codon of a coding region of an RNA. In some embodiments, “5’ UTR” refers to a sequence of an RNA molecule that begins at a transcription start site and ends one nucleotide (nt) before a start codon (usually AUG) of a coding region of an RNA molecule, e.g., in its natural context. [0126] Fragment: The term “fragment” as used herein in the context of a nucleic acid sequence (e.g., RNA sequence) or an amino acid sequence may typically be a portion of a reference sequence. In some embodiments, a reference sequence is a full-length sequence of e.g., a nucleic acid sequence or an amino acid sequence. Accordingly, a fragment, typically, refers to a sequence that is identical to a corresponding stretch within a reference sequence. In some embodiments, a fragment comprises a continuous stretch of nucleotides or amino acid residues that corresponds to at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95% of the total length of a reference sequence from which the fragment is derived. In some embodiments, the term “fragment", with reference to an amino acid sequence (peptide or polypeptide), relates to a part of an amino acid sequence, e.g., a sequence which represents the amino acid sequence shortened at the N-terminus and/or C-terminus. In some embodiments, a fragment of an amino acid sequence comprises at least 6, in particular at least 8, at least 12, at least 15, at least 20, at least 30, at least 50, or at least 100 consecutive amino acids from an amino acid sequence. [0127] Homology: As used herein, the term “homology” or “homolog” refers to the overall relatedness between polynucleotide molecules (e.g., DNA molecules and/or RNA molecules) and/or between polypeptide molecules. In some embodiments, polynucleotide molecules (e.g., DNA molecules and/or RNA molecules) and/or polypeptide molecules are considered to be “homologous” to one another if their sequences are at least 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 99% identical. In some embodiments, polynucleotide molecules (e.g., DNA molecules and/or RNA molecules) and/or polypeptide molecules are considered to be “homologous” to one another if their sequences are at least 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 99% similar (e.g., containing residues with related chemical properties at corresponding positions). For example, as is well known by those of ordinary skill in the art, certain amino acids are typically classified as similar to one another as “hydrophobic” or “hydrophilic” amino acids, and/or as having “polar” or “non-polar” side chains. Substitution of one amino acid for another of the same type may often be considered a “homologous” substitution. [0128] Humoral immunity: As used herein, the term “humoral immunity” or “humoral immune response” refers to antibody production and the accessory processes that accompany it, including: Th2 activation and cytokine production, germinal center formation and isotype switching, affinity maturation and memory cell generation. It also refers to the effector functions of antibodies, which include pathogen neutralization, classical complement activation, and opsonin promotion of phagocytosis and pathogen elimination. [0129] Identity: As used herein, the term “identity” refers to the overall relatedness between polynucleotide molecules (e.g., DNA molecules and/or RNA molecules) and/or between polypeptide molecules. In some embodiments, polynucleotide molecules (e.g., DNA molecules and/or RNA molecules) and/or between polypeptide molecules are considered to be “substantially identical” to one another if their sequences are at least 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical. Calculation of the percent identity of two nucleic acid or polypeptide sequences, for example, can be performed by aligning the two sequences for optimal comparison purposes (e.g., gaps can be introduced in one or both of a first and a second sequence for optimal alignment and non-identical sequences can be disregarded for comparison purposes). In certain embodiments, the length of a sequence aligned for comparison purposes is at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or substantially 100% of the length of a reference sequence. The nucleotides at corresponding positions are then compared. When a position in the first sequence is occupied by the same residue (e.g., nucleotide or amino acid) as the corresponding position in the second sequence, then the molecules are identical at that position. The percent identity between the two sequences is a function of the number of identical positions shared by the sequences, taking into account the number of gaps, and the length of each gap, which needs to be introduced for optimal alignment of the two sequences. The comparison of sequences and determination of percent identity between two sequences can be accomplished using a mathematical algorithm. For example, the percent identity between two nucleotide sequences can be determined using the algorithm of Meyers and Miller, 1989, which has been incorporated into the ALIGN program (version 2.0). In some embodiments, nucleic acid sequence comparisons made with the ALIGN program use a PAM120 weight residue table, a gap length penalty of 12 and a gap penalty of 4. The percent identity between two nucleotide sequences can, alternatively, be determined using the GAP program in the GCG software package using an NWSgapdna.CMP matrix. [0130] Increased, Induced, or Reduced: As used herein, these terms or grammatically comparable comparative terms, indicate values that are relative to a comparable reference measurement. For example, in some embodiments, an assessed value achieved with a provided pharmaceutical composition (e.g., immunogenic composition, e.g., vaccine) may be “increased” relative to that obtained with a comparable reference pharmaceutical composition (e.g., immunogenic composition, e.g., vaccine). Alternatively or additionally, in some embodiments, an assessed value achieved in a subject may be “increased” relative to that obtained in the same subject under different conditions (e.g., prior to or after an event; or presence or absence of an event such as administration of a pharmaceutical composition (e.g., immunogenic composition, e.g., vaccine) as described herein, or in a different, comparable subject (e.g., in a comparable subject that differs from the subject of interest in prior exposure to a condition, e.g., absence of administration of a pharmaceutical composition (e.g., immunogenic composition, e.g., vaccine) as described herein.). In some embodiments, comparative terms refer to statistically relevant differences (e.g., that are of a prevalence and/or magnitude sufficient to achieve statistical relevance). Those skilled in the art will be aware, or will readily be able to determine, in a given context, a degree and/or prevalence of difference that is required or sufficient to achieve such statistical significance. In some embodiments, the term “reduced” or equivalent terms refers to a reduction in the level of an assessed value by at least 5%, at least 10%, at least 20%, at least 50%, at least 75% or higher, as compared to a comparable reference. In some embodiments, the term “reduced” or equivalent terms refers to a complete or essentially complete inhibition, i.e., a reduction to zero or essentially to zero. In some embodiments, the term “increased” or “induced” refers to an increase in the level of an assessed value by at least 10%, at least 20%, at least 30%, at least 40%, at least 50%, at least 80%, at least 100%, at least 200%, at least 500%, or higher, as compared to a comparable reference. [0131] Ionizable: The term “ionizable” refers to a compound or group or atom that is charged at a certain pH. In the context of an ionizable amino lipid, such a lipid or a function group or atom thereof bears a positive charge at a certain pH. In some embodiments, an ionizable amino lipid is positively charged at an acidic pH. In some embodiments, an ionizable amino lipid is predominately neutral at physiological pH values, e.g., in some embodiments about 7.0-7.4, but becomes positively charged at lower pH values. In some embodiments, an ionizable amino lipid may have a pKa within a range of about 5 to about 7. [0132] Isolated: The term “isolated” means altered or removed from the natural state. For example, a nucleic acid or a peptide naturally present in a living animal is not “isolated”, but the same nucleic acid or peptide partially or completely separated from the coexisting materials of its natural state is “isolated”. An isolated nucleic acid or protein can exist in substantially purified form, or can exist in a non-native environment such as, for example, a host cell. [0133] Lipid: As used herein, the terms “lipid” and “lipid-like material” are broadly defined as molecules which comprise one or more hydrophobic moieties or groups and optionally also one or more hydrophilic moieties or groups. Molecules comprising hydrophobic moieties and hydrophilic moieties are also typically denoted as amphiphiles. [0134] RNA lipid nanoparticle: As used herein, the term “RNA lipid nanoparticle” refers to a nanoparticle comprising at least one lipid and RNA molecule(s). In some embodiments, an RNA lipid nanoparticle comprises at least one ionizable amino lipid. In some embodiments, an RNA lipid nanoparticle comprises at least one ionizable amino lipid, at least one helper lipid, and at least one polymer-conjugated lipid (e.g., PEG-conjugated lipid). In various embodiments, RNA lipid nanoparticles as described herein can have an average size (e.g., Z-average) of about 100 nm to 1000 nm, or about 200 nm to 900 nm, or about 200 nm to 800 nm, or about 250 nm to about 700 nm. In some embodiments of the present disclosure, RNA lipid nanoparticles can have a particle size (e.g., Z- average) of about 30 nm to about 200 nm, or about 30 nm to about 150 nm, about 40 nm to about 150 nm, about 50 nm to about 150 nm, about 60 nm to about 130 nm, about 70 nm to about 110 nm, about 70 nm to about 100 nm, about 80 nm to about 100 nm, about 90 nm to about 100 nm, about 70 to about 90 nm, about 80 nm to about 90 nm, or about 70 nm to about 80 nm. In some embodiments, an average size of lipid nanoparticles is determined by measuring the particle diameter. In some embodiments, RNA lipid nanoparticles may be prepared by mixing lipids with RNA molecules described herein. [0135] Lipidoid: As used herein, a “lipidoid” refers to a lipid-like molecule. In some embodiments, a lipoid is an amphiphilic molecule with one or more lipid-like physical properties. In the context of the present disclosure, the term lipid is considered to encompass lipidoids. [0136] Nanoparticle: As used herein, the term “nanoparticle” refers to a particle having an average size suitable for parenteral administration. In some embodiments, a nanoparticle has a longest dimension (e.g., a diameter) of less than 1,000 nanometers (nm). In some embodiments, a nanoparticle may be characterized by a longest dimension (e.g., a diameter) of less than 300 nm. In some embodiments, a nanoparticle may be characterized by a longest dimension (e.g., a diameter) of less than 100 nm. In many embodiments, a nanoparticle may be characterized by a longest dimension between about 1 nm and about 100 nm, or between about 1 μm and about 500 nm, or between about 1 nm and 1,000 nm. In many embodiments, a population of nanoparticles is characterized by an average size (e.g., longest dimension) that is below about 1,000 nm, about 500 nm, about 100 nm, about 50 nm, about 40 nm, about 30 nm, about 20 nm, or about 10 nm and often above about 1 nm. In many embodiments, a nanoparticle may be substantially spherical so that its longest dimension may be its diameter. In some embodiments, a nanoparticle has a diameter of less than 100 nm as defined by the National Institutes of Health. [0137] Naturally occurring: The term “naturally occurring” as used herein refers to an entity that can be found in nature. For example, a peptide or nucleic acid that is present in an organism (including viruses) and can be isolated from a source in nature and which has not been intentionally modified by man in the laboratory is naturally occurring. [0138] Neutralization: As used herein, the term “neutralization” refers to an event in which binding agents such as antibodies bind to a biological active site of a virus such as a receptor binding protein, thereby inhibiting the parasitic infection of cells. In some embodiments, the term “neutralization” refers to an event in which binding agents eliminate or significantly reduce ability of infecting cells. [0139] Nucleic acid particle: A “nucleic acid particle” can be used to deliver nucleic acid to a target site of interest (e.g., cell, tissue, organ, and the like). A nucleic acid particle may comprise at least one cationic or cationically ionizable lipid or lipid-like material, at least one cationic polymer such as protamine, or a mixture thereof and nucleic acid. In some embodiments, a nucleic acid particle is a lipid nanoparticle. In some embodiments, a nucleic acid particle is a lipoplex particle. [0140] Nucleic acid/ Polynucleotide: As used herein, the term “nucleic acid” refers to a polymer of at least 10 nucleotides or more. In some embodiments, a nucleic acid is or comprises DNA. In some embodiments, a nucleic acid is or comprises RNA. In some embodiments, a nucleic acid is or comprises peptide nucleic acid (PNA). In some embodiments, a nucleic acid is or comprises a single stranded nucleic acid. In some embodiments, a nucleic acid is or comprises a double-stranded nucleic acid. In some embodiments, a nucleic acid comprises both single and double-stranded portions. In some embodiments, a nucleic acid comprises a backbone that comprises one or more phosphodiester linkages. In some embodiments, a nucleic acid comprises a backbone that comprises both phosphodiester and non-phosphodiester linkages. For example, in some embodiments, a nucleic acid may comprise a backbone that comprises one or more phosphorothioate or 5'-N-phosphoramidite linkages and/or one or more peptide bonds, e.g., as in a “peptide nucleic acid”. In some embodiments, a nucleic acid comprises one or more, or all, natural residues (e.g., adenine, cytosine, deoxyadenosine, deoxycytidine, deoxyguanosine, deoxythymidine, guanine, thymine, uracil). In some embodiments, a nucleic acid comprises on or more, or all, non-natural residues. In some embodiments, a non-natural residue comprises a nucleoside analog (e.g., 2-aminoadenosine, 2- thiothymidine, inosine, pyrrolo-pyrimidine, 3 -methyl adenosine, 5-methylcytidine, C-5 propynyl-cytidine, C-5 propynyl-uridine, 2-aminoadenosine, C5-bromouridine, C5-fluorouridine, C5-iodouridine, C5-propynyl-uridine, C5 - propynyl-cytidine, C5-methylcytidine, 2-aminoadenosine, 7-deazaadenosine, 7-deazaguanosine, 8-oxoadenosine, 8- oxoguanosine, 6-O-methylguanine, 2-thiocytidine, methylated bases, intercalated bases, and combinations thereof). In some embodiments, a non-natural residue comprises one or more modified sugars (e.g., 2'-fluororibose, ribose, 2'-deoxyribose, arabinose, and hexose) as compared to those in natural residues. In some embodiments, a nucleic acid has a nucleotide sequence that encodes a functional gene product such as an RNA or polypeptide. In some embodiments, a nucleic acid has a nucleotide sequence that comprises one or more introns. In some embodiments, a nucleic acid may be prepared by isolation from a natural source, enzymatic synthesis (e.g., by polymerization based on a complementary template, e.g., in vivo or in vitro, reproduction in a recombinant cell or system, or chemical synthesis. In some embodiments, a nucleic acid is at least 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 225, 250, 275, 300, 325, 350, 375, 400, 425, 450, 475, 500, 600, 700, 800, 900, 1000, 1500, 2000, 2500, 3000, 3500, 4000, 4500, 5000, 5500, 6000, 6500, 7000, 7500, 8000, 8500, 9000, 9500, 10,000, 10,500, 11,000, 11,500, 12,000, 12,500, 13,000, 13,500, 14,000, 14,500, 15,000, 15,500, 16,000, 16,500, 17,000, 17,500, 18,000, 18,500, 19,000, 19,500, or 20,000 or more residues or nucleotides long. [0141] Nucleotide: As used herein, the term “nucleotide” refers to its art-recognized meaning. When a number of nucleotides is used as an indication of size, e.g., of a polynucleotide, a certain number of nucleotides refers to the number of nucleotides on a single strand, e.g., of a polynucleotide. [0142] Patient: As used herein, the term “patient” refers to any organism who is suffering or at risk of a disease or disorder or condition. Typical patients include animals (e.g., mammals such as mice, rats, rabbits, non- human primates, and/or humans). In some embodiments, a patient is a human. In some embodiments, a patient is suffering from or susceptible to one or more diseases or disorders or conditions. In some embodiments, a patient displays one or more symptoms of a disease or disorder or condition. In some embodiments, a patient has been diagnosed with one or more diseases or disorders or conditions. In some embodiments, a disease or disorder or condition that is amenable to provided technologies is or includes an HSV infection. In some embodiments, a patient is receiving or has received certain therapy to diagnose and/or to treat a disease, disorder, or condition. In some embodiments, a patient is a patient suffering from or susceptible to an HSV infection. [0143] PEG-conjugated lipid: The term “PEG-conjugated lipid" refers to a molecule comprising a lipid portion and a polyethylene glycol portion. [0144] Pharmaceutical composition: As used herein, the term “pharmaceutical composition” refers to an active agent, formulated together with one or more pharmaceutically acceptable carriers. In some embodiments, active agent is present in unit dose amount appropriate for administration in a therapeutic regimen that shows a statistically significant probability of achieving a predetermined therapeutic effect when administered to a relevant population. In some embodiments, pharmaceutical compositions may be specially formulated for parenteral administration, for example, by subcutaneous, intramuscular, or intravenous injection as, for example, a sterile solution or suspension formulation. [0145] Pharmaceutically effective amount: The term “pharmaceutically effective amount” or “therapeutically effective amount” refers to the amount which achieves a desired reaction or a desired effect alone or together with further doses. In the case of the treatment of a particular disease, a desired reaction in some embodiments relates to inhibition of the course of the disease. In some embodiments, such inhibition may comprise slowing down the progress of a disease and/or interrupting or reversing the progress of the disease. In some embodiments, a desired reaction in a treatment of a disease may be or comprise delay or prevention of the onset of a disease or a condition. An effective amount of pharmaceutical compositions (e.g., immunogenic compositions, e.g., vaccines) described herein will depend, for example, on a disease or condition to be treated, the severity of such a disease or condition, individual parameters of the patient, including, e.g., age, physiological condition, size and weight, the duration of treatment, the type of an accompanying therapy (if present), the specific route of administration and similar factors. Accordingly, doses of pharmaceutical compositions (e.g., immunogenic compositions, e.g., vaccines) described herein may depend on various of such parameters. In the case that a reaction in a patient is insufficient with an initial dose, higher doses (or effectively higher doses achieved by a different, more localized route of administration) may be used. [0146] Poly(A) sequence: As used herein, the term “poly(A) sequence” or “poly-A tail” refers to an uninterrupted or interrupted sequence of adenylate residues which is typically located at the 3'-end of an RNA molecule. Poly(A) sequences are known to those of skill in the art and may follow the 3’-UTR in the RNAs described herein. An uninterrupted poly(A) sequence is characterized by consecutive adenylate residues. In nature, an uninterrupted poly(A) sequence is typical. RNAs disclosed herein can have a poly(A) sequence attached to the free 3'-end of the RNA by a template-independent RNA polymerase after transcription or a poly(A) sequence encoded by DNA and transcribed by a template-dependent RNA polymerase. [0147] Polypeptide: As used herein, the term “polypeptide” refers to a polymeric chain of amino acids. In some embodiments, a polypeptide has an amino acid sequence that occurs in nature. In some embodiments, a polypeptide has an amino acid sequence that does not occur in nature. In some embodiments, a polypeptide has an amino acid sequence that is engineered in that it is designed and/or produced through action of the hand of man. In some embodiments, a polypeptide may comprise or consist of natural amino acids, non-natural amino acids, or both. In some embodiments, a polypeptide may comprise or consist of only natural amino acids or only non-natural amino acids. In some embodiments, a polypeptide may comprise D-amino acids, L-amino acids, or both. In some embodiments, a polypeptide may comprise only D-amino acids. In some embodiments, a polypeptide may comprise only L-amino acids. In some embodiments, a polypeptide may include one or more pendant groups or other modifications, e.g., modifying or attached to one or more amino acid side chains, at the polypeptide’s N-terminus, at the polypeptide’s C-terminus, or any combination thereof. In some embodiments, such pendant groups or modifications comprise acetylation, amidation, lipidation, methylation, pegylation, etc., including combinations thereof. In some embodiments, a polypeptide may be cyclic, and/or may comprise a cyclic portion. In some embodiments, a polypeptide is not cyclic and/or does not comprise any cyclic portion. In some embodiments, a polypeptide is linear. In some embodiments, a polypeptide may be or comprise a stapled polypeptide. In some embodiments, the term “polypeptide” may be appended to a name of a reference polypeptide, activity, or structure; in such instances it is used herein to refer to polypeptides that share the relevant activity or structure and thus can be considered to be members of the same class or family of polypeptides. For each such class, the present specification provides and/or those skilled in the art will be aware of exemplary polypeptides within the class whose amino acid sequences and/or functions are known; in some embodiments, such exemplary polypeptides are reference polypeptides for the polypeptide class or family. In some embodiments, a member of a polypeptide class or family shows significant sequence homology or identity with, shares a common sequence motif (e.g., a characteristic sequence element) with, and/or shares a common activity (in some embodiments at a comparable level or within a designated range) with a reference polypeptide of the class; in some embodiments with all polypeptides within the class). For example, in some embodiments, a member polypeptide shows an overall degree of sequence homology or identity with a reference polypeptide that is at least about 30-40%, and is often greater than about 50%, 60%, 70%, 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more and/or includes at least one region (e.g., a conserved region that may in some embodiments be or comprise a characteristic sequence element) that shows very high sequence identity, often greater than 90% or even 95%, 96%, 97%, 98%, or 99%. Such a conserved region usually encompasses at least 3-4 and often up to 20 or more amino acids; in some embodiments, a conserved region encompasses at least one stretch of at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15 or more contiguous amino acids. In some embodiments, a relevant polypeptide may comprise or consist of a fragment of a parent polypeptide. [0148] Prevent: As used herein, the term “prevent” or “prevention” when used in connection with the occurrence of a disease, disorder, and/or condition, refers to reducing the risk of developing the disease, disorder and/or condition and/or to delaying onset of one or more characteristics or symptoms of the disease, disorder, or condition. Prevention may be considered complete when onset of a disease, disorder or condition has been delayed for a predefined period of time. [0149] Recombinant: The term “recombinant” in the context of the present disclosure means “made through genetic engineering”. In some embodiments, a “recombinant” entity such as a recombinant nucleic acid in the context of the present disclosure is not naturally occurring. [0150] Reference: As used herein, the term “reference” describes a standard or control relative to which a comparison is performed. For example, in some embodiments, an agent, animal, individual, population, sample, sequence or value of interest is compared with a reference or control agent, animal, individual, population, sample, sequence or value. In some embodiments, a reference or control is tested and/or determined substantially simultaneously with the testing or determination of interest. In some embodiments, a reference or control is a historical reference or control, optionally embodied in a tangible medium. Typically, as would be understood by those skilled in the art, a reference or control is determined or characterized under comparable conditions or circumstances to those under assessment. Those skilled in the art will appreciate when sufficient similarities are present to justify reliance on and/or comparison to a particular possible reference or control. [0151] Ribonucleic acid (RNA): As used herein, the term “RNA” or “polyribonucleotide” refers to a polymer of ribonucleotides. In some embodiments, an RNA is single stranded. In some embodiments, an RNA is double stranded. In some embodiments, an RNA comprises both single and double stranded portions. In some embodiments, an RNA can comprise a backbone structure as described in the definition of “Nucleic acid / Polynucleotide” above. An RNA can be a regulatory RNA (e.g., siRNA, microRNA, etc.), or a messenger RNA (mRNA). In some embodiments where an RNA is a mRNA. In some embodiments where an RNA is a mRNA, a RNA typically comprises at its 3’ end a poly(A) region. In some embodiments where an RNA is a mRNA, an RNA typically comprises at its 5’ end an art-recognized cap structure, e.g., for recognizing and attachment of a RNA to a ribosome to initiate translation. In some embodiments, an RNA is a synthetic RNA. Synthetic RNAs include RNAs that are synthesized in vitro (e.g., by enzymatic synthesis methods and/or by chemical synthesis methods). [0152] Ribonucleotide: As used herein, the term “ribonucleotide” encompasses unmodified ribonucleotides and modified ribonucleotides. For example, unmodified ribonucleotides include the purine bases adenine (A) and guanine (G), and the pyrimidine bases cytosine (C) and uracil (U). Modified ribonucleotides may include one or more modifications including, but not limited to, for example, (a) end modifications, e.g., 5' end modifications (e.g., phosphorylation, dephosphorylation, conjugation, inverted linkages, etc.), 3' end modifications (e.g., conjugation, inverted linkages, etc.), (b) base modifications, e.g. , replacement with modified bases, stabilizing bases, destabilizing bases, or bases that base pair with an expanded repertoire of partners, or conjugated bases, (c) sugar modifications (e.g., at the 2' position or 4' position) or replacement of the sugar, and (d) internucleoside linkage modifications, including modification or replacement of the phosphodiester linkages. The term “ribonucleotide” also encompasses ribonucleotide triphosphates including modified and non-modified ribonucleotide triphosphates. [0153] Risk: As will be understood from context, “risk” of a disease, disorder, and/or condition refers to a likelihood that a particular individual will develop the disease, disorder, and/or condition. In some embodiments, risk is expressed as a percentage. In some embodiments, risk is from 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, 60, 70, 80, 90 up to 100%. In some embodiments risk is expressed as a risk relative to a risk associated with a reference sample or group of reference samples. In some embodiments, a reference sample or group of reference samples have a known risk of a disease, disorder, condition and/or event. In some embodiments a reference sample or group of reference samples are from individuals comparable to a particular individual. In some embodiments, relative risk is 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, or more. In some embodiments, risk may reflect one or more genetic attributes, e.g., which may predispose an individual toward development (or not) of a particular disease, disorder and/or condition. In some embodiments, risk may reflect one or more epigenetic events or attributes and/or one or more lifestyle or environmental events or attributes. [0154] RNA lipoplex particle: As used herein, the term “RNA lipoplex particle” refers to a complex comprising liposomes, in particular cationic liposomes, and RNA molecules. Without wishing to bound by a particular theory, electrostatic interactions between positively charged liposomes and negatively charged RNA results in complexation and spontaneous formation of RNA lipoplex particles. In some embodiments, positively charged liposomes may comprise a cationic lipid, such as in some embodiments DOTMA, and additional lipids, such as in some embodiments DOPE. In one embodiment, an RNA lipoplex particle is a nanoparticle. [0155] Selective or specific: The term “selective” or “specific”, when used herein in reference to an agent having an activity, is understood by those skilled in the art to mean that the agent discriminates between potential target entities, states, or cells. For example, in some embodiments, an agent is said to bind “specifically” to its target if it binds preferentially with that target in the presence of one or more competing alternative targets. In many embodiments, specific interaction is dependent upon the presence of a particular structural feature of the target entity (e.g., an epitope, a cleft, a binding site). It is to be understood that specificity need not be absolute. In some embodiments, specificity may be evaluated relative to that of a target-binding moiety for one or more other potential target entities (e.g., competitors). In some embodiments, specificity is evaluated relative to that of a reference specific binding moiety. In some embodiments, specificity is evaluated relative to that of a reference non- specific binding moiety. [0156] Subject: As used herein, the term “subject” refers to an organism to be administered with a composition described herein, e.g., for experimental, diagnostic, prophylactic, and/or therapeutic purposes. Typical subjects include animals (e.g., mammals such as mice, rats, rabbits, non-human primates, domestic pets, etc.) and humans. In some embodiments, a subject is a human subject. In some embodiments, a subject is suffering from a disease, disorder, or condition (e.g., an HSV infection). In some embodiments, a subject is susceptible to a disease, disorder, or condition (e.g., an HSV infection). In some embodiments, a subject displays one or more symptoms or characteristics of a disease, disorder, or condition (e.g., an HSV infection). In some embodiments, a subject displays one or more non-specific symptoms of a disease, disorder, or condition (e.g., an HSV infection). In some embodiments, a subject does not display any symptom or characteristic of a disease, disorder, or condition (e.g., an HSV infection). In some embodiments, a subject is someone with one or more features characteristic of susceptibility to or risk of a disease, disorder, or condition (e.g., an HSV infection). In some embodiments, a subject is a patient. In some embodiments, a subject is an individual to whom diagnosis and/or therapy is and/or has been administered. [0157] Suffering from: An individual who is “suffering from” a disease, disorder, and/or condition has been diagnosed with and/or displays one or more symptoms of a disease, disorder, and/or condition. [0158] Susceptible to: An individual who is “susceptible to” a disease, disorder, and/or condition is one who has a higher risk of developing the disease, disorder, and/or condition than does a member of the general public. In some embodiments, an individual who is susceptible to a disease, disorder and/or condition may not have been diagnosed with the disease, disorder, and/or condition. In some embodiments, an individual who is susceptible to a disease, disorder, and/or condition may exhibit symptoms of the disease, disorder, and/or condition. In some embodiments, an individual who is susceptible to a disease, disorder, and/or condition may not exhibit symptoms of the disease, disorder, and/or condition. In some embodiments, an individual who is susceptible to a disease, disorder, and/or condition will develop the disease, disorder, and/or condition. In some embodiments, an individual who is susceptible to a disease, disorder, and/or condition will not develop the disease, disorder, and/or condition. [0159] Synthetic: As used herein, the term “synthetic” refers to an entity that is artificial, or that is made with human intervention, or that results from synthesis rather than naturally occurring. For example, in some embodiments, a synthetic nucleic acid or polynucleotide refers to a nucleic acid molecule that is chemically synthesized, e.g., in some embodiments by solid-phase synthesis. In some embodiments, the term “synthetic” refers to an entity that is made outside of biological cells. For example, in some embodiments, a synthetic nucleic acid or polynucleotide refers to a nucleic acid molecule (e.g., an RNA) that is produced by in vitro transcription using a template. [0160] Therapy: The term “therapy” refers to an administration or delivery of an agent or intervention that has a therapeutic effect and/or elicits a desired biological and/or pharmacological effect (e.g., has been demonstrated to be statistically likely to have such effect when administered to a relevant population). In some embodiments, a therapeutic agent or therapy is any substance that can be used to alleviate, ameliorate, relieve, inhibit, prevent, delay onset of, reduce severity of, and/or reduce incidence of one or more symptoms or features of a disease, disorder, and/or condition. In some embodiments, a therapeutic agent or therapy is a medical intervention (e.g., surgery, radiation, phototherapy) that can be performed to alleviate, relieve, inhibit, present, delay onset of, reduce severity of, and/or reduce incidence of one or more symptoms or features of a disease, disorder, and/or condition. [0161] Three prime untranslated region: As used herein, the terms “three prime untranslated region” or “3' UTR” refer to a sequence of an RNA molecule that begins following a stop codon of a coding region of an open reading frame sequence. In some embodiments, the 3' UTR begins immediately after a stop codon of a coding region of an open reading frame sequence, e.g., in its natural context. In other embodiments, the 3' UTR does not begin immediately after stop codon of the coding region of an open reading frame sequence, e.g., in its natural context. [0162] Threshold level (e.g., acceptance criteria): As used herein, the term “threshold level” refers to a level that are used as a reference to attain information on and/or classify the results of a measurement, for example, the results of a measurement attained in an assay. For example, in some embodiments, a threshold level means a value measured in an assay that defines the dividing line between two subsets of a population (e.g., a batch that satisfy quality control criteria vs. a batch that does not satisfy quality control criteria). Thus, a value that is equal to or higher than the threshold level defines one subset of the population, and a value that is lower than the threshold level defines the other subset of the population. A threshold level can be determined based on one or more control samples or across a population of control samples. A threshold level can be determined prior to, concurrently with, or after the measurement of interest is taken. In some embodiments, a threshold level can be a range of values. [0163] Treat: As used herein, the term “treat,” “treatment,” or “treating” refers to any method used to partially or completely alleviate, ameliorate, relieve, inhibit, prevent, delay onset of, reduce severity of, and/or reduce incidence of one or more symptoms or features of a disease, disorder, and/or condition. Treatment may be administered to a subject who does not exhibit signs of a disease, disorder, and/or condition. In some embodiments, treatment may be administered to a subject who exhibits only early signs of the disease, disorder, and/or condition, for example for the purpose of decreasing the risk of developing pathology associated with the disease, disorder, and/or condition. In some embodiments, treatment may be administered to a subject at a later-stage of disease, disorder, and/or condition. [0164] Vaccination: As used herein, the term “vaccination” refers to the administration of a composition intended to generate an immune response, for example to a disease-associated (e.g., disease-causing) agent. In some embodiments, vaccination can be administered before, during, and/or after exposure to a disease-associated agent, and in certain embodiments, before, during, and/or shortly after exposure to the agent. In some embodiments, vaccination includes multiple administrations, appropriately spaced in time, of a vaccine composition. In some embodiments, vaccination generates an immune response to an infectious agent. [0165] Vaccine: As used herein, the term “vaccine” refers to a composition that induces an immune response upon administration to a subject. In some embodiments, an induced immune response provides protective immunity. [0166] Variant: As used herein in the context of molecules, e.g., nucleic acids, proteins, or small molecules, the term “variant” refers to a molecule that shows significant structural identity with a reference molecule but differs structurally from the reference molecule, e.g., in the presence or absence or in the level of one or more chemical moieties as compared to the reference entity. In some embodiments, a variant also differs functionally from its reference molecule. In general, whether a particular molecule is properly considered to be a “variant” of a reference molecule is based on its degree of structural identity with the reference molecule. As will be appreciated by those skilled in the art, any biological or chemical reference molecule has certain characteristic structural elements. A variant, by definition, is a distinct molecule that shares one or more such characteristic structural elements but differs in at least one aspect from the reference molecule. In some embodiments, a variant polypeptide or nucleic acid may differ from a reference polypeptide or nucleic acid as a result of one or more differences in amino acid or nucleotide sequence and/or one or more differences in chemical moieties (e.g., carbohydrates, lipids, phosphate groups) that are covalently components of the polypeptide or nucleic acid (e.g., that are attached to the polypeptide or nucleic acid backbone). In some embodiments, a variant polypeptide or nucleic acid shows an overall sequence identity with a reference polypeptide or nucleic acid that is at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, or 99%. In some embodiments, a variant polypeptide or nucleic acid does not share at least one characteristic sequence element with a reference polypeptide or nucleic acid. In some embodiments, a reference polypeptide or nucleic acid has one or more biological activities. In some embodiments, a variant polypeptide or nucleic acid shares one or more of the biological activities of the reference polypeptide or nucleic acid. In some embodiments, a variant polypeptide or nucleic acid lacks one or more of the biological activities of the reference polypeptide or nucleic acid. In some embodiments, a variant polypeptide or nucleic acid shows a reduced level of one or more biological activities as compared to the reference polypeptide or nucleic acid. In some embodiments, a polypeptide or nucleic acid of interest is considered to be a “variant” of a reference polypeptide or nucleic acid if it has an amino acid or nucleotide sequence that is identical to that of the reference but for a small number of sequence alterations at particular positions. Typically, fewer than about 20%, about 15%, about 10%, about 9%, about 8%, about 7%, about 6%, about 5%, about 4%, about 3%, or about 2% of the residues in a variant are substituted, inserted, or deleted, as compared to the reference. In some embodiments, a variant polypeptide or nucleic acid comprises about 10, about 9, about 8, about 7, about 6, about 5, about 4, about 3, about 2, or about 1 substituted residues as compared to a reference. Often, a variant polypeptide or nucleic acid comprises a very small number (e.g., fewer than about 5, about 4, about 3, about 2, or about 1) number of substituted, inserted, or deleted, functional residues (i.e., residues that participate in a particular biological activity) relative to the reference. In some embodiments, a variant polypeptide or nucleic acid comprises not more than about 5, about 4, about 3, about 2, or about 1 addition or deletion, and, in some embodiments, comprises no additions or deletions, as compared to the reference. In some embodiments, a variant polypeptide or nucleic acid comprises fewer than about 25, about 20, about 19, about 18, about 17, about 16, about 15, about 14, about 13, about 10, about 9, about 8, about 7, about 6, and commonly fewer than about 5, about 4, about 3, or about 2 additions or deletions as compared to the reference. In some embodiments, a reference polypeptide or nucleic acid is one found in nature. [0167] Vector: as used herein, refers to a nucleic acid molecule capable of transporting another nucleic acid to which it has been linked. One type of vector is a “plasmid”, which refers to a circular double stranded DNA loop into which additional DNA segments may be ligated. Another type of vector is a viral vector, wherein additional DNA segments may be ligated into the viral genome. Certain vectors are capable of autonomous replication in a host cell into which they are introduced (e.g., bacterial vectors having a bacterial origin of replication and episomal mammalian vectors). Other vectors (e.g., non-episomal mammalian vectors) can be integrated into the genome of a host cell upon introduction into the host cell, and thereby are replicated along with the host genome. Moreover, certain vectors are capable of directing the expression of genes to which they are operatively linked. Such vectors are referred to herein as “expression vectors.” In some embodiments, known techniques may be used, for example, for generation or manipulation of recombinant DNA, for oligonucleotide synthesis, and for tissue culture and transformation (e.g., electroporation, lipofection). Enzymatic reactions and purification techniques may be performed according to manufacturer's specifications or as commonly accomplished in the art or as described herein. The foregoing techniques and procedures may be generally performed according to conventional methods well known in the art and as described in various general and more specific references that are cited and discussed throughout the present specification. See e.g., Sambrook et al., Molecular Cloning: A Laboratory Manual (4th ed., Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y. (2012)), which is incorporated herein by reference for any purpose. [0168] All literature and similar material cited in this application, including, but not limited to, patents, patent applications, articles, books, treatises, and web pages, regardless of the format of such literature and similar materials, are expressly incorporated by reference in their entirety. In the event that one or more of the incorporated literature and similar materials differs from or contradicts this application, including but not limited to defined terms, term usage, described techniques, or the like, this application controls. The section headings used herein are for organizational purposes only and are not to be construed as limiting the subject matter described in any way. DETAILED DESCRIPTION OF CERTAIN EMBODIMENTS [0169] As discussed above, the present disclosure provides pharmaceutical compositions (e.g., immunogenic compositions, e.g., vaccines) for delivering herpes simplex virus (HSV) antigen constructs (e.g., HSV-1 antigen constructs, HSV-2 antigen constructs, or a combination thereof) to a subject (e.g., a patient) and related technologies (e.g., methods). In particular, the present disclosure provides HSV (e.g., HSV-1, HSV-2, or both) compositions (e.g., immunogenic compositions, e.g., vaccines) and related technologies (e.g., methods). [0170] The present disclosure provides for example, polyribonucleotides that encode one or more GP polypeptides that each comprise an HSV glycoprotein or an antigenic portion thereof. The present disclosure also provides, for example, polyribonucleotides that encode one or more T-cell string polypeptides that each comprises one or more HSV T-cell antigens or antigenic portions threof. In some embodiments, a polyribonucleotide provided herein can be part of an RNA construct. In some embodiments, a polyribonucleotide or RNA construct as described herein can be part of a composition (e.g., a pharmaceutical composition, e.g., an immunogenic composition, e.g., a vaccine). [0171] In some embodiments, technologies provided herein are directed against HSV (e.g., HSV-1, HSV-2 or both). A description of HSV and certain exemplary features is described below. I. Herpes Simplex Virus (HSV) [0172] Herpes simplex virus (HSV) belongs to the alpha subfamily of the human herpesvirus family and includes two types: HSV-1 and HSV-2. The structure of HSV-1 and HSV-2 mainly include (from inside to outside) a DNA core, capsid, tegument and envelope. Each of HSV-1 and HSV-2 have a double stranded DNA genome of about 153kb, encoding at least 80 genes. The DNA core is enclosed by an icosapentahedral capsid composed of 162 capsomeres, 150 hexons and 12 pentons, made of six different viral proteins. The DNA is surrounded by at least 20 different viral tegument proteins that have structural and regulatory roles. Some of them participating in capsid transport to the nucleus and other organelles, viral DNA entry into the nucleus, activation of early genes transcription, suppression of cellular protein biosynthesis, and RNA degradation. The viral envelope surrounding the tegument has at least 12 different glycoproteins (B-N) on their surface. The glycoproteins may exist as heterodimers (H/L and E/I) with most existing as monomers. [0173] HSV-1 and HSV-2 are responsible for a number of minor, moderate and severe pathologies, including oral and genital ulceration, virally induced blindness, viral encephalitis and disseminated infection of neonates. HSV-1 and HSV-2 are usually transmitted by different routes and affect different areas of the body, but the signs and symptoms that they cause can overlap. Infections caused by HSV-1 represent one of the more widespread infections of the orofacial region and commonly causes herpes labialis, herpetic stomatitis, and keratitis. HSV-2 typically causes genital herpes and is transmitted primarily by direct sexual contact with lesions. Most genital HSV infections are caused by HSV-2, however, an increasing number of genital HSV infections have been attributed to HSV-1. Genital HSV-1 infections are typically less severe and less prone to occurrence than genital HSV-2 infections. [0174] HSV infections are transmitted through contact with herpetic lesions, mucosal surfaces, genital secretions, or oral secretions. The average incubation period after exposure is typically 4 days, but may range between 2 and 12 days. HSV particles can infect neuronal prolongations enervating peripheral tissues and establish latency in these cells, namely in the trigeminal ganglia and dorsal root ganglia of the sacral area from where they can sporadically reactivate. Additionally, similar to other herpesviruses, HSV infections are lifelong and generally asymptomatic. Without wishing to be bound by any particular theory, it is understood that HSV particles can be shed from infected individuals independent of the occurrence of clinical manifestations. [0175] HSV infections are rarely fatal, but are characterized by blisters that can rupture and become painful. There are few clear differences in clinical presentation based on the type of infecting virus. However, as discussed above, HSV-1 infections tend to be less severe than HSV-2 infections, and patients infected with HSV-2 generally have more outbreaks. A. Lifecycle [0176] As described herein, to initiate infection, an HSV (HSV-1 or HSV-2) particle binds to the cell surface using the viral glycoproteins and fuses its envelope with the plasma membrane (see, e.g., FIG.2, Step 1). After the fusion of membranes, the viral capsid and tegument proteins are internalized in the cytoplasm (see, e.g., FIG.2, Step 2). Once in the cytoplasm, the viral capsid accumulates in the nucleus and releases viral DNA into the nucleus (see, e.g., FIG.2, Step 3). HSV replicates by three rounds of transcription that yield: α (immediate early) proteins that mainly regulate viral replication; β (early) proteins that synthesize and package DNA; and γ (late) proteins, most of which are virion proteins (see, Whitley et.al., Lancet 2001 May 12;357(9267); Taylor et.al., Front Biosci. 2002 Mar 1;7:d752-64; and Ibáñez et.al., Front Microbiol. 2018 Oct 11;9:2406; each of which is incorporated herein by reference in its entirety) (see, e.g., FIG.2, Steps 4-6). [0177] The HSV capsids are assembled within the nucleus of infected cells (see, e.g., FIG..2, Step 7). Once the assembly of viral capsids has been completed in the nucleus, these particles will continue their maturation process in this same compartment through the acquisition of tegument proteins. After leaving the nucleus, additional tegument proteins will be added to the capsids. Meanwhile, the glycoproteins are translated and glycosylated in the endoplasmic reticulum and processed in the trans-Golgi network (TGN) and then directed to multivesicular bodies (see, e.g., FIG.2, Step 8). Then, they are exported to the plasma membrane glycoproteins within early endosomes (see, e.g., FIG.2, Step 9). Viral capsids in the cytoplasm will then fuse with HSV-glycoprotein-containing endosomes to form infectious virions within vesicles (see, e.g., FIG.2, Steps 10-12). [0178] HSV (HSV-1 or HSV-2) are able to establish a latent infection. After primary infection, HSV either replicates productively in epithelial cells or enters sensory neuron axons and moves to the neuronal cell nucleus. There, the viral DNA remains as circular, extra-chromosomal DNA, and does not possess any lytic gene expression; however, latency associated transcripts are expressed and then spliced to produce RNA. This general transcriptional silence may allow the virus to remain hidden in the cell by avoiding immune surveillance. In some aspects, provided herein are technologies (e.g., compositions and methods) for augmenting, inducing, promoting, enhancing and/or improving an immune response against HSV (e.g., HSV-1 and/or HSV-2) or a component thereof (e.g., a protein or portion thereof). In some embodiments, technologies provided herein are designed to augment, induce, promote, enhance and/or improve immunological memory against HSV or a component thereof (e.g., a protein or portion thereof). In some embodiments, technologies described herein are designed to act as an immunological boost to a primary composition (e.g., immunogenic composition, e.g., vaccine), such as a composition (e.g., immunogenic composition, e.g., vaccine) directed to an epitope and/or epitopes of HSV (e.g., HSV-1 and/or HSV-2). [0179] The virus remains in this state for the lifetime of the host, or until the proper signals reactivate the virus and new progeny are generated. Progeny virus then travel through the neuron axis to the site of the primary infection to re-initiate a lytic replication cycle. B. HSV Genome [0180] The genome of HSV-1 and the genome of HSV-2 are both approximately 150 kb long of double- stranded DNA, varying slightly between subtypes and strains. The genome encodes more than 80 genes and has high GC contents: 67 and 69% for HSV-1 and HSV-2, respectively (see, Whitley et.al., Lancet 2001 May 12;357(9267); Taylor et.al., Front Biosci. 2002 Mar 1;7:d752-64; and Jiao et.al., Microbiol Resour Announc. 2019 Sep; 8(39): e00993-19, which is incorporated herein by reference in its entirety). [0181] The genome is organized as unique long region (UL) and a unique short region (US). The UL is typically bounded by terminal long (TRL) and internal long (IRL) repeats. The US is typically bounded by terminal short (IRS) and internal short (TRS) repeats. The genes found in the unique regions are present in the genome as a single copy, but genes that are encoded in the repeat regions are present in the genome in two copies (see, Whitley et.al., Lancet 2001 May 12;357(9267); Taylor et.al., Front Biosci. 2002 Mar 1;7:d752-64; and Jiao et.al., Microbiol Resour Announc. 2019 Sep; 8(39): e00993-19, which is incorporated herein by reference in its entirety). [0182] HSV contains three origins of replication within the genome that are named depending upon their location in either the Long (oriL) or Short (oriS) region of the genome. OriL is found as a single copy in the UL segment, but oriS is located in the repeat region of the Short segment; thus, it is present in the genome in two copies. Both oriL and oriS are palindromic sequences consisting of an AT-rich center region flanked by inverted repeats that contain multiple binding sites of varying affinity for the viral origin binding protein (UL9). Either oriL or one of the oriS sequences is sufficient for viral replication (see, Whitley et.al., Lancet 2001 May 12;357(9267); Taylor et.al., Front Biosci. 2002 Mar 1;7:d752-64; and Jiao et.al., Microbiol Resour Announc. 2019 Sep; 8(39): e00993-19, which is incorporated herein by reference in its entirety). [0183] The viral genome also contains signals that orchestrate proper processing of the newly synthesized genomes for packaging into pre-formed capsids. Progeny genomes are generated in long concatemers that require cleavage into unit-length monomers. For this purpose, the viral genome contains two DNA sequence elements, pac1 and pac2, that ensure proper cleavage and packaging of unit-length progeny genomes. These elements are located within the direct repeats (DR) found within the inverted repeat regions at the ends of the viral genome (see, Whitley et.al., Lancet 2001 May 12;357(9267); Taylor et.al., Front Biosci. 2002 Mar 1;7:d752-64; and Jiao et.al., Microbiol Resour Announc. 2019 Sep; 8(39): e00993-19, which is incorporated herein by reference in its entirety). C. Certain HSV Proteins 1. ICP0 [0184] Infected cell protein 0 (ICP0) of herpes simplex virus 1 (HSV-1) is an α (immediate-early) protein of herpes simplex virus 1, and is capable of activating HSV-1 gene expression, disrupt nuclear domain (ND) 10 structures, mediate the degradation of cellular proteins, and evade the host cell’s intrinsic and innate antiviral defenses (see., Smith et.al., Future Virol. 2011 Apr; 6(4): 421–429). 2. ICP22 [0185] Infected cell protein 22 (ICP22) is expressed from an immediate–early (IE) gene during the replication cycle of HSV-1 and HSV-2. ICP22 can generally regulate viral and host gene transcription by changing the phosphorylation status of host RNA polymerase II (RNA pol II) and can also facilitate the nuclear egress complex (NEC) accurately locate to the nuclear membrane to promote nuclear budding (see, Wu et.al., Front Microbiol. 2021 Jun 7;12:668461). 3. VP16 [0186] The UL48 gene encodes VP16 or alpha-gene-transactivating factor (α-TIF). VP16 is an important transactivator that can activate the transcription of viral immediate-early genes, and in the late stage of viral replication. Additionally, VP16, as a tegument, is involved in viral assembly (see Fan, et.al., Front Microbiol. 2020; 11: 1910). [0187] In the early stage of viral infection, VP16 released by invading virions binds to the immediate-early (IE) gene promoter to stimulate the transcription of IE genes as a transactivating factor that acts specifically on IE genes (see Fan, et.al., Front Microbiol. 2020; 11: 1910). In the late stage, VP16 assembles into the tegument to participate in the assembly of virions and promote their maturation (see Fan, et.al., Front Microbiol. 2020; 11: 1910). 4. Glycoproteins [0188] In order to replicate, enveloped HSV must be able to fuse with the membrane of a living cell and deliver their genetic material into its cytoplasm. The HSV viral envelope surrounding the tegument has at least 12 different glycoproteins (gB-gN) on their surface. The glycoproteins may exist as heterodimers (gH/gL and gE/gI) with most existing as monomers. HSV gC, gB, gD, gH, and gL are involved in the process of viral cell entry. Initial attachment is mediated by gC, followed by gD. Then gH/gL pull the virus and the cell membrane together, and then gB triggers the membrane fusion. (Reske et. al., Rev Med Virol. May-Jun 2007; and Arii et. al., Adv Exp Med Biol. 2018;1045:3-21). [0189] The present disclosure provides HSV glycoprotein (e.g., gB, gC, gD, gE, gG, gH, gI, and/or gL) antigens and antigenic fragments thereof can be useful in preventing or treating HSV, e.g., in HSV antigen constructs and/or HSV compositions (e.g., immunogenic compositions, e.g., vaccines) as further disclosed herein. 5. Glycoprotein C (gC) [0190] Mature HSV glycoprotein C (gC) is a 56 kDa protein that plays a role in initial cell attachment. Glycoprotein C is a type I membrane glycoprotein and is considered a significant attachment protein and principle viral ligand for binding heparin sulfate proteoglycans (HSPGs) on a cell surface. This binding can occur by gC interaction with HSPG rich regions found on F-actin rich membrane protrusions referred to as filopodia. [0191] Glycoprotein C has also been shown to be involved in regulation of cell entry and infection by increasing pH threshold for acid-induced conformational changes of gB. Low pH induces reversible conformational changes to gB domains I and V, the functional region containing hydrophobic loops important in cell fusion. By positively regulating low-pH-induced conformational changes of gB, gC can enhance HSV’s ability to invade cell types, like epithelial cells, that require a low-pH mechanism for invasion. [0192] Glycoprotein C has also been shown to play a role in immune evasion, in addition to its role in attachment. Glycoprotein C is a target for lymphocyte cytotoxicity in certain cell types and is able to bind complement component C3b to inhibit compliment activation. Furthermore, neutralizing epitopes that exist on other HSV glycoproteins, like gB, can be protected by gC, preventing immune responses from blocking fusion. 6. Glycoprotein D (gD) [0193] HSV glycoprotein D (gD) is a 46 kDA type I membrane glycoprotein. The N-terminal ectodomain is comprised of 316 amino acids. Glycoprotein D facilitates invasion by interacting with several cell surface receptors, including herpesvirus entry mediator (HVEM), nectin-1 or nectin-2, and heparin sulfate that contain specific modifications. These cellular receptors do not function as co-receptors, as each glycoprotein interaction with a cell’s receptor occurs independently of each other. Binding of gD to one of these cellular receptors causes a conformational change that converts gD’s auto-inhibitory closed state into an active state that transmits one of two signals believed to be required for gH/gL complex activation. HVEM, the first gD receptor identified, belongs to the tumor necrosis factor receptor family and is commonly found on T cells, B cells, dendritic cells, natural killer cells, macrophages, as well as non-immune cell types like neurons and epithelial cells. Within the N-terminus of gD, there is a 37 residue hairpin structure that forms the entire site for binding to HVEM. Specifically, residues 1-32 of gD’s N-terminal domain bind HVEM’s cysteine-rich domain 1. When not in contact with HVEM, this N-terminal extension adopts an extended and flexible conformation. [0194] Clinical strains of HSV use nectin-1 for cell entry; however, several mutant strains of HSV utilize nectin-2. Furthermore, heparin sulfate is utilized by HSV-1 but not HSV-2. Glycoprotein D interaction with net-1 has been shown to be essential in some cell types such as neurons, even when other receptors are present on a cell surface. 7. Glycoprotein H (gH)/Glycoprotein L (gL) Complex [0195] Glycoprotein H (gH) is an essential 56kD protein that exists as a heterodimeric complex with 25 kDa glycoprotein L (gL) (complex referred to herein as gH/gL). The gH/gL complex is required for cell fusion and entry. gH/gL does not share any structural similarities with documented fusion proteins and likely does not function as a cofusogen with gB. Instead, gH/gL may act as a regulator of fusion and important component in stabilizing contact between HSV and a cell. Glycoprotein H receives a signal from gD through its H1 domain, and transmits this signal to membrane proximal H3 domain, which in turn propagates that signal to gH’s cytoplasmic tail. Once gH’s cytoplasmic tail receives this signal, it releases strain on the pre-fusion conformation of gB, which favors attachment of gB’s fusion loop to a cell surface, promoting gB mediated membrane fusion. Mutations in gH’s C-terminal tail have been shown to reduce fusion activity. Furthermore, antibody responses directed towards gH have been shown capable of inhibiting fusion processes mediated by gB-gH-gL. In addition to this essential role, gH contains an arginylglycylaspartic acid (RGD) motif that can bind integrin receptors found on cells. Interaction of gH with integrin is believed to trigger intracellular signals which facilitate capsid transport. 8. Glycoprotein B (gB) [0196] Glycoprotein B is a protein that has an apparent molecular weight of approximately 95-100 kDa and consists of an extended rod or spike-like ectodomain, a hydrophobic membrane proximal region (MPR), a transmembrane region (TMR), and a C-terminal domain (CTD). The ectodomain is well characterized to actively participate in fusion, while MPR, TMR, and CTD can play roles in regulation fusion. Glycoprotein B is a class III fusogen. Glycoprotein B ectodomain architecture shares conformational similarity with fusogens from viruses not belonging to the herpesvirdae family. Glycoprotein B is activated through its interaction with gH/gL, but HSV cannot fuse with a target cell through activation of gB alone and requires gB interaction to specific receptors for fusion to be completed. A well-known receptor target of gB is cell-surface heparin sulfate, an interaction that is not essential for HSV fusion, but is known to promote viral adhesion to a cell surface. Glycoprotein B can also interact with paired immunoglobulin-like type 2 receptor, most commonly found on monocytes, macrophages, and dendritic cells. [0197] HSV gB exists in two forms, a pre-fusion and post fusion form. Several changes in the pre-fusion form of gB are thought to lead to its active and post-fusion state. The first change occurs at domain V or at MPR, which allows fusion loops to point towards a cell membrane and away from a viral membrane. This change can produce a compacting intermediate conformation 1 that does not yet attach to a cell membrane surface. The next change occurs at domain III and involves gB adopting an extended intermediate conformation 2 that allows its fusion loop to attach to a cell membrane surface. Lastly, changes in domain V convert gB to its post-fusion conformation that favors membrane fusion. [0198] The post-fusion form of HSV-1 gB has an ectodomain that exists as three protomers that interact to produce a rod-like trimeric structure. Each promoter is comprised of five distinct domains with linker regions that individually form a hairpin shape. Each domain of an individual protomer interacts with the same domain of an adjacent protomer to form the described trimeric structure. Domain I houses an important fusion loop and is commonly referred to as the fusion domain. Domain II facilitates interactions with gH/gL and is referred to as the gH/gL domain. Domain III is comprised of alpha helices that help form the trimeric coil-coil central core of this protein. Domain IV is referred to as the crown domain and sits on top of the post-fusion form; it is believed to bind with cellular receptors. Antibodies that bind to the crown domain can disrupt gB binding to cellular receptors. Domain V consists of a long extension and connects protomers together. 9. Glycoprotein E and glycoprotein I (gE/gI) [0199] Glycoprotein E is approximately 53 kDa and Glycoprotein I is approximately 141 kDa. Both proteins interact to form a heterodimeric complex (complex referred to herein as gE/gI) that plays a role in cell-to-cell spread and virus induced fusion. The gE/gI complex, unlike gB, gD, and gH/gL, is not required for fusion and entrance into a cell, but is important for cell-to-cell spread. Disruption of gE/gI formation has effects on HSV proliferation, as this virus relies on cell-to-cell spread for its lytic cycle. The mechanism in which gE/gI facilitate cell-to-cell spread is thought to be reliant on several tegument polypeptides. Cooperation of tegument polypeptides, UL11, UL16, and UL21 may play a role in processing, transport, and biological activity of gE. 10. Glycoprotein G [0200] Glycoprotein G (gG) from both HSV-1 (gG1) and HSV-1 (gG2) is the first viral chemokine-binding protein shown to potentiate chemokine function of a cell. Glycoprotein G varies in size significantly between HSV-1 and HSV-2, with a 76 kDa and 43 kDa size, respectively. Glycoprotein G is unique in that its soluble form (SgG2) can have immune modulatory capacity through its extracellular activity. Once extracellular, SgG2 binds chemokines through the glycosaminoglycan (GAG)-binding domain of a chemokine without interfering with chemokine’s G protein coupled receptors (GPCRs) binding site. SgG2’s interaction with GAG containing proteins allows initiation of lipid raft formation and accumulation, which produces a clustering of chemokine receptors into this micro domain. Clustering of chemokine receptors, in turn, increases local concentration of chemokines on a host cell’s extracellular surface and allows these chemokines to interact with GPCRs. This interaction likely leads to increased immune signaling responses and chemokine stimulation. This combination of receptor relocalization and presentation of the chemokine complex with SgG2 provides a molecular rationale for enhancement of chemokine function during HSV infection. This immune modulation is in contrast to what is seen in other viruses that inhibit chemokine function, as in this case, chemokine function is potentiated by SgGs. Without being bound to any particular theory, it is thought that an overall manipulation of endogenous immune signaling may be overall favorable to HSV. 11. ICP47 [0201] Infected cell protein 47 (ICP47) encoded by gene US12, is a polymorphous protein and could block RNA splicing in early infection, and then, shuttle viral RNA from nucleus to cytoplasm in late infection. ICP47 directly binds antigen-dependent transporter (TAP), limiting antigen trafficking, leading to the occurrence of empty MHC-I (Cheng et.al., Virol J. 2020 Jul 10;17(1):101). The binding of ICP47 to TAP stabilizes the inward conformation, therefore blocking the translocation pathway points to the endoplasmic reticulum (ER) cavity. By blocking the entry of viral antigens into ER, HSV could avoid the attack of cytotoxic T lymphocytes, which may lead to immune escape of HSV and establish lifelong infection in the host cells (Cheng et.al., Virol J. 2020 Jul 10;17(1):101). 12. VHS [0202] The virion-host shutoff (VHS) protein is viral protein synthesized with late kinetics and packaged into mature virion particles. Functionally, VHS is a viral RNase that preferentially degrade both host and viral RNA species. VHS has been reported to interfere with dendritic cells (DC) activation during both productive and nonproductive HSV infection (Cotter et.al., J Virol. 2011 Dec; 85(23): 12662–12672.). 13. US3 [0203] All members of the Alphaherpesvirinae subfamily encode a serine/threonine kinase, designated US3. US3 is a significant virulence factor for herpes simplex virus type 1 (HSV-1), and is a multifunctional polypeptide that plays various roles in the viral life cycle by phosphorylating a number of viral and cellular substrates (Kato et.al., Adv Exp Med Biol. 2018;1045:45-62.). D. HSV Compositions [0204] Several HSV compositions, mainly targeting HSV-2 and primarily focused on the generation of neutralizing antibodies (nAbs) targeting the viral envelope glycoprotein D as the correlate of immune protection, have been developed and evaluated in human clinical trial, see Table 1 below. Despite these compositions exhibiting protection against HSV in preclinical studies and in some cases Phase 2 studies, none of these compositions has demonstrated sufficient efficacy for further development or commercialization. [0205] The present disclosure provides an insight that many prior strategies for developing pharmaceutical compositions (e.g., immunogenic compositions, e.g., vaccines) for treatment of and/or protection from HSV infection have focused primarily, or even almost exclusively, on development of neutralizing antibodies that target surface glycoproteins. The present disclosure identifies a problem with such strategies including, for example, that they may fail to appreciate value or even criticality of ensuring that an induced immune response includes significant T cell activity (in some embodiments, CD4 T cell activity, in some embodiments CD8 T cell activity, in some embodiments, both). In some embodiments, pharmaceutical compositions (e.g., immunogenic compositions, e.g., vaccines) that comprise or deliver CD4 and CD8 epitope(s) of one or more HSV antigens (e.g., HSV-1 antigens, HSV-2 antigens, or a combination thereof), e.g., in addition to one or more B cell antigens and/or epitopes may be used in treatment of and/or protection from HSV infection. Table 1: Certain HSV Compositions Under Clinical Development Name Platform Antigens Immune Clinical results Response is , is c c c y
Figure imgf000041_0001
Failed to reduce shedding c
Figure imgf000042_0001
[0206] The present disclosure provides the recognition that constructs and/or compositions described herein may be administered as part of regimen with other therapeutic agents. The present disclosure also recognizes that subjects that are administered constructs and/or compositions described herein may have previously been administered other therapeutic agents. [0207] In some embodiments, for example, a subject may be receiving or had previously received an anti- viral agent for HSV. In some embodiments, an anti-viral agent can be administered to treat HSV-1 or HSV-2 infection or recurrent episodes. In some embodiments, an anti-viral agent is or comprises acyclovir, valacyclovir, famciclovir, or a combination thereof. Table 2 below provides certain information about select anti-viral agents. Table 2: Antiviral Drugs for Treating HSV Medication Dosage Most common adverse effects Approximate price for com lete dosin
Figure imgf000042_0002
Apply to affected area on 1-Docosanol face or lips at the first sign (Treatment for Redness or swelling of cold sore/fe er blister
Figure imgf000043_0001
[0208] The present invention provides combinations of nucleotides (e.g., polyribonucleotides) that can be used to express and/or deliver one or more HSV polypeptides or an antigenic portion thereof. [0209] In some embodiments, the present disclosure provides a combination comprising a plurality of polyribonucleotides. In some embodiments the plurality of polyribonucleotides comprise a first set of polyribonucleotides. In some embodiments the plurality of polyribonucleotides comprise a first set of polyribonucleotides and a second set of polyribonucleotides. In some embodiments, a first set of polyribonucleotides encode one or more glycoprotein (GP) polypeptides. In some embodiments, a second set of polyribonucleotides encode one or more T-cell string polypeptides. [0210] The present disclosure provides the insight that delivering one or more GP polypeptides and one or more T-cell string polypeptides to a subject can induce a more robust immune response to HSV (e.g., HSV-1, HSV-2 or both). For example, GP polypeptides can induce humoral immune responses (e.g., B cell response) to portions of an HSV particle that are surface exposed in a subject prior to infection, while T-cell string polypeptides can induce cell-mediated immune responses (e.g., T cell response) to HSV molecules (e.g., proteins) produced inside a subject’s cells following infection. By inducing multiple immune system responses to HSV in a subject, additional protection against HSV can be achieved. A humor immune response, for instance, can be helpful for preventing an infection from occurring and/or reducing cell-to-cell transfer of HSV following infection, while a cell-mediated immune response can be helpful for preventing an early infection from taking off and/or clearing an active infection. A. HSV glycoproteins and antigenic fragments thereof [0211] In some embodiments, a first set of polyribonucleotides can include one or more polyribonucleotides. In some embodiments, a polyribonucleotide in a first set can encode a glycoprotein (GP) polypeptide. In some embodiments, a GP polypeptide comprises an HSV glycoprotein or an antigenic portion thereof. [0212] In some embodiments, a first set of polyribonucleotides encodes two or more GP polypeptides. For instance, in some embodiments, a first set of polyribonucleotides comprises two or more polyribonucleotides. In some embodiments, two or more GP polypeptides comprise an HSV glycoprotein or an antigenic portion thereof. In some embodiments, the two or more GP polypeptides differ. [0213] In some embodiments, a first set of polyribonucleotides encode three or more GP polypeptides. In some embodiments, a first set of polyribonucleotides comprises three or more polyribonucleotides. In some embodiments, three or more GP polypeptides comprise an HSV glycoprotein or an antigenic portion thereof. In some embodiments, the three or more GP polypeptides differ. [0214] In some embodiments, a first set of polyribonucleotides encode four or more GP polypeptides. In some embodiments, a first set of polyribonucleotides comprises four or more polyribonucleotides. In some embodiments, four or more GP polypeptides comprise an HSV glycoprotein or an antigenic portion thereof. In some embodiments, the four or more GP polypeptides differ. [0215] In some embodiments, a first set of polyribonucleotides encode one or more GP polypeptides, wherein the one or more GP polypeptides comprise an HSV glycoprotein C (gC) or an antigenic portion thereof, an HSV glycoprotein D (gD) or an antigenic portion thereof, an HSV glycoprotein E (gE) or an antigenic portion thereof, an HSV glycoprotein B (gB) or an antigenic portion thereof, an HSV glycoprotein I (gI) or an antigenic portion thereof, an HSV glycoprotein G (gG) or an antigenic portion thereof, an HSV glycoprotein H (gH) or an antigenic portion thereof, an HSV glycoprotein L (gL) or an antigenic portion thereof, or a combination thereof. [0216] In some embodiments, a first set of polyribonucleotides comprises (i) a polyribonucleotide encoding an HSV gC or an antigenic portion thereof, (ii) a polyribonucleotide encoding an HSV gD or an antigenic portion thereof, (iii) a polyribonucleotide encoding an HSV gE or an antigenic portion thereof, or (iv) a combination thereof. [0217] In some embodiments, a first set of polyribonucleotides comprises (i) a polyribonucleotide encoding an HSV gB or an antigenic portion thereof, (ii) a polyribonucleotide encoding an HSV gC or an antigenic portion thereof, (iii) a polyribonucleotide encoding an HSV gD or an antigenic portion thereof, or (iv) a combination thereof. [0218] In some embodiments, a first set of polyribonucleotides comprises (i) a polyribonucleotide encoding an HSV gC or an antigenic portion thereof, (ii) a polyribonucleotide encoding an HSV gD or an antigenic portion thereof, (iii) a polyribonucleotide encoding an HSV gE or an antigenic portion thereof, (iv) a polyribonucleotide encoding an HSV gB or an antigenic portion thereof, or (v) a combination thereof. [0219] In some embodiments, a first set of polyribonucleotides comprises: (i) a polyribonucleotide that encoding an antigenic portion of HSV gC, (ii) a polyribonucleotide that encoding an antigenic portion of HSV gD, (iii) a polyribonucleotide that encoding an antigenic portion of HSV gE, or (iv) a combination thereof. [0220] In some embodiments, a first set of polyribonucleotides comprises: (i) a polyribonucleotide that encoding an antigenic portion of HSV gC, (ii) a polyribonucleotide that encoding an antigenic portion of HSV gD, (iii) a polyribonucleotide that encoding an antigenic portion of HSV gE, (iv) a polyribonucleotide that encoding an antigenic portion of HSV gB, or (v) a combination thereof. [0221] In some embodiments, the first set of polyribonucleotides comprises (i) a polyribonucleotide that encoding an antigenic portion of HSV gC, (ii) a polyribonucleotide that encoding an antigenic portion of HSV gD, and (iii) a polyribonucleotide that encoding an antigenic portion of HSV gE. [0222] In some embodiments, the first set of polyribonucleotides comprises (i) a polyribonucleotide that encoding an antigenic portion of HSV gC, (ii) a polyribonucleotide that encoding an antigenic portion of HSV gD, (iii) a polyribonucleotide that encoding an antigenic portion of HSV gE, and (iv) a polyribonucleotide that encoding an antigenic portion of HSV gB. [0223] The present invention provides one or more glycoprotein (GP) polypeptides each comprising an HSV glycoprotein or an antigenic portion thereof. [0224] In some embodiments, an HSV glycoprotein or an antigenic portion thereof is an HSV-1 glycoprotein or an antigenic portion thereof. In some embodiments, an HSV glycoprotein or an antigenic portion thereof is an HSV-2 glycoprotein or an antigenic portion thereof. [0225] An overview of exemplary HSV glycoproteins with or without a secretory signal is provided in Table 3 below. Table 3: Exemplary HSV Glycoprotein, Secretory Signal, and Versions HSV-2 Secretory Amino nt Version 1 Version 2 Version 3 Version 4 Glycoprotein signal acid SEQ ID nt nt nt nt SEQ ID NO SEQ ID SEQ ID SEQ ID SEQ ID
Figure imgf000045_0001
HSV-2 gE – 327 325 LVVV (SEQ ID NO:
Figure imgf000046_0001
Table 4 below, example deoxyribonucleic acid sequences encoding certain HSV gB, gC, gD, and gE glycoproteins are provided in Table 5, and example ribonucleic acid sequences encoding certain HSV gB, gC, gD, and gE glycoproteins are provided in Table 6 below. Table 4: Example HSV Glycoproteins or Antigenic Portions SEQ Glycoprotein or Amino Acid Sequence ID antigenic NO fr m nt SE G RP T T A P RP T T A C T RI A A R R P HA P F P N K S F R A F N SN
Figure imgf000046_0002
PFGALAVGLLVLAGLAAAFFAFRYVMRLQSNPMKALYPLTTKELKNPTNPDASGEGEEGGDFDEA KLAEAREMIRYMALVSAMERTEHKAKKKGTSALLSAKVTDMVMRKRRNTNYTQVPNKDGDADE DDL K T K H A AI S M Q Y LR LA A K T K H A AI LS M Q Y LR LA A KK E A G LT TF LA I M EY DI LL Q P F P N K S F R A F N SN A
Figure imgf000047_0001
KLAEAREMIRYMALVSAMERTEHKAKKKGTSALLSAKVTDMVMRKRRNTNYTQVPNKDGDADE DDL 384 HSV-2 gB / UL27 MRGGGLICALVVGALVAAVASAAPAAPAAPRASGGVAATVAANGGPASRPPPVPSPATTKARKRK T K H A AI S M Q Y LR LA A
Figure imgf000048_0001
SEQ Antigen / Version Polynucleotide Sequence ID NO ORF 4 H 2 T A AT A T AA T AA CC C C C C C C CC T G C A AC G T C C C T T G CA T A T C T C G G G C A CA G AA
Figure imgf000048_0002
GGGTGTTCGACCCCGCTCAGATCCACACACAGACCCAAGAGAACCCCGACGGCT TTAGCACCGTGTCCACAGTGACATCTGCCGCCGTTGGAGGACAGGGCCCTCCTA GAACCTTTACCTGCCAGCTGACCTGGCACAGAGACAGCGTGTCCTTCAGCAGAA T C C C G CT AC C G CA TG CC G TG C G A A G G A CA G G C A GT C C C G C C T TC C G C A C G A A A TC TC C T A C C C G
Figure imgf000049_0001
GCATGACCCACTTGCACGCTATGGGTCAAGAGTCCAGATACGGTGTCGCTTCCC TAACAGTACAAGGACTGAGTTTCGGCTGCAGATCTGGCGTTATGCCACAGCTAC TGACGCAGAGATTGGTACCGCCCCCAGTTTGGAAGAGGTGATGGTCAACGTGT TC C G C A C G A A A TC TC C T A C CT T AT C AC C A G G A C A T A CT G G TT G CC C C CT T AT C AC C A G G A C A T A CT G
Figure imgf000050_0001
CCTCTGGAACCGCCAGTGTGTTGCCGAGGCCGACTATCACGATGGAATTCACAG GCGATCATGCCGTCTGTACTGCCGGCTGTGTGCCAGAAGGCGTAACCTTCGCTT GGTTTCTCGGGGATGACTCAAGTCCTGCAGAGAAAGTGGCTGTGGCCTCTCAG CC C C CT T AT C AC C A G G A C A T A CT G G TT G CC C C CT T AT C AC C A G G A C A T A CT G G TT G CC C CC T T A C C C
Figure imgf000051_0001
CGCCCCTCCTGGCGGCCAGCTGGTGTACGATTCCGCCCCCAACAGAACCGATCC CCACGTGATCTGGGCCGAAGGCGCCGGCCCCGGCGCCTCCCCCAGACTGTACT CCGTGGTGGGCCCCCTGGGCAGACAGAGACTGATCATCGAAGAACTGACCCTG C A C G T A A C T T G A CA C C G T G TC G G C G C T T A T C G T A T G CC CC C G CA G T C T A T CT C T C T G G C G
Figure imgf000052_0001
ACTTCCTGTGGACGTCCTGGAACTGCGACAATCCGAAGCACACTGCCGGTTTCC TACGAGCAGACGGAGTACATATGCCGCCTTGCAGGCTACCCCGATGGAATTCCA GTCCTTGAGCACCATTGA C C C A AA A G C A A A C TT A C C A C G C TT A C C C CA AA A G C C A T C T A C CC T A C A C C C C C A A GA G C T A
Figure imgf000053_0001
GACACAGGGTATGTACTACTGGGTCTGGGGCAGAACCGACAGGCCATCTGCTT ACGGGACATGGGTCCGCGTTCGAGTATTTCGGCCACCCTCACTGACCATACATC CCCATGCCGTTCTTGAAGGGCAGCCTTTCAAGGCAACCTGTACTGCTGCCACAT G T C A C G C A TC CT T A T C C C A G A A CC TA T A C AC C C A T TC C G C C C C C C CC TC C A T C T T C G C C A
Figure imgf000054_0001
AGCACACCTACAACCTGACAATCGCCTGGTACAGAATGGGCGACAACTGCGCCA TTCCTATCACCGTGATGGAGTACACCGAGTGTCCCTACAACAAGAGCCTGGGCG TGTGCCCCATCAGAACACAGCCTAGATGGTCCTACTACGACAGCTTCAGCGCCG G A C T G A G CT T G G C CA A T T T G A C CA T T C C T G G C CA A T T T G A TC CA T T C C T G G C CA A T T T G A
Figure imgf000055_0001
TCCTGGAACATAGGGCCAGAGCCAGCTGCAAGTATGCCCTTCCCCTGCGGATTC CGCCTGCAGCATGTCTGACCTCAAAAGCCTACCAGCAAGGGGTGACTGTGGACA GCATTGGCATGCTGCCTCGTTTCATTCCCGAGAATCAACGGACAGTGGCTCTGT T C C T A GA C AC AA C G G C C T CC C G C CC C C G C A A A C C G C G G C T C T C G G C A C G C A T T C T TC T C
Figure imgf000056_0001
GTGACCGTGCACATGGAAACACCTGAGGCCATCCTGTTCGCCCCTGGCGAGACA TTTGGCACCAACGTGTCCATCCACGCTATCGCCCACGACGATGGCCCTTACGCC ATGGATGTCGTGTGGATGAGATTCGACGTGCCCAGCAGCTGTGCCGAGATGAG C T A TC C G CT C G AC A T T C T TC T C CA C G C T A TC C G CT A A C C T C TC C A AG CG AT A G A A T CA A A A A A C C
Figure imgf000057_0001
CTCTGGCCATCGCCTATAGTCCCCCTTTTCCCGCTGGCGACGAGGGGCTTTACT CCGAACTGGCCTGGCGGGATAGGGTGGCGGTGGTGAACGAGAGCCTCGTCATC TACGGTGCTCTGGAAACCGACTCAGGACTGTATACGCTCAGCGTTGTTGGCCTC C AG T GT TT T A G T G AG T T C A A C C T C TC C A AG CG AT A G A A T CA A A A A A C C T C TC C A AG CG AT A G A A T CA A
Figure imgf000058_0001
TCACATCCATGCGTGGGGGCATATGACCATCAGCACAGCTGCCCAGTACCGCAA TGCCGTCGTGGAGCAGCACCTCCCCCAACGGCAGCCAGAACCAGTGGAGCCCA CTCGGCCTCATGTGCGAGCCTGATAA C G CC A C T C C G T C C C G C TA T C T A C C G AC A T T CT C T G G T A AA G G G C C G CA C G A A A A CC A T C G AC
Figure imgf000059_0001
TTTTGGGACCAATGTGAGCATTCACGCCATAGCTCACGATGACGGGCCCTATGC CATGGACGTGGTGTGGATGAGGTTCGATGTGCCCTCATCATGCGCTGAGATGC GGATCTACGAAGCTTGCCTGTATCACCCACAGCTTCCCGAGTGCTTGTCTCCCG CT AA C C G C CA G T C T T AC T CT GC T C G C G A A G G G C C G G CC G AC G T C T GC A AC T C CT C A A C G C GC C G C C
Figure imgf000060_0001
CCTGGTCAGCTTTCGGTACGAAGACCAGGGCCCGTTGGTCGAGGGGCAGCTGG GGGAGAACAACGAGCTGCGGCTGACGCGCGATGCGATCGAGCCGTGCACCGTG GGACACCGGCGCTACTTCACCTTCGGTGGGGGCTACGTGTACTTCGAGGAGTA T A C G G G G T C G G A C G T C C G C G C T CT C A G A G C G CA G A C G A T C T C C T GC G G G A A A A C C C T T
Figure imgf000061_0001
GCCGTGTCTGGGGTTTCCAGCTTCATGAGCAATCCCTTTGGAGCCCTGGCCGT GGGACTGCTGGTTTTGGCTGGACTTGCCGCTGCCTTCTTCGCCTTTAGATACGT GATGCGGCTCCAGAGCAACCCCATGAAGGCCCTGTATCCACTGACCACCAAAGA G A G A G AT A C C A T T C CA A T C C T C G A G G C G T A T A A C G G T A TC A A C CA T AA T G A C G T T GT C T
Figure imgf000062_0001
744 HSV-1 gB GCTCCAACCTCTCCTGGAACCCCTGGAGTGGCTGCTGCAACCCAGGCTGCAAAT GGAGGACCTGCAACCCCTGCTCCTCCTCCTCTGGGAGCTGCTCCAACAGGAGAT CCAAAACCTAAAAAGAATAAAAAGCCTAAAAATCCAACCCCTCCAAGACCTGCTG A A A C G T C A G G TA T G C G G G GA GC C A C C T A C G A T G T G G A AT G T A G G AA A G A G GT A G G T C A C G G
Figure imgf000063_0001
GGGGTCTGCCGCTCCACGGCCAAGTACGTGCGGAACAACATGGAGACCACCGC GTTTCACCGGGACGACCACGAGACCGACATGGAGCTCAAGCCGGCGAAGGTCG CCACGCGCACGAGCCGGGGGTGGCACACCACCGACCTCAAGTACAACCCCTCGC G A CA G GA C G C G T C G G G G G G T G G C T CA T G A T T A G G C C T T C T C AC C CC AC A T A G C G A GC TA A A T C
Figure imgf000064_0001
AAGTGGCAAGAGGTGGACGAGATGCTGAGAGCCGAGTACGGCGGCAGCTTCAG ATTCAGCTCTGACGCCATCAGCACCACCTTCACCACCAATCTGACCGAGTACAG CCTGTCCAGAGTGGACCTGGGCGATTGCATCGGCAGAGATGCCAGAGAAGCCA C A A C TC G A G A C G T C C T G C TT GT TT T C A GC C CA T T C C G C A G C T T A C G A T C AC A A T T C A T C G T G
Figure imgf000065_0001
TGGCCTGGTGCGAGTTGCAGAACCACGAACTGACTCTGTGGAATGAGGCCCGT AAGCTGAACCCAAATGCCATAGCCAGTGCCACGGTAGGCAGGAGAGTTTCAGCA CGGATGCTTGGTGATGTGATGGCCGTGAGTACATGCGTGCCAGTAGCACCAGA T G T TT C G A GT A C TT T C G C CA T A T C C C C C AA CT A C A C T A G T A A G T C A T A AA G A T C G T C G G C A C
Figure imgf000066_0001
CAGACATGAAATCAAAGATTCTGGACTGCTGGATTACACAGAAGTGCAGAGAAG AAATCAGCTGCATGATCTGAGATTTGCTGATATTGATACAGTGATCAGAGCTGA TGCAAATGCTGCAATGTTTGCTGGACTGTGTGCTTTCTTTGAAGGAATGGGAGA G T A A G C AA A A C CT T AT C AC C A G G A C A T A CT G G TT G CC C G G C G G C CA A T T T G A C CA T T C C T A G
Figure imgf000067_0001
AGCCCAGATGGCTCCGAAGCGTCTGCGATTGCCCCACATACGGGACGACGACG CCCCTCCATCCCATCAACCCCTGTTTTACTGATAA 759 HSV-2 gE CGCACCTCTTGGAAACGCGTTACTTCCGGGGAGGACGTTGTCCTCCTTCCAGCA A C C T C TC C A AG CG AT A G A A T CA A A A GA C C A G CC C C
Figure imgf000068_0001
SEQ Antigen / Versions Polynucleotide Sequence ID NO ORF C C C C C C C C C U C C C GA C A U C C A A
Figure imgf000068_0002
UCCGCUCCACCCUGCCCGUGUCCUACGAGCAGACCGAGUACAUCUGCCGCCUG GCCGGCUACCCCGACGGCAUCCCCGUGCUGGAGCACCACUAA 17 HSV-2 gC Version 1 AGCGCUUCUCCCGGCAGAACCAUCACAGUGGGCCCUAGAGGCAACGCCUCUAA A U G A C G A C G C U C U A G AC G U C C G C C U G CC C G C A C G C G G A A U A U C AC C C C U A C A U A C G G
Figure imgf000069_0001
ACCGCCCUUCCGCAUAUGGCACUUGGGUGAGAGUUCGCGUCUUUCGGCCCCC UUCUCUCACCAUCCAUCCUCAUGCCGUGCUCGAAGGCCAGCCCUUUAAGGCCA CAUGCACUGCUGCGACCUACUACCCUGGCAACAGAGCCGAGUUUGUCUGGUU G G C A G U A C C C U A C A U A C A G C A U G G C A G U A C GC C U G C G A GA G G C C U A A G G C U G C CA
Figure imgf000070_0001
290 HSV-2 gC Version 2.3 AGCCCCGGCAGAACCAUAACAGUAGGGCCACGGGGGAAUGCUUCCAAUGCUGC [UL44] ACCUUCCGCUUCACCGAGGAAUGCUUCUGCCCCAAGAACUACCCCCACUCCUC CUCAACCCAGGAAAGCGACAAAGUCCAAGGCCAGCACCGCAAAACCCGCUCCU G C G A GA G A C C U A A G G C U G C CA GC C U G C G A GA G A C C U A A G G C U G C CA GC C U G C G A GA G A C C U
Figure imgf000071_0001
GCACUGCUGCGACCUACUACCCUGGCAACAGAGCCGAGUUUGUCUGGUUUGA GGAUGGUCGGCGAGUAUUCGAUCCAGCCCAGAUUCACACACAAACGCAGGAAA AUCCGGACGGCUUCAGCACAGUGUCCACGGUGACCUCUGCUGCAGUUGGUGG G C U G C CA C C C A UC C G C U G C U A U C G C A C A U G C C C A C A G C A C G U G A AA A U A G C A G C CC C
Figure imgf000072_0001
CACCCAAAACCGGACCUCCUAAGACCAGCUCUGAACCGGUGCGGUGUAAUAGG CACGACCCAUUGGCGCGAUAUGGCAGUAGGGUCCAGAUACGGUGCAGAUUCC CAAACAGCACAAGAACAGAAUUCCGGCUGCAAAUCUGGCGAUAUGCAACGGCC G A C U C C G G AA U U G G C G G G A C A U G A C A A C U A G G G G C G A G C G C U G U C C A GC C G U G G A C
Figure imgf000073_0001
CAAGGACCUCCCCGUACCUUCACAUGCCAGCUUACAUGGCACCGGGACUCAGU AAGCUUUUCACGUCGUAAUGCAUCCGGUACUGCUUCUGUGCUGCCUCGACCC ACCAUCACCAUGGAGUUCACAGGGGAUCACGCAGUGUGUACGGCAGGCUGCG C C G C CC C G C C C A G G A C U A GA G CG C C G C A C C U G U C C A G A C G U A A G G C U G C CC G C C C C G
Figure imgf000074_0001
CGCCAUCCCCAUCACCGUGAUGGAGUACACCGAGUGCCCCUACAACAAGUCCC UGGGCGUGUGCCCCAUCCGCACCCAGCCCCGCUGGUCCUACUACGACUCCUUC UCCGCCGUGUCCGAGGACAACCUGGGCUUCCUGAUGCACGCCCCCGCCUUCG C C G C C CA C A G A U G C U C U U A G G A U C U U G AC C C A G C U C A U G A C G G C G C C C A G C U C A U
Figure imgf000075_0001
CCCCUGCGGAUUCCGCCUGCAGCAUGUCUGACCUCAAAAGCCUACCAGCAAGG GGUGACUGUGGACAGCAUUGGCAUGCUGCCUCGUUUCAUUCCCGAGAAUCAA CGGACAGUGGCUCUGUAUUCCCUGAAGAUCGCAGGAUGGCAUGGGCCCAAAC G G C G C C C A G C U C A U G A C G G C G A C C C G C UC GA A C U A U A CG G CC C AC G C C U G C C C A U C C C
Figure imgf000076_0001
UGUGUCCUCCCAGAUACCACCCAAUUGGCACAUUCCUUCCAUUCAGGACGUAG CUCCGCAUCACUGA 24 HSV-2 gE CGCACCUCCUGGAAGCGCGUGACCUCCGGCGAGGACGUGGUGCUGCUGCCCG C C C C U G G A C C G C U C C C U C A G G U C C CC U C G A U C C A C C A U G G A U G CU C C CC U C G A U C C A
Figure imgf000077_0001
CGAUGGCCCUUACGCCAUGGAUGUCGUGUGGAUGAGAUUCGACGUGCCCAGC AGCUGCGCCGAGAUGAGAAUCUACGAGGCCUGCCUGUAUCACCCUCAGCUGC CCGAAUGUCUGAGCCCUGCUGAUGCCCCUUGUGCCGUUAGCAGCUGGGCCUA U G G A C UA G C G C G C G U G A G U A U C A A G G A C G C G C G C G U GA A C G U U C U C C U C C G C G C G
Figure imgf000078_0001
GCUUUACUCCGAACUGGCCUGGCGGGAUAGGGUGGCGGUGGUGAACGAGAGC CUCGUCAUCUACGGUGCUCUGGAAACCGACUCAGGACUGUAUACGCUCAGCG UUGUUGGCCUCUCCGAUGAGGCUCGACAGGUUGCCUCCGUAGUGCUGGUCGU G A G U A U C A A G G A C A G C G C G C G U G A G U A U C A A G G A C A G C C C C U G G A C C GC G C G A G C
Figure imgf000079_0001
ACCGUGAACCUGGAAUUCCAGCACGCCAGCCCCCAGCACGCCGGCCUGUACCU GUGCGUGGUGUACGUGGAUGAUCACAUCCACGCCUGGGGCCACAUGACCAUC UCCACCGCCGCCCAGUACAGAAACGCCGUGGUGGAACAGCACCUGCCCCAGAG G U C C A C U U G A C U A C G U A G U G C U G C G CC C U U G U CG AC C U U CC C G A A C C C G C C C G G C C
Figure imgf000080_0001
GGACCGCGCCCCCGUCCCCUUCGAGGAGGUGAUCGACAAGAUCAACG CCAAGGGGGUCUGUCGGUCCACGGCCAAGUACGUGCGCAACAACCUG GAGACCACCGCGUUUCACCGGGACGACCACGAGACCGACAUGGAGCU A C A G A G C C G G C C C C C C G C G G C C G G C A C G G G U A A A A C A C G U G A G A C C G C U C C A
Figure imgf000081_0001
CGGCACCACCGUGAACUGCAUCGUGGAAGAAGUGGACGCCAGAAGCG UGUACCCCUACGACGAAUUUGUGCUGGCCACCGGCGACUUCGUGUAC AUGAGCCCUUUCUACGGCUACAGAGAGGGCAGCCACACCGAGCACAC C C A C A U G C A C C G A A C C U A A A G C C C G C U C C G A C U C G U G A G A U C C C A A G U G
Figure imgf000082_0001
CGAAACCUCCUCACCACUCCCAAAUUCACUGUGGCUUGGGACUGGGU ACCCAAAAGGCCAAGUGUGUGUACCAUGACCAAGUGGCAGGAAGUC GACGAAAUGCUCCGCUCUGAGUAUGGCGGAAGCUUUCGCUUCUCUAG C A U A A C G G C G G G G C C G U C A C G G U U A C G G G U U A A U U G A C G A A U
Figure imgf000083_0001
GCAAGAGAUGCAAUGGAUAGAAUCUUUGCAAGAAGAUACAAUGCAA CCCACAUCAAAGUGGGACAGCCUCAGUACUACCAGGCAAAUGGAGGA UUUCUGAUUGCUUACCAGCCUCUGCUGUCAAAUACCCUGGCUGAACU U G U U G C A U U G G U U G A A A C C U A A C U A G G C C G C A C C A G A A G A C G C C G G C C G A G G C A
Figure imgf000084_0001
AGACCACCUCCUCGAUCGAGUUCGCCCGGCUGCAGUUUACGUAUAAC CACAUACAGCGCCACGUGAACGACAUGCUGGGGCGCAUCGCCGUCGC GUGGUGCGAGCUGCAGAACCACGAGCUGACUCUCUGGAACGAGGCCC G U A G A C G G G G G A G G C U C G C A C C C U U G C C U G C G G C U G C C U C G G A A U A G C A C U G U
Figure imgf000085_0001
GUGUUCCUGUGGCUCCCGACAACGUGAUCGUGCAGAACAGCAUGCGG GUGUCCAGCAGACCUGGCACCUGUUACUCUAGACCCCUGGUGUCCUU CAGAUACGAGGAUCAGGGCCCACUGAUCGAGGGACAGCUGGGCGAG GU G C A G C C C U G C A G U A C C C G U G U C G U G U A C G C G C G C G G A A C U U U U C C A U
Figure imgf000086_0001
ACGUUUACUUUGAGGAAUACGCUUACUCUCACCAGCUCUCAAGAGCC GACGUAACGACAGUCUCCACAUUUAUCGACUUGAACAUCACUAUGCU GGAGGAUCAUGAAUUUGUACCACUGGAGGUGUAUACGCGGCACGAA A G A U G A A G A A C C C U U G C A A U U G C A A G G A G C C G C C G C G C U U G U C A A C C G A
Figure imgf000087_0001
UACAGUGAUCAGAGCUGAUGCAAAUGCUGCAAUGUUUGCUGGACUG UGUGCUUUCUUUGAAGGAAUGGGAGAUCUGGGGAGAGCUGUGGGAA AAGUGGUGAUGGGAGUGGUGGGAGGAGUGGUGUCUGCUGUGUCUGG A U G U A C A A A A C G G U A C C C U U U C U G A C C U G U G U A G C U A C C U G C C U C C C C C G U
Figure imgf000088_0001
CCAUUCAGGACGUAGCUCCGCAUCACGCCCCAGCUGCACCCAGCAAC CCUGGCCUGAUCAUCGGUGCACUUGCCGGAUCUACCCUCGCUGUGCU GGUCAUUGGCGGGAUUGCGUUCUGGGUUCGCAGGAGAGCCCAGAUG C U C C A C G G C C G A U C G U C G C C C G C G G U U G C C G
Figure imgf000089_0001
v p g pu y p y u p t set of polyribonucleotides. In some embodiments, a first set of polyribonucleotides encode one or more glycoprotein (GP) polypeptides. [0228] In some embodiments, one or more GP polypeptides each comprise an HSV glycoprotein, a variant thereof, or an antigenic portion thereof. In some embodiments, one or more GP polypeptides comprise an HSV glycoprotein, a variant thereof, or an antigenic portion thereof as set forth in Table 4. [0229] In some embodiments, a GP polypeptide comprises an HSV glycoprotein C (gC) or antigenic portions thereof. In some embodiments, a GP polypeptide comprises an antigenic portion of HSV gC. In some embodiments, an antigenic portion of HSV gC comprises an amino acid sequence that is at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to the amino acid sequence of SEQ ID NO: 1 or a portion thereof. In some embodiments, an antigenic portion of HSV gC has an amino acid sequence that is identical to the amino acid sequence of SEQ ID NO: 1. In some embodiments, an antigenic portion of HSV gC comprises an amino acid sequence that is at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to the amino acid sequence of SEQ ID NO: 260 or a portion thereof. In some embodiments, an antigenic portion of HSV gC has an amino acid sequence that is identical to the amino acid sequence of SEQ ID NO: 260. [0230] In some embodiments, at least one polyribonucleotide of the first set of polyribonucleotides comprises a ribonucleic acid sequence that is at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to the ribonucleic acid sequence of any one of SEQ ID NO: 16-19, 147 and 274-281. [0231] In some embodiments, a GP polypeptide comprises an HSV glycoprotein D (gD) or antigenic portions thereof. In some embodiments, a GP polypeptide comprises an antigenic portion of HSV gD. In some embodiments, an antigenic portion of HSV gD comprises an amino acid sequence that is at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to the amino acid sequence of SEQ ID NO: 2 or a portion thereof. In some embodiments, an antigenic portion of HSV gD has an amino acid sequence that is identical to the amino acid sequence of SEQ ID NO: 2. [0232] In some embodiments, at least one polyribonucleotide of the first set of polyribonucleotides comprises a ribonucleic acid sequence that is at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to the ribonucleic acid sequence of any one of SEQ ID NO: 20-23, 143, and 286. [0233] In some embodiments, a GP polypeptide comprises an HSV glycoprotein E (gE) or antigenic portions thereof. In some embodiments, a polypeptide comprises an HSV gE antigens or antigenic portions thereof. In some embodiments, an antigenic portion of HSV gE comprises an amino acid sequence that is at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to the amino acid sequence of SEQ ID NO: 3 or a portion thereof. In some embodiments, an antigenic portion of HSV gE has an amino acid sequence that is identical to the amino acid sequence of SEQ ID NO: 3. [0234] In some embodiments, at least one polyribonucleotide of the first set of polyribonucleotides comprises a ribonucleic acid sequence that is at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to the ribonucleic acid sequence of any one of SEQ ID NO: 24-27 and 282-285. [0235] In some embodiments, the present disclosure provides a combination comprising a plurality of polyribonucleotides. In some embodiments, a plurality of polyribonucleotides comprise a first set of polyribonucleotides. In some embodiments, a first set of polyribonucleotides encode one or more glycoprotein (GP) polypeptides. In some embodiments, at least one polyribonucleotide of the polyribonucleotides in the first set encodes a GP polypeptide that comprises an HSV glycoprotein (gB) variant thereof, or one or more antigenic portions thereof. In some embodiments, at least one polyribonucleotide of the polyribonucleotides in the first set encodes a GP polypeptide that comprises an antigenic portion of HSV glycoprotein (gB) or an antigenic portion of a variant of HSV gB. [0236] In some embodiments, a GP polypeptide comprises an HSV glycoprotein B (gB), variant thereof, or antigenic portions thereof. In some embodiments, a GP polypeptide comprises an antigenic portion of HSV gB. In some embodiments, an HSV gB comprises an amino acid sequence that is at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to the amino acid sequence of any one of SEQ ID NO: 379 or a portion thereof. In some embodiments, an antigenic portion of HSV gB comprises an amino acid sequence that is at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to the amino acid sequence of any one of SEQ ID NO: 380 or a portion thereof. In some embodiments, an HSV gB comprises an amino acid sequence that is at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to the amino acid sequence of any one of SEQ ID NO: 381 or a portion thereof. In some embodiments, an antigenic portion of HSV gB comprises an amino acid sequence that is at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to the amino acid sequence of any one of SEQ ID NO: 382 or a portion thereof. In some embodiments, an HSV gB comprises an amino acid sequence that is at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to the amino acid sequence of any one of SEQ ID NO: 383 or a portion thereof. In some embodiments, HSV gB comprises an amino acid sequence that is at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to the amino acid sequence of any one of SEQ ID NO: 384 or a portion thereof. [0237] In some embodiments, an HSV gB comprises an amino acid sequence that is identical to the amino acid of SEQ ID NO: 379. In some embodiments, an HSV gB comprises an amino acid sequence that is identical to the amino acid of SEQ ID NO: 380. In some embodiments, an HSV gB has an amino acid sequence that is identical to the amino acid of SEQ ID NO: 381. In some embodiments, an HSV gB has an amino acid sequence that is identical to the amino acid of SEQ ID NO: 382. In some embodiments, an HSV gB has an amino acid sequence that is identical to the amino acid of SEQ ID NO: 383. In some embodiments, an HSV gB has an amino acid sequence that is identical to the amino acid of SEQ ID NO: 384. [0238] Further, an issue with current HSV compositions (e.g., immunogenic compositions, e.g., vaccines) is that they have not been able to fully leverage antigenic potential of HSV gB (HSV-1 gB, HSV-2 gB, or both). One reason for these results is that HSV gB can be unstable. In particular, cell entry of enveloped viruses requires specialized viral proteins that mediate fusion with the host membrane. During this process, the viral proteins, including gB, undergo substantial structural rearrangements from a metastable prefusion conformation to a stable postfusion conformation (FIG.16). This metastability renders the herpes simplex virus (e.g., HSV-1, HSV-2, or both) fusion gB highly unstable. [0239] As used herein, the term “stable”, when applied to glycoprotein B, means that glycoprotein B maintains one of more aspects of a physical structure (e.g., maintains a specific conformation) and/or activity for a specific period of time. In some embodiments, a stable glycoprotein B has been modified (e.g., certain mutations) so that its structure is stabilized. In some embodiments, a stable glycoprotein B structure is maintained for a specific period of time. In some embodiments, a stable glycoprotein B is in a stable prefusion conformation. In some embodiments, a stable glycoprotein B is in a stable postfusion conformation. In some embodiments, a stable glycoprotein B maintains a biological relevant activity (e.g., antigenic potential). In some embodiments, a stable glycoprotein B is in a stable prefusion conformation and maintains antigenic potential. [0240] The present disclosure encompasses a recognition that stabilization of gB or antigenic portions thereof can be useful or advantageous for eliciting an immune response. The present disclosure further provides the recognition that stabilization of HSV gB or antigenic portions thereof can be particularly advantageous for use, e.g., in preventing or treating HSV, e.g., in HSV antigen constructs and/or HSV compositions (e.g., immunogenic compositions, e.g., vaccines) as further disclosed herein. Accordingly, the present disclosure provides certain mutations that can stabilize HSV gB or antigenic portions thereof. [0241] Provided herein is a polyribonucleotide encoding a GP polypeptide that comprises an HSV glycoprotein B (gB), variant thereof, or one or more antigenic portions thereof. In some embodiments, an HSV gB or antigenic portion thereof comprises one or more mutations that stabilize the HSV gB or antigenic portion thereof relative to a comparable HSV gB or antigenic portion thereof that does not comprise the one or more mutations. In some embodiments, one or more mutations are one or more amino acid substitutions. In some embodiments, one or more amino acid substitutions comprise 120C, 181C, 238C, 251C, 259C, 290C, 291C, 391C, 526C, 571C, 610C, 630C, 636C, 676C, 677C, 680C, 714C, 718C, 725C, 758C, and combinations thereof, wherein the numbering is with reference to SEQ ID NO: 379. [0242] In some embodiments, an HSV gB comprises an amino acid sequence according to SEQ ID NO: 379 or antigenic portion thereof, except that the HSV gB or antigenic portion thereof comprises one or more mutations comprising 120C, 181C, 238C, 251C, 259C, 290C, 291C, 391C, 526C, 571C, 610C, 630C, 636C, 676C, 677C, 680C, 714C, 718C, 725C, 758C, and combinations thereof. [0243] In some embodiments, an HSV gB has an amino acid sequence that (i) is at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the amino acid SEQ ID NO: 379 and (ii) comprises one or more mutations, where the one or more mutations comprise 120C, 181C, 238C, 251C, 259C, 290C, 291C, 391C, 526C, 571C, 610C, 630C, 636C, 676C, 677C, 680C, 714C, 718C, 725C, 758C, and combinations thereof. [0244] In some embodiments, an HSV gB has an amino acid sequence that comprises one or more mutations, where the one or more mutations comprise: (a) 120C and 677C, (b) 181C and 725C, (c) 238C and 610C, (d) 251C and 718C, (e) 259C and 758C, (f) 290C and 680C, (g) 291C and 636C, (h) 391C and 526C, (i) 571C and 676C, (j) 571C and 680C, and/or (k) 630C and 714C, where the numbering is with reference to SEQ ID NO: 379. [0245] In some embodiments, an HSV gB has an amino acid sequence that (i) is at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the amino acid SEQ ID NO: 379 and (ii) comprises one or more mutations, where the one or more mutations comprise: (a) 120C and 677C, (b) 181C and 725C, (c) 238C and 610C, (d) 251C and 718C, (e) 259C and 758C, (f) 290C and 680C, (g) 291C and 636C, (h) 391C and 526C, (i) 571C and 676C, (j) 571C and 680C, and/or (k) 630C and 714C, where the numbering is with reference to SEQ ID NO: 379. [0246] In some embodiments, an HSV gB or antigenic portion thereof does not comprise an 516P mutation, where the numbering is with reference to SEQ ID NO: 379. [0247] In some embodiments, the present disclosure provides a plurality of polyribonucleotides, wherein the plurality of polyribonucleotides comprises a first set of polyribonucleotides that comprises: (i) a polyribonucleotide that encoding an antigenic portion of HSV gC as provided herein, (ii) a polyribonucleotide that encoding an antigenic portion of HSV gD as provided herein, (iii) a polyribonucleotide that encoding an antigenic portion of HSV gE as provided herein, and (iv) a polyribonucleotide that encoding an antigenic portion of HSV gB or variant thereof as provided herein. [0248] In some embodiments, a first set of polyribonucleotides further comprises a polyribonucleotide encoding an HSV glycoprotein G (gG) or an antigenic portion thereof, a polyribonucleotide encoding an HSV glycoprotein H (gH) or an antigenic portion thereof, a polyribonucleotide encoding an HSV glycoprotein I (gI) or an antigenic portion thereof, a polyribonucleotide encoding an HSV glycoprotein L (gL) or an antigenic portion thereof, or a combination thereof. B. HSV T-cell antigens and antigenic portions thereof [0249] The present disclosure provides combinations comprising a first set of polyribonucleotides. In some embodiments, combinations provided herein further comprises a second set of polyribonucleotides. In some embodiments, a second set of polyribonucleotides encode one or more T-cell string polypeptides. In some embodiments, one or more T-cell string polypeptides comprises one or more HSV T-cell antigens or antigenic portions thereof. [0250] In some embodiments, one or more HSV T-cell antigens comprises (i) one or more HSV RS1 polypeptides or antigenic portions thereof, (ii) one or more HSV RL2 polypeptides or antigenic portions thereof, (iii) one or more HSV UL1 polypeptides or antigenic portions thereof, (iv) one or more HSV UL5 polypeptides or antigenic portions thereof, (v) one or more HSV UL9 polypeptides or antigenic portions thereof, (vi) one or more HSV UL19 polypeptides or antigenic portions thereof, (vii) one or more HSV UL21 polypeptides or antigenic portions thereof, (viii) one or more HSV UL25 polypeptides or antigenic portions thereof, (ix) one or more HSV UL27 polypeptides or antigenic portions thereof, (x) one or more HSV UL29 polypeptides or antigenic portions thereof, (xi) one or more HSV UL30 polypeptides or antigenic portions thereof, (xii) one or more HSV UL39 polypeptides or antigenic portions thereof, (xiii) one or more HSV UL40 polypeptides or antigenic portions thereof, (xiv) one or more HSV UL46 polypeptides or antigenic portions thereof, (xv) one or more HSV UL47 polypeptides or antigenic portions thereof, (xvi) one or more HSV UL48 polypeptides or antigenic portions thereof, (xvii) one or more HSV UL49 polypeptides or antigenic portions thereof, (xviii) one or more HSV UL52 polypeptides or antigenic portions thereof, (xix) one or more HSV UL54 polypeptides or antigenic portions thereof, (xx) one or more HSV US10 polypeptides or antigenic portions thereof, (xxi) one or more HSV US12 polypeptides or antigenic portions thereof, (xxii) one or more HSV UL26 polypeptides or antigenic portions thereof, (xxiii) one or more HSV UL50 polypeptides or antigenic portions thereof, or (xxiv) a combination thereof. [0251] Example antigen amino acid sequences are shown in Table 7 below. Table 7: Example amino acid antigen sequences SEQ Antigen Strain Amino Acid Sequence ID NO 476 UL1 HG52 MGFVCLFGLVVMGAWGAWGGSQATEYVLRSVIAKEVGDILRVPCMRTPADDVS F R G G V A G L D G G V A G L D G G V A G L D G A P S F A L R Y T T
Figure imgf000094_0001
GPULIEGQLGENNELRLTRDALEPCTVGHRRYFIFGGGYVYFEEYAYSHQLSRADVT TVSTFIDLNITMLEDHEFVPLEVYTRHEIKDSGLLDYTEVQRRNQLHDLRFADIDTVI RADANAAMFAGLCAFFEGMGDLGRAVGKVVMGVVGGVVSAVSGVSSFMSNPFGAL G R A P S F A L R Y T T T R A G Y T G V D V A D G E R E I N L E H R A D V A D G E R E
Figure imgf000095_0001
GQLGENNELRLTRDALEPCTVGHRRYFIFGGGYVYFEEYAYSHQLSRADVTTVSTFI DLNITMLEDHEFVPLEVYTRHEIKDSGLLDYTEVQRRNQLHDLRFADIDTVIRADAN AAMFAGLCAFFEGMGDLGRAVGKVVMGVVGGVVSAVSGVSSFMSNPFGALAVGLL E V T CI P A I D L C N L V G V N R S Q Q L A V T CI P A I D L C N L V G V N Y A R P Q K
Figure imgf000096_0001
486 UL29 MS MDTKPKTTTTVKVPPGPMGYVYGRACPAEGLELLSLLSARSGDADVAVAPLIVGLTV ESGFEANVAAVVGSRTTGLGGTAVSLKLMPSHYSPSVYVFHGGRHLAPSTQAPNLT RLCERARRHFGFSDYAPRPCDLKHETTGDALCERLGLDPDRALLYLVITEGFREAVCI P A I D L C N L V G V N R S Q Q L G Y V R D S FL R S M T S L A C T R P D L Y V R T G V R F
Figure imgf000097_0001
EEWMRSKEVDLDFGLTERLREHEAQLMILAQALNPYDCLIHSTPNTLVERGLQSALK YEEFYLKRFGGHYMESVFQMYTRIAGFLACRATRGMRHIALGRQGSWWEMFKFFF HRLYDHQIVPSTPAMLNLGTRNYYTSSCYLVNPQATTNQATLRAITGNVSAILARNG A L D VL HI L L L T Y V R T G V A C A N A K A SV M M S A T F A T T E F A T T E E V S
Figure imgf000098_0001
IPEKFILMILIEGVFFAASFAAIAYLRINNLLRVTCQSNDLISRDEAVHTTASCYIYNNY LGGHAKPEAARVYRLFREAVDIEIGFIRSQAPTDSSILSPGALAAIENYVRFSADRLLG LIHMQPLYSAPAPDASFPLSLMSTDKHTNFFECRSTSYAGAVVNDL Q Q L P S G F P A D H Y L Q Q L P S G F P A D A I LL Q G R S G Q K PP P D EI D E V A G A R A
Figure imgf000099_0001
RYAGRMTYIATGALLARFNPGAVKCVLPREAAFAGRVLDVLAVLAEQTVQW LSVVVGARLHPHSAHPAFADVEQEALFRALPLGSPGVVAAEHEALGDTAARRLLATS GLNAVLGAAVYALHTALATVTLKYALACGDARRRRDDAAAARAVLATGLILQRLLGL A F E V A G A R A W S L A F E V A G A R A W S L A F P L R F N D A P L R F N D A
Figure imgf000100_0001
501 UL48 MS MDLLVDDLFADADGVSPPPPRPAGGPKNTPAAPPLYATGRLSQAQLMPSPPMPVPP AALFNRLLDDLGFSAGPALCTMLDTWNEDLFSGFPTNADMYRECKFLSTL PSDVIDWGDAHVPERSPIDIRAHGDVAFPTLPATRDELPSYYEAMAQFFR F N D A F G H D F A K D A F A K D A P V D R V Y A F D A G G D T T M L L E P
Figure imgf000101_0001
LVLRDDADAGPPPQIRWASAAGRAGTVLAAAGGGVEVVGTAAGLATPPRREPVDM DAELEDDDDGLFGE 506 RS1 333 MSAEQRKKKKTTTTTQGRGAEVAMADEDGGRLRAAAETTGGPGSPDPADGPPPTP V A R R T A G A A V V P V L E A N F G P N P D A A Y N R G L D F P A R S R C P E
Figure imgf000102_0001
FCARALLEPDGDAPPLVLRDDADAGPPPQIRWASAAGRAGTVLAAAGGGVEVVGAA AGLATPPRREPVDMDAELEDDDDGLFGE 508 RL2 HG52 MEPRPGTSSRADPGPERPPRQTPGTQPAAPHAWGMLNDMQWLASSDSEEETEVGI G T T G S G A V R A K V P P GI G N L A R SS H A G G T V N GI G N L A R S H A D A A R Q VL P
Figure imgf000103_0001
ASLEDLQRRDLTYYWEVILDITKRALAAHGGEDARNEFHALTALEQTLGLGQGALTR LASVTHGALPAFTRSNIIVIDEAGLLGRHLLTTVVYCWWMINALYHTPQYAGRLRPV LVCVGSPTQTASLESTFEHQKLRCSVRQSENVLTYLICNRTLREYTRLSHSWAIFINN R Q Q F N L Q N Q VL P R V N R Q Q F N L Q N Q VL P R V N R Q Q F N L Q N L R LI L L S
Figure imgf000104_0001
STVSFAEVVARFCRQFTDRVLLLHSLTPPGDVTTWGRYRVVIYTTVVTVGL SFDPPHFDSMFAYVKPMNYGPDMVSVYQSLGRVRTLRKGELLIYMDGSG ARSEPVFTPMLLNHVVSASGQWPAQFSQVTNLLCRRFKGRCDASHADAAQARGS R R L L Q R L T P L VL Q G S L A S M G R L T P L VL Q G S L A S M G R L P V D Q E L N R
Figure imgf000105_0001
QRLAHGRVRWVAECQMTAEQFMQPDNANLALELHPAFDFFAGVADVELPGGEVPP AGPGAIQATWRVVNGNLPLALCPVAFRDARGLELGVGRHAMAPATIAAVRGAFEDR SYPAVFYLLQAAIHGNEHVFCALARLVTQCITSYWNNTRCAAFVNDYSLVSYIVTYL P T VV A Y A F D F T L Q A C IK E D E P D YL P Q V H F E R G H A L E L Q A C IK E D E P
Figure imgf000106_0001
PAGPGAIQATWRVVNGNLPLALCPVAFRDARGLELGVGRHAMAPATIAAVRGAFED RSYPAVFYLLQAAIHGSEHVFCALARLVTQCITSYWNNTRCAAFVNDYSLVSYIVTYL GGDLPEECMAVYRDLVAHVEALAQLVDDFTLPGPELGGQAQAELNHLMRDPALLPP Q V H F E R G H A L E A R G L D D R SS A R G L D D R SS N A R G L D D R SS N T D A
Figure imgf000107_0001
QLHERFMDAITPAGTVITLLGLTPEGHRVAVHVYGTRQYFYMNKAEVDRHLQCRAP RDLCERLAAALRESPGASFRGISADHFEAEVVERADVYYYETRPTLYYRVFVRSGRAL AYLCDNFCPAIRKYEGGVDATTRFILDNPGFVTFGWYRLKPGRGNAPAQPRPPTAFG Q G I P V SL C S K E T L A M T D A P AL G Q G I P V L C S K E T L A M T D A P AL G Q
Figure imgf000108_0001
MLLAFMTFVKQYGPEFVTGYNIINFDWPFVLTKLTEIYKVPLDGYGRMNG RGVFRVWDIGQSHFQKRSKIKVNGMVNIDMYGIITDKVKLSSYKLNAVAEAVLK DKKKDLSYRDIPAYYASGPAQRGVIGEYCVQDSLLVGQLFFKFLPHLELSAVARLAGI P F S L M K E V P C L V Q L A L P M A L G G H A F F R D P A R D E D E E T P E Q VI A
Figure imgf000109_0001
LEHFGRRETLTEVLGRYDVRPDAGETVEGFASELLGRIVACIEAHFPEHAREYQAVSV RRAVIKDDWVLLQLIPGRGALNQSLSCLRFKHGRASRATARTFLALSVGTNNRLCAS LCQQCFATKCDNNRLHTLFTVDAGTPCSRSAPSSTSRPSSS D P A R D E D E E T P A VI A SV S G R R SL T C G R S L P A T C G R S L P A T
Figure imgf000110_0001
ARLANRVERGVSEIDYTTVGVGAGETMHFYIPGACMAGLIEILDTHRQEC SSRVCELTASHTIAPLYVHGKYFYCNSLF 532 US1 HG52 MADIPPDPPALNTTPANHAPPSPPPGSRKRRRPVLPSSSESEGKPDTESESSSTESSE V D R L S P E D L S P E V D R L S P E V D R L S P E D P M Y A G L T L R E H G G
Figure imgf000111_0001
DTGTPAPASGERAPPNSTRSASESRHRLTVAQVIQIAIPASIIAFVFLGSCICFIHRCQ RRYRRPRGQIYNPGGVSCAVNEAAMARLGAELRSHPNTPPKPRRRSSSSTTMPSLT SIAEESEPGPVVLLSVSPRPRSGPTAPQEV E H G G R T E H G G Q T A P S A S A G G W N E G A A H A P S A S A G R L F Q V
Figure imgf000112_0001
IPEVSHVRGVTVHMETPEAILFAPGETFGTNVSIHAIAHDDGPYAMDVVWMRFDVP SSCAEMRIYEACLYHPQLPECLSPADAPCAVSSWAYRLAVRSYAGCSRTTPPPRCFA EARMEPVPGLAWLASTVNLEFQHASPQHAGLYLCVVYVDDHIHAWGHMTISTAAQ A P VL P P P A R L A L G A F L P P P A R L A L G A F L P P P A R L A L G A F
Figure imgf000113_0001
LRYTPAGEVMSVLLVDTDATQQQLAQGPVAGTPNVFSSDVPSVALLLFPNGTVIHLL AFDTLPIATIAPGFLAASALGVVMITAALAGILRVVRTCVPFLWRRE 548 UL22 G MGPGLWVVMGVLVGVAGGHDTYWTEQIDPWFLHGLGLARTYWRDTNTGRLWLP P P A R L A L G V V L V C T AL P T A R A Q V P R D R A T C D LP P LF D L W L
Figure imgf000114_0001
554 US10 G MIRRRGNVEIRVYYESVRPSRSRSHLKPSDHQEFPGHHVSPGSPGFPESPGNREFHD LPENPGSRAYPGTRDPHDPHGCPGSLDPHGSPDPSSPRQRTYVLPRVGIHNAPASD TRAPKRAHSRHRADRPPESPGSELYPLNAQALAHLQMLPADHRAFFRTVIEVSRLCA Q G P G A A H P P G A A H P P E A I L G P S TI Y A E A I L G P P S TI Y A E A I
Figure imgf000115_0001
GRRLGTIVTYDTSLDAAIAPFRHLDPATREGVRREAAEAELALAGRTWAPGVEAL THTLLSTAVNNMMLRDRWSLVAERRRQAGIAGHTYLQASEKFKIWGAESAPAPERG YKTGAPGAMDTSPAASVPAPQVAVRARQVASSSSSSSSFPAPADMNPVSASGAPAP P S TI Y A E A I L G P P TI Y A
Figure imgf000116_0001
In some embodiments, a T-cell string polypeptide comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 T-cell antigens or antigenic portions thereof, HSV-2 T-cell antigens or antigenic portions thereof, or a combination thereof). In some embodiments, such T-cell string polypeptides can be particularly useful in effective vaccination. [0253] In some embodiments, a T-cell string polypeptide includes a plurality of HSV T-cell antigens or antigenic portions thereof (e.g., a plurality of HSV antigens that are or include one or more T cell and/or B cell antigens for HSV). As disclosed herein, T cell antigens include, e.g., CD4 T cell antigens and/or CD8 T cells. In some embodiments, an HSV antigen is a T cell antigen. In some embodiments, an HSV antigen is a B cell antigen. [0254] In some embodiments, an HSV T-cell antigen comprises at least one of UL1, UL21, UL27, UL29, UL39, UL40, UL46, UL47, UL48, UL49, RS1, RL2, UL5, UL9, UL19, UL25, UL30, UL52, US1, US7, US8, UL22, US10, US12, UL26, UL50, and/or UL54 or antigenic portion thereof. [0255] In some embodiments, a T-cell polypeptide comprises a plurality of (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19 of) UL1, UL21, UL27, UL29, UL39, UL40, UL46, UL47, UL48, UL49, RS1, RL2, UL5, UL9, UL19, UL25, UL30, UL52, US1, US7, US8, UL22, US10, US12, UL26, UL50, and/or UL54 or antigenic portions thereof. [0256] In some embodiments, a T-cell polypeptide comprises and/or encodes UL54, UL29, UL39, UL9, UL30a (UL30.1), UL40, UL5a (UL5.1), UL21, and/or UL46 or fragments thereof. [0257] In some embodiments, a T-cell polypeptide comprises and/or encodes UL54, UL29, UL40, and/or UL47 or fragments thereof. [0258] In some embodiments, a T-cell polypeptide comprises and/or encodes at least UL47 or fragments thereof. [0259] In some embodiments, a T-cell polypeptide comprises and/or encodes at least UL40 or fragments thereof. [0260] In some embodiments, a T-cell polypeptide comprises and/or encodes one or more of RL2, UL54, UL9, UL39, UL29, UL5, UL40, UL30, UL49, UL46 and/or UL21 or fragments thereof. [0261] The UL1 open reading frame encodes HSV gL (also referred to herein as UL1 polypeptide). In some embodiments, an HSV antigen (e.g., a T-cell or B cell antigen for HSV) is or includes a UL1 polypeptide or fragment thereof. In various embodiments, a UL1 polypeptide or fragment thereof has at least 80% sequence identity with a UL1 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity). Examples of UL1 polypeptides known in the art include UL1 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains. In some embodiments, a UL1 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 476. [0262] In some embodiments, an HSV antigen (e.g., a T cell or B cell antigen for HSV) is or includes a UL21 polypeptide or fragment thereof. In various embodiments, a UL21 polypeptide or fragment thereof has at least 80% sequence identity with a UL21 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity). Examples of UL21 polypeptides known in the art include UL21 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains. In some embodiments, a UL21 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 477, 478 and/or 479. [0263] The UL27 open reading frame encodes HSV gB (also referred to herein as UL27 polypeptide). In some embodiments, an HSV antigen (e.g., a T cell or B cell antigen for HSV) is or includes a UL27 polypeptide or fragment thereof. In various embodiments, a UL27 polypeptide or fragment thereof has at least 80% sequence identity with a UL27 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity). Examples of UL27 polypeptides known in the art include UL27 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains. In some embodiments, a UL27 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 480, 481, 482 and/or 483. [0264] In some embodiments, an HSV antigen (e.g., a T cell or B cell antigen for HSV) is or includes a UL29 polypeptide or fragment thereof. In various embodiments, a UL29 polypeptide or fragment thereof has at least 80% sequence identity with a UL29 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity). Examples of UL29 polypeptides known in the art include UL29 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains. In some embodiments, a UL29 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 484, 485, and/or 486. [0265] In some embodiments, an HSV antigen (e.g., a T cell or B cell antigen for HSV) is or includes a UL39 polypeptide or fragment thereof. In various embodiments, a UL39 polypeptide or fragment thereof has at least 80% sequence identity with a UL39 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity). Examples of UL39 polypeptides known in the art include UL39 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains. In some embodiments, a UL39 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 487, 488 and/or 489. [0266] In some embodiments, an HSV antigen (e.g., a T cell or B cell antigen for HSV) is or includes a UL40 polypeptide or fragment thereof. In various embodiments, a UL40 polypeptide or fragment thereof has at least 80% sequence identity with a UL40 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity). Examples of UL40 polypeptides known in the art include UL40 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains. In some embodiments, a UL40 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 490, 491 and/or 492. [0267] In some embodiments, an HSV antigen (e.g., a T cell or B cell antigen for HSV) is or includes a UL46 polypeptide or fragment thereof. In various embodiments, a UL46 polypeptide or fragment thereof has at least 80% sequence identity with a UL46 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity). Examples of UL46 polypeptides known in the art include UL46 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains. In some embodiments, a UL46 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 493, 494 and/or 495. [0268] In some embodiments, an HSV antigen (e.g., a T cell or B cell antigen for HSV) is or includes a UL47 polypeptide or fragment thereof. In various embodiments, a UL47 polypeptide or fragment thereof has at least 80% sequence identity with a UL47 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity). Examples of UL47 polypeptides known in the art include UL47 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains. In some embodiments, a UL47 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 496, 497 and/or 498. [0269] In some embodiments, an HSV antigen (e.g., a T cell or B cell antigen for HSV) is or includes a UL48 polypeptide or fragment thereof. In various embodiments, a UL48 polypeptide or fragment thereof has at least 80% sequence identity with a UL48 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity). Examples of UL48 polypeptides known in the art include UL48 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains. In some embodiments, a UL48 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 499, 500 and/or 501. [0270] In some embodiments, an HSV antigen (e.g., a T cell or B cell antigen for HSV) is or includes a UL49 polypeptide or fragment thereof. In various embodiments, a UL49 polypeptide or fragment thereof has at least 80% sequence identity with a UL49 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity). Examples of UL49 polypeptides known in the art include UL49 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains. In some embodiments, a UL49 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 502, 503 and/or 504. [0271] In some embodiments, an HSV antigen (e.g., a T cell or B cell antigen for HSV) is or includes a RS1 polypeptide or fragment thereof. In various embodiments, a RS1 polypeptide or fragment thereof has at least 80% sequence identity with a RS1 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity). Examples of RS1 polypeptides known in the art include RS1 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains. In some embodiments, a RS1 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 505, 506 and/or 507. [0272] In some embodiments, an HSV antigen (e.g., a T cell or B cell antigen for HSV) is or includes a RL2 polypeptide or fragment thereof. In various embodiments, a RL2 polypeptide or fragment thereof has at least 80% sequence identity with a RL2 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity). Examples of RL2 polypeptides known in the art include RL2 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains. In some embodiments, a RL2 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 508, 509 and/or 510. [0273] In some embodiments, an HSV antigen (e.g., a T cell or B cell antigen for HSV) is or includes a UL5 polypeptide or fragment thereof. In various embodiments, a UL5 polypeptide or fragment thereof has at least 80% sequence identity with a UL5 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity). Examples of UL5 polypeptides known in the art include UL5 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains. In some embodiments, a UL5 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 511, 512 and/or 513. [0274] In some embodiments, an HSV antigen (e.g., a T cell or B cell antigen for HSV) is or includes a UL9 polypeptide or fragment thereof. In various embodiments, a UL9 polypeptide or fragment thereof has at least 80% sequence identity with a UL9 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity). Examples of UL9 polypeptides known in the art include UL9 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains. In some embodiments, a UL9 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 514, 515 and/or 516. [0275] In some embodiments, an HSV antigen (e.g., a T cell or B cell antigen for HSV) is or includes a UL19 polypeptide or fragment thereof. In various embodiments, a UL19 polypeptide or fragment thereof has at least 80% sequence identity with a UL19 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity). Examples of UL19 polypeptides known in the art include UL19 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains. In some embodiments, a UL19 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 517, 518 and/or 519. [0276] In some embodiments, an HSV antigen (e.g., a T cell or B cell antigen for HSV) is or includes a UL25 polypeptide or fragment thereof. In various embodiments, a UL25 polypeptide or fragment thereof has at least 80% sequence identity with a UL25 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity). Examples of UL25 polypeptides known in the art include UL25 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains. In some embodiments, a UL25 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 520, 521 and/or 522. [0277] In some embodiments, an HSV antigen (e.g., a T cell or B cell antigen for HSV) is or includes a UL30 polypeptide or fragment thereof. In various embodiments, a UL30 polypeptide or fragment thereof has at least 80% sequence identity with a UL30 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity). Examples of UL30 polypeptides known in the art include UL30 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains. In some embodiments, a UL30 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 523, 524 and/or 525. [0278] In some embodiments, an HSV antigen (e.g., a T cell or B cell antigen for HSV) is or includes a UL52 polypeptide or fragment thereof. In various embodiments, a UL52 polypeptide or fragment thereof has at least 80% sequence identity with a UL52 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity). Examples of UL52 polypeptides known in the art include UL52 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains. In some embodiments, a UL52 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 526, 527 and/or 528. [0279] In some embodiments, an HSV antigen (e.g., a T cell or B cell antigen for HSV) is or includes a US1 polypeptide or fragment thereof. In various embodiments, a US1 polypeptide or fragment thereof has at least 80% sequence identity with a US1 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity). Examples of US1 polypeptides known in the art include US1 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains. In some embodiments, a US1 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 532, 533, 534, 535 and/or 536. [0280] The US7 open reading frame encodes HSV gI (also referred to herein as US7 polypeptide). In some embodiments, an HSV antigen (e.g., a T cell or B cell antigen for HSV) is or includes a US7 polypeptide or fragment thereof. In various embodiments, a US7 polypeptide or fragment thereof has at least 80% sequence identity with a US7 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity). Examples of US7 polypeptides known in the art include US7 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains. In some embodiments, a US7 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 537, 538, 539 and/or 540. [0281] The US8 open reading frame encodes HSV gE (also referred to herein as US8 polypeptide). In some embodiments, an HSV antigen is (e.g., a T cell or B cell antigen for HSV) or includes a US8 polypeptide or fragment thereof. In various embodiments, a US8 polypeptide or fragment thereof has at least 80% sequence identity with a US8 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity). Examples of US8 polypeptides known in the art include US8 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains. In some embodiments, a US8 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 541, 542, 543 and/or 544. [0282] The UL22 open reading frame encodes HSV gH (also referred to herein as UL22 polypeptide). In some embodiments, an HSV antigen is (e.g., a T cell or B cell antigen for HSV) or includes a UL22 polypeptide or fragment thereof. In various embodiments, a UL22 polypeptide or fragment thereof has at least 80% sequence identity with a UL22 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity). Examples of UL22 polypeptides known in the art include UL22 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains. In some embodiments, a UL22 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 545, 546, 547 and/or 548. [0283] In some embodiments, an HSV antigen (e.g., a T cell or B cell antigen for HSV) is or includes a US10 polypeptide or fragment thereof. In various embodiments, a US10 polypeptide or fragment thereof has at least 80% sequence identity with a US10 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity). Examples of US10 polypeptides known in the art include US10 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains. In some embodiments, a US10 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 551, 552, 553 and/or 554. [0284] In some embodiments, an HSV antigen (e.g., a T cell or B cell antigen for HSV) is or includes a US12 polypeptide or fragment thereof. In various embodiments, a US12 polypeptide or fragment thereof has at least 80% sequence identity with a US12 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity). Examples of US12 polypeptides known in the art include US12 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains. In some embodiments, a US12 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NO: 555. [0285] In some embodiments, an HSV antigen (e.g., a T cell or B cell antigen for HSV) is or includes a UL26 polypeptide or fragment thereof. In various embodiments, a UL26 polypeptide or fragment thereof has at least 80% sequence identity with a UL26 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity). Examples of UL26 polypeptides known in the art include UL26 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains. In some embodiments, a UL26 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 558, 559, 560, and/or 561. [0286] In some embodiments, an HSV antigen (e.g., a T cell or B cell antigen for HSV) is or includes a UL50 polypeptide or fragment thereof. In various embodiments, a UL50 polypeptide or fragment thereof has at least 80% sequence identity with a UL50 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity). Examples of UL50 polypeptides known in the art include UL50 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains. In some embodiments, a UL50 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 556 and/or 557. [0287] In some embodiments, an HSV antigen (e.g., a T cell or B cell antigen for HSV) is or includes a UL54 polypeptide or fragment thereof. In various embodiments, a UL54 polypeptide or fragment thereof has at least 80% sequence identity with a UL54 amino acid sequence set forth in Table 7 or otherwise known in the art, or a corresponding fragment thereof (e.g., at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity). Examples of UL54 polypeptides known in the art include UL54 polypeptides encoded by known HSV strains such as, without limitation, HG52, G, 333, and MS strains. In some embodiments, a UL54 polypeptide or fragment thereof has at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with an amino acid sequence as set forth in SEQ ID NOs: 529, 530, and/or 531. [0288] In some embodiments, at least one of the polypeptides in the second set of polynucleotides encode one or more T-cell string polypeptides. In some embodiments, a T-cell string polypeptide can include one or more HSV antigens including one or more T cell antigens (e.g., CD4 and/or CD8 T cell antigens) for HSV of the present disclosure and one or more HSV antigens that is not a T cell antigen of the present disclosure. In some embodiments, a polyribonucleotide encoding a T-cell string polypeptide can include and/or encode one or more HSV antigens including one or more B cell antigens for HSV of the present disclosure and one or more HSV antigens that is not a B cell antigen of the present disclosure. In some embodiments, a polyribonucleotide encoding a T-cell string polypeptide can include and/or encode one or more HSV antigens including one or more T cell antigens for HSV of the present disclosure and one or more HSV antigens that is a B cell antigen for HSV (e.g., an antigen that is or includes a B cell epitope disclosed herein or otherwise known in the art). In some embodiments, a polyribonucleotide encoding a T-cell string polypeptide can include and/or encode one or more HSV antigens including one or more T cell antigens for HSV of the present disclosure and one or more HSV antigens selected from HSV glycoproteins or fragments thereof. In some embodiments, a polyribonucleotide encoding a T-cell string polypeptide can include and/or encode one or more HSV antigens including one or more T cell antigens for HSV of the present disclosure and one or more HSV antigens selected from an HSV gD protein or an antigenic fragment thereof, an HSV gB protein or an antigenic fragment thereof, an HSV gE protein or an antigenic fragment thereof, an HSV gG protein or an antigenic fragment thereof, an HSV gI protein or an antigenic fragment thereof, an HSV gH protein or an antigenic fragment thereof, an HSV gL protein or an antigenic fragment thereof, an HSV ICP4 protein or an antigenic fragment thereof, or an ICP8 protein or an antigenic fragment thereof. [0289] In various embodiments, a polyribonucleotide encoding a T-cell string polypeptide can be present in a composition for delivery of the HSV antigen construct to a subject. In various embodiments, an HSV antigen construct can be present in a composition for delivery of one or more HSV antigens and/or epitopes to a subject. In various embodiments, a polyribonucleotide encoding a T-cell string polypeptide can be or include an RNA molecule that encodes one or more antigens and/or epitopes. [0290] Compositions for delivery of a polyribonucleotide encoding a T-cell string polypeptide, in some embodiments, advantageously include, for example, one or more B cell antigens for HSV and one or more T cell antigens (e.g., CD4 and/or CD8 T cell antigens) for HSV. Without wishing to be bound by any particular scientific theory, and without suggesting other embodiments are also advantageous, combination of B cell antigens and T cell antigens can be advantageous in promoting immune system defenses against HSV at multiple lifecycle points include without limitation prior to cellular entry and after cellular entry. [0291] Among other things, the present disclosure provides an insight that many prior strategies for developing pharmaceutical compositions (e.g., immunogenic compositions, e.g., vaccines) for treatment of and/or protection from viral infection have focused primarily, or even almost exclusively, on development of neutralizing antibodies that target surface glycoproteins. The present disclosure identifies a problem with such strategies including, for example, that they may fail to appreciate value or even criticality of ensuring that an induced immune response includes significant T cell activity (in some embodiments, CD4 T cell activity, in some embodiments CD8 T cell activity, in some embodiments, both). [0292] Alternatively or additionally, the present disclosure provides an insight that consideration of expression of HSV proteins (e.g., at particular periods of the HSV life cycle and/or in particular tissues or compartments of an infected subject) can improve composition (e.g., immunogenic composition, e.g., vaccine) effectiveness. [0293] In some embodiments, the present disclosure provides technologies for identifying, selecting, and/or characterizing HSV protein sequences (e.g., HSV-1 protein sequences, HSV-2 protein sequences, or a combination thereof), and combinations thereof, particularly useful for inclusion in pharmaceutical compositions (e.g., immunogenic compositions, e.g., vaccines) as described herein. [0294] In some embodiments, pharmaceutical compositions (e.g., immunogenic compositions, e.g., vaccines) that comprise or deliver CD4 and CD8 antigen(s) of one or more HSV proteins (e.g., HSV-1 proteins, HSV-2 proteins, or a combination thereof), e.g., in addition to one or more B cell antigens. Among other things, the present disclosure provides HSV antigen constructs (e.g., HSV-1 antigen constructs, HSV-2 antigen constructs, or a combination thereof) and compositions (e.g., pharmaceutical compositions, e.g., immunogenic compositions, e.g., vaccines) that comprise and/or deliver antigen constructs that induce both neutralizing antibodies and T cells (e.g., CD4 and/or CD8 T cells). Such neutralizing antibodies and T cells (e.g., CD4 and/or CD8 T cells) can target, for example, an HSV glycoprotein and, in some embodiments, one or more additional HSV proteins. In some embodiments, the present disclosure provides such polyribonucleotides and compositions that induce particularly strong neutralizing antibody responses and/or particularly diverse T cell responses (e.g., targeting multiple T cell antigens). [0295] In some embodiments, the present disclosure provides such polyribonucleotides and compositions that induce robust B cell responses. In some embodiments, a B cell response includes the production of a diverse, specific repertoire of antibodies. [0296] In some embodiments, the present disclosure provides such polyribonucleotides and compositions that induce T cell and B cell responses to HSV antigens and/or epitopes. [0297] The present disclosure provides the recognition, for example, that polyribonucleotides and compositions comprising RNA molecules as described herein (e.g., encoding for one or more HSV (e.g., HSV-1 and/or HSV-2) antigens and/or epitopes) may result in a higher degree of antigen presentation to various immune system components and/or pathways. In some embodiments, administration of such constructs or compositions may induce T cell and/or B cell responses. The present disclosure provides the insight that, e.g., in some embodiments in which T cell and B cell responses are induced in a subject, the subject may have a more sustained, long-term immune response. Such an immune response can be beneficial, e.g., for preventing HSV (e.g., HSV-1 and/or HSV-2) reactivation with a single administration, which may increase vaccination rates and subject compliance as compared with presently available vaccines that require dosing every few years. In some embodiments, constructs and compositions comprising RNA molecules as described herein (e.g., encoding for one or more HSV (e.g., HSV-1, HSV- 2, or a combination thereof) antigens and/or epitopes) can provide more diverse protection (e.g., protection against HSV (e.g., HSV-1 and/or HSV-2) variants) because, without wishing to be bound to any particular theory, the constructs and compositions can induce multiple immune system responses. [0298] The present disclosure also provides the recognition that, by administering polyribonucleotides and compositions that encode HSV (e.g., HSV-1 and/or HSV-2) antigens and/or epitopes, the polyribonucleotides and compositions described herein avoid administering HSV (e.g., HSV-1 and/or HSV-2) virions, which may infect the subject, go into latency, and reactivate to cause a flare-up. [0299] Still further, the present disclosure provides an insight (and also identifies a source of a problem in some prior HSV vaccination strategies) that, in some embodiments, particularly effective pharmaceutical compositions (e.g., immunogenic compositions, e.g., vaccines) alter one or more characteristics of the innate immune system. The present disclosure provides some such compositions, including, for example, compositions that comprise RNA construct(s) encoding HSV (e.g., HSV-1 and/or HSV-2) protein(s) (e.g., HSV antigens or HSV epitopes) as described herein. [0300] Separately, in some embodiments, the present disclosure provides particular pharmaceutical composition (e.g., immunogenic composition, e.g., vaccine) formats including, for example, RNA pharmaceutical compositions (e.g., immunogenic compositions, e.g., vaccines) comprising particular elements and/or sequences useful for vaccination. [0301] The present disclosure provides a variety of insights and technologies related to such HSV (e.g., HSV-1 and/or HSV-2) antigen constructs and compositions (e.g., RNA vaccine). [0302] As described herein, in many embodiments, provided compositions (e.g. pharmaceutical compositions, e.g., immunogenic compositions, e.g., vaccines) include an RNA active encoding one or more HSV (e.g., HSV-1 and/or HSV-2) polypeptides or antigenic fragments thereof; in some embodiments such RNA active is a modified RNA format in that its uridine residues are substituted with uridine analog(s) such as pseudouridine; alternatively or additionally, in some embodiments, such RNA active includes particular elements (e.g., cap, 5’UTR, 3’UTR, polyA tail, etc) and/or characteristics (e.g., codon optimization) identified, selected, characterized, and/or demonstrated to achieve significant (e.g., elevated) translatability (e.g., in vitro) and/or expression (i.e., in a subject to whom it has been administered) of encoded protein(s). Still further alternatively or additionally, in some embodiments, such RNA active includes particular elements and/or characteristics identified, selected, characterized, and/or demonstrated to achieve significant RNA stability and/or efficient manufacturing, particularly at large scale (e.g., 0.1-10 g, 10-500 g, 500 g-1 kg, 750 g-1.5 kg; those skilled in the art will appreciate that different products may be manufactured at different scales, e.g., depending on patient population size). In some embodiments, such RNA manufacturing scale may be within a range of about 0.01 g/hr RNA to about 1 g/hr RNA, 1 g/hr RNA to about 100 g/hr RNA, about 1 g RNA/hr to about 20 g RNA/hr, or about 100 g RNA/hr to about 10,000 g RNA/hr. In some embodiments, such RNA manufacturing scale may be tens or hundreds of milligrams to tens or hundreds of grams (or more) of RNA per batch. In some embodiments, such RNA manufacturing scale may allow a batch size within a range of about 0.01 g to about 500 g RNA, about 0.01 g to about 10 g RNA, about 1 g to about 10 g RNA, about 10 g to about 500 g RNA, about 10 g to about 300 g RNA, about 10 g to about 200 g RNA or about 30 g to about 60 g RNA. [0303] Still further, in many embodiments, provided compositions (e.g., pharmaceutical compositions, e.g., immunogenic compositions, e.g., vaccines) that include one or more polyribonucleotides are prepared, formulated, and/or utilized in particular LNP compositions, as described herein. [0304] Among other things, the present disclosure provides technologies for rapid development of a pharmaceutical composition (e.g., immunogenic composition, e.g., HSV vaccine) for delivering particular HSV (e.g., HSV-1 and/or HSV-2) antigen constructs to a subject. [0305] Additionally, the present disclosure provides, for example, nucleic acid constructs encoding HSV (e.g., HSV-1 and/or HSV-2) antigens as described herein, expressed HSV (e.g., HSV-1 and/or HSV-2) proteins, and various methods of production and/or use relating thereto, as well as compositions developed therewith and methods relating thereto. [0306] For example, the present disclosure provides technologies for preventing, characterizing, treating, and/or monitoring HSV (e.g., HSV-1 and/or HSV-2) outbreaks and/or infections including, as noted, various nucleic acid constructs and encoded proteins, as well as agents (e.g., antibodies) that bind to such proteins, and compositions that comprise and/or deliver them. [0307] In some embodiments, provided herein are technologies (e.g., compositions and methods) for augmenting, inducing, promoting, enhancing and/or improving an immune response against HSV (e.g., HSV-1 and/or HSV-2) or a component thereof (e.g., a protein or fragment thereof). In some embodiments, technologies provided herein are designed to augment, induce, promote, enhance and/or improve immunological memory against HSV (e.g., HSV-1 and/or HSV-2) or a component thereof (e.g., a protein or fragment thereof). In some embodiments, technologies described herein are designed to act as an immunological boost to a primary composition (e.g., immunogenic composition, e.g., vaccine), such as a composition (e.g., immunogenic composition, e.g., vaccine) directed to an antigen and/or epitopes of HSV (e.g., HSV-1 and/or HSV-2). In some embodiments, compositions of the present disclosure comprise one or more polynucleotide constructs (e.g., one or more string constructs) that encode one or more antigens from HSV (e.g., HSV-1 and/or HSV-2). In some embodiments, the present disclosure provides compositions comprising nucleic acids encoding such HSV (e.g., HSV-1 and/or HSV-2) antigens; those skilled in the art will appreciate from context when reference to a particular polynucleotide (e.g., a DNA or RNA) as “encoding” such antigens in fact is referencing a coding strand or its complement. [0308] The present disclosure provides pharmaceutical compositions (e.g., immunogenic compositions, e.g., vaccines) that deliver particular HSV antigen constructs to a subject (e.g., a patient) and related technologies (e.g., methods). In some embodiments, the present disclosure provides pharmaceutical compositions (e.g., immunogenic compositions, e.g., vaccines) that deliver particular HSV-1 antigen constructs to a subject (e.g., a patient) and related technologies (e.g., methods). In some embodiments, the present disclosure provides pharmaceutical compositions (e.g., immunogenic compositions, e.g., vaccines) that deliver particular HSV-2 antigen constructs to a subject (e.g., a patient) and related technologies (e.g., methods). In some embodiments, the present disclosure provides pharmaceutical compositions (e.g., immunogenic compositions, e.g., vaccines) that deliver particular HSV-1 and HSV-2 antigen constructs to a subject (e.g., a patient) and related technologies (e.g., methods). [0309] The present disclosure further provides the recognition that some HSV antigens are common to both HSV-1 and HSV-2. The present disclosure also provides the recognition that some HSV antigens include sequences conserved between HSV-1 and HSV-2. In addition, the present disclosure recognizes that some HSV-1 antigens have, e.g., at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to comparable HSV-2 antigens. [0310] In some embodiments, the present disclosure provides some HSV antigen constructs particularly useful in effective vaccination. In some embodiments, HSV antigen constructs are HSV-1 antigen construct, HSV-2 antigen constructs, or a combination thereof. [0311] Antigens utilized in accordance with the present disclosure are or include HSV (e.g., HSV-1 and/or HSV-2) components (e.g., antigenic fragments thereof, including epitopes that may comprise non-amino acid, e.g., carbohydrate moieties), which components induce immune responses when administered to humans (or other animals such as rodents and non-human primates susceptible to HSV (e.g., HSV-1 and/or HSV-2) infection). [0312] In many embodiments, antigens utilized in provided pharmaceutical compositions (e.g., immunogenic compositions, e.g., vaccines) include both B-cell and T-cell antigens and/or epitopes, as described herein. In some particular embodiments, delivered antigens include both B-cell and T cell (e.g., CD4 and/or CD8 T cell) antigens and/or epitopes, optionally together in a single antigen polypeptide. In some embodiments, antigens utilized in provided pharmaceutical compositions (e.g., immunogenic compositions, e.g., vaccines) include T cell antigens and/or epitopes. In some embodiments, antigens utilized in provided pharmaceutical compositions (e.g., immunogenic composition, e.g., vaccine), together, include B cell, CD4 T cell and CD8 T cell epitopes. Indeed, in some embodiments, the present disclosure defines particularly useful epitopes for inclusion in HSV (e.g., HSV-1 and/or HSV-2) compositions (e.g., immunogenic compositions, e.g., vaccines), and/or provides antigens that include them. [0313] Exemplary antigens and/or epitopes for use in compositions described herein included those provided in, e.g., Tables 7 and 8 herein and antigenic fragments thereof. In some embodiments, exemplary antigens disclosed in Tables 7 and 8, and/or fragments and/or epitopes thereof, can be useful for compositions described herein. [0314] In some embodiments, a provided pharmaceutical composition (e.g., immunogenic composition, e.g., HSV (e.g., HSV-1 and/or HSV-2) vaccine) comprises or delivers (e.g., causes expression of in a recipient organism, for example by administration of a nucleic acid construct, such as an RNA construct as described herein, that encodes it) an antigen that is or comprises one or more epitopes (e.g., one or more B-cell and/or one or more T-cell antigens and/or epitopes) of an HSV (e.g., HSV-1 and/or HSV-2) protein. In some embodiments, a pharmaceutical composition described herein induces a relevant immune response effective against HSV (e.g., by targeting an HSV-1 protein, an HSV-2 protein, or a combination thereof). [0315] In some embodiments, a provided pharmaceutical composition (e.g., immunogenic composition, e.g., HSV (e.g., HSV-1 and/or HSV-2) vaccine) comprises or delivers an antigen that is or comprises a full-length HSV (e.g., HSV-1 and/or HSV-2) protein. In some embodiments, a provided pharmaceutical composition (e.g., immunogenic composition, e.g., HSV (e.g., HSV-1 and/or HSV-2) vaccine) comprises or delivers an antigen that is or comprises a fragment of an HSV (e.g., HSV-1 and/or HSV-2) protein that is less than a full-length HSV (e.g., HSV-1 and/or HSV-2) protein. In some embodiments, a provided pharmaceutical composition (e.g., immunogenic composition, e.g., HSV (e.g., HSV-1 and/or HSV-2) vaccine) comprises or delivers a chimeric polypeptide that is or comprises part or all of an HSV (e.g., HSV-1 and/or HSV-2) protein and one or more heterologous polypeptide elements. [0316] In some embodiments, an antigen that is included in and/or delivered by a provided pharmaceutical composition (e.g., immunogenic composition, e.g., HSV (e.g., HSV-1 and/or HSV-2) vaccine) is or comprises one or more peptide fragments of an HSV (e.g., HSV-1 and/or HSV-2) antigen; in some such embodiments, each of the one or more peptide fragments includes at least one epitope (e.g., one or more B cell epitopes and/or one or more T cell epitopes), for example as may be predicted, selected, assessed and/or characterized as described herein. [0317] In some embodiments, a T-cell string polypeptide that is included in and/or delivered by a provided pharmaceutical composition (e.g., immunogenic composition, e.g., HSV (e.g., HSV-1 and/or HSV-2) vaccine) is or comprises a plurality of peptide fragments (e.g., HSV T-cell antigens) of one or more HSV (e.g., HSV-1 and/or HSV- 2) antigens. In some embodiments, a single polypeptide antigen may include a plurality of such fragments, e.g., presented as a string of antigens or fragments thereof as described herein (e.g., in that a single polypeptide includes a plurality of amino acid sequences derived from distinct HSV antigens or fragments thereof, optionally separated by or otherwise associated with amino acid linkers or other intervening or terminal amino acid sequences). In some embodiments, a single RNA antigen construct may include a plurality of sequences encoding HSV antigens, e.g., presented as a string of antigen encoding sequences as described herein (e.g., in that a single RNA molecule includes a plurality of nucleic acid sequences encoding distinct HSV antigens or fragments thereof, optionally separated by or otherwise associated with nucleic acid linkers or other intervening or terminal nucleic acid sequences). [0318] In some embodiments, one or more HSV (e.g., HSV-1 and/or HSV-2) antigens or antigenic fragments thereof may be linked with one or more sequences with which it is linked in nature. In some such embodiments, such sequence(s) may be or comprise one or more heterologous elements (e.g., one or more elements, not naturally found in the relevant HSV (e.g., HSV-1 and/or HSV-2) such as a polypeptide or antigenic fragment thereof not naturally found to be directly linked to the relevant HSV (e.g., HSV-1 and/or HSV-2) antigen(s)). For example, in some embodiments, an antigen peptide provided and/or utilized in accordance with the present disclosure may include one or more linker elements and/or one or more membrane association elements and/or one or more secretion elements, etc. In some embodiments, an antigenic polypeptide may comprise a plurality of HSV (e.g., HSV-1 and/or HSV-2) protein fragments or epitopes separated from one another by linkers. [0319] In some embodiments, an HSV (e.g., HSV-1 and/or HSV-2) polypeptide, or fragment or epitope thereof, utilized in a construct as described herein (or encoded by a polyribonucleotide describe herein) may include one or more sequence alterations relative to a particular reference HSV (e.g., HSV-1 and/or HSV-2) polypeptide, or fragment or epitope thereof. For example, in some embodiments, a utilized antigen may include one or more sequence variations found in circulating strains or predicted to arise, e.g., in light of assessments of sequence conservation and/or evolution of HSV (e.g., HSV-1 and/or HSV-2) polypeptides over time and/or across strains. Alternatively or additionally, in some embodiments, a utilized antigen may include one or more sequence variations selected, for example, to impact stability, folding, processing and/or display of the antigen or any epitope thereof. [0320] In some embodiments, an HSV (e.g., HSV-1 and/or HSV-2) polypeptide, or fragment or epitope thereof, utilized in an antigen as described herein shows at least 70%, 71%, 72%, 73%, 74%, 75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% amino acid sequence identity with a relevant corresponding reference (e.g., wild type) polypeptide, fragment or epitope. In some embodiments, an HSV (e.g., HSV-1 and/or HSV-2) polypeptide, or fragment or epitope thereof, utilized in an antigen as described herein shows at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence homology (i.e., identity or conservative substitution as is understood in the art) amino acid sequence identity with a relevant corresponding reference (e.g., wild type) protein, fragment or epitope. Moreover, in some embodiments, an HSV (e.g., HSV-1 and/or HSV-2) polypeptide, or fragment or epitope thereof, utilized in an antigen as described herein shares conserved amino acid residues (e.g., at corresponding positions) with a relevant corresponding reference (e.g., wild type) polypeptide, fragment or epitope. Those skilled in the art will appreciate that, in general, lower percent identity or homology may be tolerated for shorter peptides, as a single change will by definition have a larger impact on percent identity or homology when considered relative to a smaller number of residues. For example, those skilled in the art will appreciate that, for sequences longer than about 20 amino acids, percent identity or homology is typically greater than about 80%; for sequences longer than about 50 amino acids, percent identity or homology is typically greater than about 90%. [0321] In some embodiments, assessments of degree of conservation may consider the physiochemical difference between two amino acids as described, for example, in WO2014/180569, which is incorporated herein by reference in its entirety. It is well known in molecular evolution that amino acids that interchange frequently are likely to have chemical and physical similarities whereas amino acids that interchange rarely are likely to have different physico-chemical properties. The likelihood for a given substitution to occur in nature compared with the likelihood for this substitution to occur by chance can measured by log-odds matrices. The patterns observed in log- odds matrices imposed by natural selection "reflect the similarity of the functions of the amino acid residues in their weak interactions with one another in the three dimensional conformation of proteins" (see Dayhoff et al. Atlas of protein sequence and structure 5:345, 1978180569, which is incorporated herein by reference in its entirety). In some embodiments, evolutionary based log-odds matrices, which may be referred to as "T scores" can be used to
Figure imgf000130_0001
re ect extent to w c a sequence variation might impact T cell recognition. Substitutions with positive T scores (i.e., log-odds) are likely to occur in nature, and hence correspond to two amino acids that have similar physico-chemical properties. Substitutions with positive T scores would have a lower likelihood of altering immunogenicity. Conversely, substitutions with negative T scores reflect substitutions that are unlikely to occur in nature and hence correspond to two amino acids that have significantly different physico-chemical properties. Such substitutions would have a greater chance of altering immunogenicity. In some embodiments, presence of negative T score substitutions within a sequence, even if it is otherwise highly conserved, may indicate that it would be relatively less useful in a composition antigen as described herein. [0322] In various embodiments, an HSV antigen construct includes and/or encodes a plurality of HSV antigens (e.g., a plurality of HSV antigens that are or include one or more T cell antigens for HSV) provided in Table 7, or fragments thereof. In some embodiments, an HSV antigen construct can include and/or encode at least one HSV antigen provided in Table 7, or fragments thereof. In some embodiments, an HSV antigen construct can include and/or encode at least one T cell antigen for HSV provided in Table 7, or fragments thereof. [0323] In some embodiments, an HSV antigen construct can include and/or encode a plurality of (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19) HSV antigens provided in Table 7, or fragments thereof. [0324] In some embodiments, an HSV antigen construct can include and/or encode a plurality of (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19) T cell antigens for HSV selected from HSV antigens provided in Table 7, or fragments thereof. [0325] In some embodiments, an antigen utilized in accordance with the present disclosure includes HSV (e.g., HSV-1 and/or HSV-2) protein sequences identified and/or characterized by one or more of: 1) HLA-I or HLA-II binding (e.g., to HLA allele(s) present in a relevant population) 2) HLA ligandomics data, optionally confirmed by mass spectrometry 3) Relatively high expression 4) Sequence conservation 5) Surface exposure 6) Serum reactivity 7) Immunogenicity (e.g., presence of one or more B-cell and/or T-cell antigens and/or epitopes; evidence of ability to induce sterile protection in model systems including, e.g., humans, non-human primates, and/or mice). 8) Absence of sequences that overlap with human proteome [0326] In some embodiments, such characteristics are experimentally or computationally assessed. In some embodiments, such characteristics are assessed by consultation with published reports. [0327] For example, in some embodiments, HLA-I and/or HLA-II binding is experimentally assessed; in some embodiments it is predicted. [0328] In some embodiments, predicted HLA-I or HLA-II binding is assessed using an algorithm such as neonmhc 1 and/or neonmhc2, which predict and/or characterize likelihood of MHC class I and MHC class II binding, respectively. Alternatively or additionally, in some embodiments, an MHC-peptide presentation prediction algorithm or MHC-peptide presentation predictor is or comprises NetMHCpan or NetMHCIIpan. In some embodiments, a hidden Markov model approach may be utilized for MHC-peptide presentation prediction and/or characterization. In some embodiments, the peptide prediction model MARIA may be utilized. In some embodiments, NetMHCpan is not utilized to predict or characterize likelihood of MHC binding for peptides as described herein. In some embodiments, the peptide prediction model MARIA may be utilized. In some embodiments, NetMHCIIpan is not utilized to predict or characterize likelihood of MHC binding for peptides as described herein. In some embodiments, neither NetMHCpan nor NetMHCIIpan is utilized to predict or characterize likelihood of MHC binding for peptides as described herein. In some embodiments, an MHC-peptide presentation prediction algorithm or MHC-peptide presentation predictor is or comprises RECON® (Real-time Epitope Computation for ONcology), which offers high quality MHC-peptide presentation prediction based on expression, processing and binding capabilities. See, for example, Abelin et al., Immunity 21:315, 2017; Abelin et al., Immunity 15:766, 2019. [0329] In some embodiments, HLA binding and/or ligandomics assessments will consider the geographic region of subjects to be immunized. For example, in some embodiments, HLA allelic diversity will be considered. In some embodiments, antigen(s) included in a provided pharmaceutical composition (e.g., immunogenic composition, e.g., vaccine) will be or comprise peptides (e.g., epitopes) expected or determined, when considered together, to bind to a significant percentage (e.g., at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or more) of HLA alleles expected or known to be present in a relevant region or population. In some embodiments, antigen(s) included in a provided pharmaceutical composition (e.g., immunogenic composition, e.g., vaccine) will be or comprise peptides expected or determined, when considered together, to bind to the most prevalent (e.g., the 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 most prevalent, or at least 1, 2, 3, 4, or 5 of the 10 most prevalent, etc.) HLA alleles expected or known to be present in a relevant region or population). [0330] In some embodiments, expression level is experimentally determined (e.g., in a model system or in infected humans). In some embodiments, expression level is a reported level (e.g., in a published or presented report). In some embodiments, expression level is assessed as RNA (e.g., via RNASeq). In some embodiments (and typically preferably), expression levels is assessed as protein. [0331] In some embodiments, sequence conservation is assessed, for example, using publicly available sequence evaluation software (such as, for example, multiple sequence alignment programs MAFFT, Clustal Omega, etc.). In some embodiments, sequence conservation is determined by consultation with published resources (e.g., sequences). In some embodiments, sequence conservation includes consideration of currently or recently detected strains (e.g., in an active outbreak). [0332] In some embodiments, surface exposure is assessed by reference to publicly available database and/or software. [0333] In some embodiments, serum reactivity is assessed by contacting serum samples from infected individuals with polypeptides including sequences of interest (e.g., as may be displayed via, for example, phage display or peptide array, etc.; see, for example, Whittemore et al “ A General Method to Discover Epitopes from Sera” PlosOne, 2016; https://doi.org/10.1371/journal.pone.0157462). In some embodiments, serum reactivity is assessed by consultation with literature reports and or database data indicating serum-recognized sequences. [0334] In some embodiments, assessment of immunoreactivity and/or of presence of an epitope may be or comprise consultation with the Immune Epitope Database (IEDB) which those skilled in the art will be aware is a freely available resource funded by NIAID that catalogs experimental data on antibody and T cell epitopes (see iedb.org). [0335] In some embodiments, antigen(s) utilized in accordance with the present disclosure are characterized by dendritic cell presentation which, in turn may be indicative of HLA binding and/or of immunogenicity. [0336] In some embodiments, antigen(s) utilized in accordance with the present disclosure are or comprises sequences (e.g., epitopes, fragments, complete proteins) of HSV proteins found in the HSV envelope. In some embodiments, antigen(s) utilized in accordance with the present disclosure are or comprises sequences (e.g., epitopes, fragments, complete proteins) of HSV proteins found in the HSV tegument. [0337] Among other things, the present disclosure provides an insight that, in some embodiments, it may be desirable to include two or more different epitopes, optionally from two or more different HSV (e.g., HSV-1 and/or HSV-2) proteins, in pharmaceutical composition (e.g., immunogenic composition, e.g., vaccine) compositions, which can be useful in the treatment of HSV. Table 8: Exemplary antigen fragment Antigen Sequence (Amino Acid) SEQ ID Fragment NO: RL2.1 CTDEIAPPLRCQSFPCLHPFCIPCMKTWIPLRNTCPLCNTPVAYLIVGVTASGSFSTIPIVN 562 Fragment DPRTRVEAEAAVRAGTAVDFIWTGNPRTAPRSLS RL2.2 LPIAGVSSVVALAPYVNKTVTGDCLPVLDMETGHIGAYVVLVDQTGNVADLLRAAAPAWS 563 Fragment RRTLLPEHARNCVRPPDYPTPPASEWNSLWMTPVGNMLFDQGTLVG RS1 RAAAWMRQVPDPEDVRVVILYSPLPGEDLAAGRAGGGPPPEWSAERGGLSCLLAALGNR 564 Fragment LCGPATAAWAGNWTGAPDVSALGAQGVLLLSTRDLAFAGAVEFLGLLAGACDRRLIVVN AVRAADWPADGPVVSRQHAYLACEVLPAVQCAVRWPAARDLRRTVLASGRVFGPGVFA RVEAAHARLYPDAPPLRLCRGANVRYRVRTRFGPDTLVPMSPREYRRAVLPALDGRAAAS UL54 ETLVAHGPSLYRTFAANPRAASTAKAMRDCVLRQENLIEALASADETLAWCKMCIHHNLP 565 fragment LRPQDPIIGTAAAVLENLATRLRPFLQCYLKARGLCGLDDLCSRRRLSDIKDIASFVLVILA RLANRVERGVSEIDYTTVGVGAGETMHFYIPGACMAGLIEILDTHRQECSSRVCELTASH TIAPLYVHGKYFYCNSLF UL29 REDIETIAFIKRFSLDYGAINFINLAPNNVSELAMYYMANQILRYCDHSTYFINTLTAVIAG 566 Fragment SRRPPSVQAAAAWAPQGGAGLEAGARALMDSLDAHPGAWTSMFASCNLLRPVMAARP MVVLGLSISKYYGMAGNDRVFQAGNWASLLGGKNACPLLIFDRTRKFVL UL39 RTFGSAPRLTEDDFGLLNYALAEMRRLCLDLPPVPPNAYTPYHLREYATRLVNGFKPLVRR 567 Fragment SARLYRILGVLVHLRIRTREASFEEWMRSKEVDLDFGLTERLREHEAQLMILAQALNPYDC LIHSTPNTLVERGLQSALKYEEFYLKRFGGHYMESVFQMYTRIAGFLA UL49 KMTRGAPKASATPATDPARGRRPAQADSAVLLDAPAPTASGRTKTPAQGLAKKLHFSTA 568 Fragment PPSPTAPWTPRVAGFNKRVFCAAVG UL9 LLNNYDVLVLDEVMSTLGQLYSPTMQQLGRVDALMLRLLRTCPRIIAMDATANAQLVDFL 569 Fragment CSLRGEKNVHVVIGEYAMPGFSARRCLFLPRLGPEVLQAALRRRGPAGGAPPPDAPPDAT FFGELEARLAGGDNVCIFSSTVSFAEVVARFCRQFTDRVLLLHSLTPPGDVTTWGRYRVVI YTTVVTVGLSFDPPHFDSMFAYVKPMNYGPDMVSVYQSLGRVRTLRKGELLIYMDGSGA RSEPV UL30.1 ISCLLYDLSTTALEHILLFSLGSCDLPESHLSDLASRGLPAPVVLEFDSEFEMLLAFMTFVK 570 Fragment QYGPEFVTGYNIINFDWPFVLTKLTEIYKVPLDGYGRMNGRGVFRVWDIGQSHF UL30.2 GLLPCLHVAATVTTIGREMLLATRAYVHARWAEFDQLLADFPEAAGMRAPGPYSM 571 Fragment UL40 TSQCPDINHLRSLSILNRWLETELVFVGDEEDVSKLSEGELGFYRFLFAFLSAADDLVTEN 572 Fragment LGGLSGLFEQKDILHYYVEQECIEVVHSRVYNIIQLVLFHNNDQARRAYVARTINHPAIRV KVDWLEARVRECDSIPEKFILMILIEGVFFAASFAAIAYLRTNNLLR UL5.1 HEFGNLMKVLEYGLPITEEHMQFVDRFVVPESYITNPANLPGWTRLFSSHKEVSAYMAKL 573 Fragment HAYLKVTREGEFVVFTLPVLTFVSVKEFDEYRRL UL5.2 ELFGEVFESAPFSTYVDNVIFRGCELLTGSPRGGLMSVALQTDNYTLMGYTYTRVFAFAEE 574 Fragment LRRRHATAGVAEFLEESPLPYIVLRDQHGFMSVVNTNI UL52 SVAAPVEVTALYATDGCVITSSLALLTNCLLGAEPLYIFSYDAYRSDAPNGPTGAPTEQER 575 fragment FEGSRALYRDAGGLNGDSFRVTFCLLGTEVGVTHHPKGRTRPMFVCRFERADDVAVLQD ALGRGTPLLPAHVTATLDLEATFALHANIIMALTVAIVHNAPARIGSGSTAPLYEPGESMR SVV UL1 RTPADDVSWRYEAPSVIDYARIDGIFLRYHCPGLDTFLWDRHAQRAYLVNPFLFAAGFLE 576 Fragment DLSHSVFPADTQETT UL19 DGRLLHNTQARAADAADDRPHRPADWTVHHKIYYYVLVPAFSRGRCCTAGVRFDRVYA 577 Fragment TLQNMVVPEIAPGEECPSDPVTDPAHPLHPANLVANTVKRMFHN UL21 SPTQKLAVYYYLIHRERRMSPFPALVRLVGRYIQRHGLYVPAPDEPTLADAMNGL 578 Fragment UL27.1 NYTEGIAVVFKENIAPYKFKATMYYKDVTVSQVWFGHRYSQFMGIFEDRAPVPFEEV 579 Fragment UL27.2 SVYPYDEFVLATGDFVYMSPFYGYREGSH 580 Fragment UL46 GLASDPHYDYIRHYASAAKQALGEVELSGGQLSRAILAQYWKYLQTVVPSGLDIPDDPAG 581 Fragment DCDPSLHVLLRPTLLPKLLVRAPFKSGAAAAKYAAAVAGLRDAAHRLQQYMFFMRPADPS RPSTDTALRLSELLAYVSVLYHWASWMLWTADKYV UL47 GPDAAVFRSSLGSLLYWPGVRALLGRDCRVAARYAGRMTYIATGALLARFNPGAVKCVLP 582 Fragment REAAFAGRVL UL48 ALFNRLLDDLGFSAGPALCTMLDTWNEDLFSGFPTNADMYRECKFLSTLPSDVIDWGDA 583 Fragment HVPERSPIDIRAHGDVAFPTLPATRDELPSYYEAMAQFFRGELRA UL25 FLWEDQTLLRATANTITALAVLRRLLANGNVYADRLDNRLQLGMLIPGAVPAEAIARGAS 584 Fragment GLDSGAIKSGDNNLEALCVNYVLPLYQADPTVELTQLFPGLAALCL US1.1 DDASDGWLVDTPPRKSKRPRINLRLTSSPDRRAGVVFPEV 585 Fragment US1.2 PASLPGIAHAHRRSARQAQMRSGAAWTLDLHYIRQCVNQL 586 Fragment US8.1 ILSPTAPSVYPHSEGRKSRRPLTTFGSGSPGRRHSQASYPSVLW 587 Fragment US8.2 GLAWLASTVNLEFQHASPQHAGLYLCVVYVDDHIHAWGHMTISTAAQYRNAVVEQHLP 588 Fragment QRQPEPVEPTRPHVRAPHPAPSARGPLRL US8.3 KLLWAAEPLDACGPLRPSWVALWPPRRVLETVVDAACMRAPEPLAIAYSPPFPAGDEGLY 589 Fragment SELAWRDRVAVVNESLVIYGALETDSGLYTLSVVGLSDEARQVASVVLVVEPAP US12 EDREAARTAVTDPELPLLCPPDV 590 Fragment UL50 ANGATVIQPSLRVLRAADGPEACYVLGRSSLNARGLLVMPTRWPSGHACAFVVCNLTGV 591 Fragment PVTL UL26 APLPDRAVPIYVAGFLALYDSGDPGELALDPDTVRAALPPENPLPINVDHRARCEVGRVLA 592 Fragment VVNDPRGPFFVGLIACVQLERVLETAASAAIFERRGPALSREERLLYLITNYLPSVSLSTKR RGDEVPPDRTLFAHVALCAIGRRLGTIVTYDTSLDAA UL26 HYPPPPAHPYPGMLFAGPSPLEAQIAALVGAIAADRQAGGLPAAAGDHGIRGSAKRRRHE 593 Fragment VEQPEYDCG US10 SSPRQRTYVLPRVGIHNAPASDTRAPKRANSRHRADRPPESPGSELYPLNAQALAHLQML 594 Fragment PADHRAFFRTVIEVSRLCALNTHDPPPPLAGARVGQEAQLVHTQWLRANRESSPLWPWR TAAMNFIAAAAPCVQTHRHMHDLLMACAFWC [0338] In some embodiments, an HSV antigen for use in accordance with the present disclosure comprises an RL2 polypeptide or antigenic fragment thereof, an RS1 polypeptide or antigenic fragment thereof, a UL54 polypeptide or antigenic fragment thereof, a UL29 polypeptide or antigenic fragment thereof, a UL39 polypeptide or antigenic fragment thereof, a UL49 polypeptide or antigenic fragment thereof, a UL9 polypeptide or antigenic fragment thereof, a UL30 polypeptide or antigenic fragment thereof, a UL40 polypeptide or antigenic fragment thereof, a UL5 polypeptide or antigenic fragment thereof, a UL52 polypeptide or antigenic fragment thereof, a UL1 polypeptide or antigenic fragment thereof, a UL19 polypeptide or antigenic fragment thereof, a UL21 polypeptide or antigenic fragment thereof, a UL27 polypeptide or antigenic fragment thereof, a UL46 polypeptide or antigenic fragment thereof, a UL47 polypeptide or antigenic fragment thereof, a UL48 polypeptide or antigenic fragment thereof, a UL25 polypeptide or antigenic fragment thereof, US7 polypeptide or antigenic fragment thereof, US8 polypeptide or antigenic fragment thereof, and UL22 polypeptide or antigenic fragment thereof, a US10 polypeptide or antigenic fragment thereof, a US12 polypeptide or antigenic fragment thereof, a UL26 polypeptide or antigenic fragment thereof, a UL50 polypeptide or antigenic fragment thereof, or a combination thereof. [0339] In some embodiments, a polyribonucleotide provided herein encodes one or more of an RL2 polypeptide or antigenic fragment thereof, an RS1 polypeptide or antigenic fragment thereof, a UL54 polypeptide or antigenic fragment thereof, a UL29 polypeptide or antigenic fragment thereof, a UL39 polypeptide or antigenic fragment thereof, a UL49 polypeptide or antigenic fragment thereof, a UL9 polypeptide or antigenic fragment thereof, a UL30 polypeptide or antigenic fragment thereof, a UL40 polypeptide or antigenic fragment thereof, a UL5 polypeptide or antigenic fragment thereof, a UL52 polypeptide or antigenic fragment thereof, a UL1 polypeptide or antigenic fragment thereof, a UL19 polypeptide or antigenic fragment thereof, a UL21 polypeptide or antigenic fragment thereof, a UL27 polypeptide or antigenic fragment thereof, a UL46 polypeptide or antigenic fragment thereof, a UL47 polypeptide or antigenic fragment thereof, a UL48 polypeptide or antigenic fragment thereof, a UL25 polypeptide or antigenic fragment thereof, a US1 polypeptide or antigenic fragment thereof, a US7 polypeptide or antigenic fragment thereof, US8 polypeptide or antigenic fragment thereof, and a UL22 polypeptide or antigenic fragment thereof, a US10 polypeptide or antigenic fragment thereof, a US12 polypeptide or antigenic fragment thereof, a UL26 polypeptide or antigenic fragment thereof, a UL50 polypeptide or antigenic fragment thereof, or a combination thereof. [0340] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide comprising one or more HSV RL2 polypeptides or antigenic fragments thereof, one or more HSV RS1 polypeptides or antigenic fragments thereof, and one or more HSV UL54 polypeptides or antigenic fragments thereof. [0341] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an RL2 polypeptide or antigenic fragment thereof, a linker, an RL2 polypeptide or antigenic fragment thereof, a linker, an RS1 polypeptide or antigenic fragment thereof, a linker, a UL54 polypeptide or antigenic fragment thereof, and a linker. [0342] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an UL54 polypeptide or antigenic fragment thereof, a linker, an RS1 polypeptide or antigenic fragment thereof, a linker, an RL2 polypeptide or antigenic fragment thereof, a linker, a RL2 polypeptide or antigenic fragment thereof, and a linker. [0343] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide comprising one or more HSV UL29 polypeptides or antigenic fragments thereof, one or more HSV UL39 polypeptides or antigenic fragments thereof, one or more HSV UL49 polypeptides or antigenic fragments thereof, and one or more HSV UL9 polypeptides or antigenic fragments thereof. [0344] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an UL29 polypeptide or antigenic fragment thereof, a linker, an UL39 polypeptide or antigenic fragment thereof, a linker, an UL49 polypeptide or antigenic fragment thereof, a linker, a UL9 polypeptide or antigenic fragment thereof, and a linker. [0345] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an UL9 polypeptide or antigenic fragment thereof, a linker, an UL49 polypeptide or antigenic fragment thereof, a linker, an UL39 polypeptide or antigenic fragment thereof, a linker, a UL29 polypeptide or antigenic fragment thereof, and a linker. [0346] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide comprising one or more HSV UL30 polypeptides or antigenic fragments thereof, one or more HSV UL40 polypeptides or antigenic fragments thereof, one or more HSV UL5 polypeptides or antigenic fragments thereof, and one or more HSV UL52 polypeptides or antigenic fragments thereof. [0347] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an UL30 polypeptide or antigenic fragment thereof, a linker, an UL30 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, a UL5 polypeptide or antigenic fragment thereof, a linker, a UL5 polypeptide or antigenic fragment thereof, a linker, a UL52 polypeptide or antigenic fragment thereof, and a linker. [0348] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an UL52 polypeptide or antigenic fragment thereof, a linker, an UL5 polypeptide or antigenic fragment thereof, a linker, an UL5 polypeptide or antigenic fragment thereof, a linker, a UL40 polypeptide or antigenic fragment thereof, a linker, a UL30 polypeptide or antigenic fragment thereof, a linker, a UL30 polypeptide or antigenic fragment thereof, and a linker. [0349] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide comprising one or more HSV UL1 polypeptides or antigenic fragments thereof, one or more HSV UL19 polypeptides or antigenic fragments thereof, one or more HSV UL21 polypeptides or antigenic fragments thereof, one or more HSV UL27 polypeptides or antigenic fragments thereof, one or more HSV UL46 polypeptides or antigenic fragments thereof, one or more HSV UL47 polypeptides or antigenic fragments thereof, one or more UL48 polypeptides or antigenic fragments thereof, and one or more HSV UL25 polypeptides or antigenic fragments thereof. [0350] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an HSV UL1 polypeptide or antigenic fragment thereof, a linker, an HSV UL19 polypeptide or antigenic fragment thereof, a linker, an HSV UL21 polypeptide or antigenic fragment thereof, a linker, an HSV UL27 polypeptide or antigenic fragment thereof, a linker, an HSV UL27 polypeptide or antigenic fragment thereof, a linker, an HSV UL46 polypeptide or antigenic fragment thereof, a linker, an HSV UL47 polypeptide or antigenic fragment thereof, a linker, an HSV UL25 polypeptide or antigenic fragment thereof, a linker, an HSV UL48 polypeptide or antigenic fragment thereof, and a linker. [0351] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, nucleotide sequences that encode an HSV-1 gD secretory signal, an HSV UL48 polypeptide or antigenic fragment thereof, a linker, an HSV UL25 polypeptide or antigenic fragment thereof, a linker, an HSV UL47 polypeptide or antigenic fragment thereof, a linker, an HSV UL46 polypeptide or antigenic fragment thereof, a linker, an HSV UL27 polypeptide or antigenic fragment thereof, a linker, an HSV UL27 polypeptide or antigenic fragment thereof, a linker, an HSV UL21 polypeptide or antigenic fragment thereof, a linker, an HSV UL19 polypeptide or antigenic fragment thereof, a linker, an HSV UL1 polypeptide or antigenic fragment thereof, and a linker, and a MITD. [0352] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide comprising, one or more HSV US1 polypeptides or antigenic fragments thereof, one or more HSV US8 polypeptides or antigenic fragments thereof, one or more HSV US12 polypeptides or antigenic fragments thereof, one or more HSV UL50 polypeptides or antigenic fragments thereof, one or more HSV UL26 polypeptides or antigenic fragments thereof, and one or more HSV US10 polypeptides or antigenic fragments thereof. [0353] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an US1 polypeptide or antigenic fragment thereof, a linker, an US1 polypeptide or antigenic fragment thereof, a linker, an US8 polypeptide or antigenic fragment thereof, a linker, a US8 polypeptide or antigenic fragment thereof, a linker, a US8 polypeptide or antigenic fragment thereof, a linker, a US12 polypeptide or antigenic fragment thereof, a linker, a UL50 polypeptide or antigenic fragment thereof, a linker, a UL26 polypeptide or antigenic fragment thereof, a linker, a UL26 polypeptide or antigenic fragment thereof, a linker, a US10 polypeptide or antigenic fragment thereof, and a linker. [0354] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an UL26 polypeptide or antigenic fragment thereof, a linker, an UL26 polypeptide or antigenic fragment thereof, a linker, an US10 polypeptide or antigenic fragment thereof, a linker, a UL50 polypeptide or antigenic fragment thereof, a linker, a US12 polypeptide or antigenic fragment thereof, a linker, a US8 polypeptide or antigenic fragment thereof, a linker, a US8 polypeptide or antigenic fragment thereof, a linker, a US8 polypeptide or antigenic fragment thereof, a linker, a US1 polypeptide or antigenic fragment thereof, a linker, a US1 polypeptide or antigenic fragment thereof, and a linker. [0355] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide comprising one or more HSV RL2 polypeptides or antigenic fragments thereof, one or more HSV UL54 polypeptides or antigenic fragments thereof, one or more HSV UL47 polypeptides or antigenic fragments thereof, one or more HSV UL46 polypeptides or antigenic fragments thereof, and one or more HSV UL21 polypeptides or antigenic fragments thereof. [0356] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an RL2 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, a UL46 polypeptide or antigenic fragment thereof, a linker, a UL21 polypeptide or antigenic fragment thereof, and a linker. [0357] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an UL21 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, an RL2 polypeptide or antigenic fragment thereof, and a linker. [0358] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide comprising one or more HSV UL29 polypeptides or antigenic fragments thereof, one or more HSV UL39 polypeptides or antigenic fragments thereof, one or more HSV UL9 polypeptides or antigenic fragments thereof, one or more HSV UL5 polypeptides or antigenic fragments thereof, one or more HSV UL40 polypeptides or antigenic fragments thereof, and one or more HSV UL30 polypeptides or antigenic fragments thereof. [0359] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an UL29 polypeptide or antigenic fragment thereof, a linker, an UL39 polypeptide or antigenic fragment thereof, a linker, an UL9 polypeptide or antigenic fragment thereof, a linker, an UL5.1 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL30.1 polypeptide or antigenic fragment thereof, and a linker. [0360] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an UL30.1 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL5.1 polypeptide or antigenic fragment thereof, a linker, an UL9 polypeptide or antigenic fragment thereof, a linker, an UL39 polypeptide or antigenic fragment thereof, a linker, an UL29 polypeptide or antigenic fragment thereof, and a linker. [0361] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide comprising one or more HSV RL2 polypeptides or antigenic fragments thereof, one or more HSV UL54 polypeptides or antigenic fragments thereof, one or more HSV UL9 polypeptides or antigenic fragments thereof, one or more HSV UL39 polypeptides or antigenic fragments thereof, and one or more HSV UL5 polypeptides or antigenic fragments thereof. [0362] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an RL2.1 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, an UL9 polypeptide or antigenic fragment thereof, a linker, a UL39 polypeptide or antigenic fragment thereof, a linker, a UL5.1 polypeptide or antigenic fragment thereof, and a linker. [0363] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an UL5.1 polypeptide or antigenic fragment thereof, a linker, an UL39 polypeptide or antigenic fragment thereof, a linker, an UL9 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, an RL2.1 polypeptide or antigenic fragment thereof, and a linker. [0364] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide comprising one or more HSV UL47 polypeptides or antigenic fragments thereof, one or more HSV UL46 polypeptides or antigenic fragments thereof, one or more HSV UL21 polypeptides or antigenic fragments thereof, one or more HSV UL5.2 polypeptides or antigenic fragments thereof, one or more HSV UL40 polypeptides or antigenic fragments thereof, one or more HSV UL30.1 polypeptides or antigenic fragments thereof, and one or more HSV UL29 polypeptides or antigenic fragments thereof. [0365] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL21 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL30.1 polypeptide or antigenic fragment thereof, a linker, an UL29 polypeptide or antigenic fragment thereof, and a linker. [0366] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an UL29 polypeptide or antigenic fragment thereof, a linker, an UL30.1 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL21 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, and a linker. [0367] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide comprising one or more HSV RL2.1 polypeptides or antigenic fragments thereof, one or more HSV UL54 polypeptides or antigenic fragments thereof, one or more HSV UL5.2 polypeptides or antigenic fragments thereof, one or more HSV UL40 polypeptides or antigenic fragments thereof, one or more HSV UL47 polypeptides or antigenic fragments thereof, and one or more HSV UL46 polypeptides or antigenic fragments thereof. [0368] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, a RL2.1 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, and a linker. [0369] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, a RL2.1 polypeptide or antigenic fragment thereof, and a linker. [0370] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide comprising one or more HSV RL2.1 polypeptides or antigenic fragments thereof, one or more HSV RS1 polypeptides or antigenic fragments thereof, and one or more HSV UL54 polypeptides or antigenic fragments thereof. [0371] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, a RL2.1 polypeptide or antigenic fragment thereof, a linker, a RS1 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, and a linker. [0372] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide comprising one or more HSV UL29 polypeptides or antigenic fragments thereof, one or more HSV UL39 polypeptides or antigenic fragments thereof, and one or more HSV UL9 polypeptides or antigenic fragments thereof. [0373] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an UL29 polypeptide or antigenic fragment thereof, a linker, an UL39 polypeptide or antigenic fragment thereof, a linker, an UL9 polypeptide or antigenic fragment thereof, and a linker. [0374] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide comprising one or more HSV UL5.1 polypeptides or antigenic fragments thereof, one or more HSV UL5.2 polypeptides or antigenic fragments thereof, one or more HSV UL40 polypeptides or antigenic fragments thereof, and one or more HSV UL30.1 polypeptides or antigenic fragments thereof. [0375] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an UL5.1 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL30.1 polypeptide or antigenic fragment thereof, and a linker. [0376] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide comprising one or more HSV UL47 polypeptides or antigenic fragments thereof, one or more HSV UL46 polypeptides or antigenic fragments thereof, and one or more HSV UL21 polypeptides or antigenic fragments thereof. [0377] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL21 polypeptide or antigenic fragment thereof, and a linker. [0378] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide comprising one or more HSV UL29 polypeptides or antigenic fragments thereof, one or more HSV UL39 polypeptides or antigenic fragments thereof, one or more HSV UL9 polypeptides or antigenic fragments thereof, one or more HSV UL5.1 polypeptides or antigenic fragments thereof, one or more HSV UL5.2 polypeptides or antigenic fragments thereof, one or more HSV UL40 polypeptides or antigenic fragments thereof, and one or more HSV UL30.1 polypeptides or antigenic fragments thereof. [0379] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an UL29 polypeptide or antigenic fragment thereof, a linker, an UL39 polypeptide or antigenic fragment thereof, a linker, an UL9 polypeptide or antigenic fragment thereof, a linker, an UL5.1 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL30.1 polypeptide or antigenic fragment thereof, and a linker. [0380] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an UL30.1 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL5.1 polypeptide or antigenic fragment thereof, a linker, an UL9 polypeptide or antigenic fragment thereof, a linker, an UL39 polypeptide or antigenic fragment thereof, a linker, an UL29 polypeptide or antigenic fragment thereof, and a linker. [0381] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide comprising one or more HSV RL2.1 polypeptides or antigenic fragments thereof, one or more HSV RS1 polypeptides or antigenic fragments thereof, one or more HSV UL54 polypeptides or antigenic fragments thereof, one or more HSV UL47 polypeptides or antigenic fragments thereof, one or more HSV UL46 polypeptides or antigenic fragments thereof, and one or more HSV UL21 polypeptides or antigenic fragments thereof. [0382] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, a RL2.1 polypeptide or antigenic fragment thereof, a linker, a RS1 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL21 polypeptide or antigenic fragment thereof, and a linker. [0383] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an UL21 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, a RS1 polypeptide or antigenic fragment thereof, a linker, a RL2.1 polypeptide or antigenic fragment thereof, and a linker. [0384] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide comprising one or more HSV UL5.1 polypeptides or antigenic fragments thereof, one or more HSV UL5.2 polypeptides or antigenic fragments thereof, one or more HSV UL40 polypeptides or antigenic fragments thereof, one or more HSV UL30.1 polypeptides or antigenic fragments thereof, one or more HSV UL47 polypeptides or antigenic fragments thereof, one or more HSV UL46 polypeptides or antigenic fragments thereof, and one or more HSV UL21 polypeptides or antigenic fragments thereof. [0385] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an UL5.1 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL30.1 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL21 polypeptide or antigenic fragment thereof, and a linker. [0386] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide comprising, in N-terminus to C-terminus order, an UL21 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL30.1 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL5.1 polypeptide or antigenic fragment thereof, and a linker. [0387] In some embodiments, a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is an RL2 polypeptide or antigenic fragment thereof. In some embodiments, an RL2 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of CTDEIAPPLRCQSFPCLHPFCIPCMKTWIPLRNTCPLCNTPVAYLIVGVTASGSFSTIPIVNDPRTRVEAEAAVRAGTAVDFIWTGNPR TAPRSLS (SEQ ID NO: 562). In some embodiments, an RL2 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of LPIAGVSSVVALAPYVNKTVTGDCLPVLDMETGHIGAYVVLVDQTGNVADLLRAAAPAWSRRTLLPEHARNCVRPPDYPTPPASEWN SLWMTPVGNMLFDQGTLVG (SEQ ID NO: 563). [0388] In some embodiments, a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is an RS1 polypeptide or antigenic fragment thereof. In some embodiments, an RS1 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of RAAAWMRQVPDPEDVRVVILYSPLPGEDLAAGRAGGGPPPEWSAERGGLSCLLAALGNRLCGPATAAWAGNWTGAPDVSALGAQ GVLLLSTRDLAFAGAVEFLGLLAGACDRRLIVVNAVRAADWPADGPVVSRQHAYLACEVLPAVQCAVRWPAARDLRRTVLASGRVF GPGVFARVEAAHARLYPDAPPLRLCRGANVRYRVRTRFGPDTLVPMSPREYRRAVLPALDGRAAAS (SEQ ID NO: 564). [0389] In some embodiments, a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is a UL54 polypeptide or antigenic fragment thereof. In some embodiments, a UL54 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of ETLVAHGPSLYRTFAANPRAASTAKAMRDCVLRQENLIEALASADETLAWCKMCIHHNLPLRPQDPIIGTAAAVLENLATRLRPFLQC YLKARGLCGLDDLCSRRRLSDIKDIASFVLVILARLANRVERGVSEIDYTTVGVGAGETMHFYIPGACMAGLIEILDTHRQECSSRVCE LTASHTIAPLYVHGKYFYCNSLF (SEQ ID NO: 565). [0390] In some embodiments, a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is a UL29 polypeptide or antigenic fragment thereof. In some embodiments, a UL29 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of REDIETIAFIKRFSLDYGAINFINLAPNNVSELAMYYMANQILRYCDHSTYFINTLTAVIAGSRRPPSVQAAAAWAPQGGAGLEAGAR ALMDSLDAHPGAWTSMFASCNLLRPVMAARPMVVLGLSISKYYGMAGNDRVFQAGNWASLLGGKNACPLLIFDRTRKFVL (SEQ ID NO: 566). [0391] In some embodiments, a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is a UL39 polypeptide or antigenic fragment thereof. In some embodiments, a UL39 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of RTFGSAPRLTEDDFGLLNYALAEMRRLCLDLPPVPPNAYTPYHLREYATRLVNGFKPLVRRSARLYRILGVLVHLRIRTREASFEEWM RSKEVDLDFGLTERLREHEAQLMILAQALNPYDCLIHSTPNTLVERGLQSALKYEEFYLKRFGGHYMESVFQMYTRIAGFLA (SEQ ID NO: 567). [0392] In some embodiments, a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is a UL49 polypeptide or antigenic fragment thereof. In some embodiments, a UL49 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of KMTRGAPKASATPATDPARGRRPAQADSAVLLDAPAPTASGRTKTPAQGLAKKLHFSTAPPSPTAPWTPRVAGFNKRVFCAAVG (SEQ ID NO: 568). [0393] In some embodiments, a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is a UL9 polypeptide or antigenic fragment thereof. In some embodiments, a UL9 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of LLNNYDVLVLDEVMSTLGQLYSPTMQQLGRVDALMLRLLRTCPRIIAMDATANAQLVDFLCSLRGEKNVHVVIGEYAMPGFSARRCL FLPRLGPEVLQAALRRRGPAGGAPPPDAPPDATFFGELEARLAGGDNVCIFSSTVSFAEVVARFCRQFTDRVLLLHSLTPPGDVTTWG RYRVVIYTTVVTVGLSFDPPHFDSMFAYVKPMNYGPDMVSVYQSLGRVRTLRKGELLIYMDGSGARSEPV (SEQ ID NO: 569). [0394] In some embodiments, a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is a UL30 polypeptide or antigenic fragment thereof. In some embodiments, a UL30 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of ISCLLYDLSTTALEHILLFSLGSCDLPESHLSDLASRGLPAPVVLEFDSEFEMLLAFMTFVKQYGPEFVTGYNIINFDWPFVLTKLTEIYK VPLDGYGRMNGRGVFRVWDIGQSHF (SEQ ID NO: 570). In some embodiments, a UL30 polypeptide or antigenic fragment thereof comprises an amino acid sequence of GLLPCLHVAATVTTIGREMLLATRAYVHARWAEFDQLLADFPEAAGMRAPGPYSM (SEQ ID NO: 571). [0395] In some embodiments, a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is a UL40 polypeptide or antigenic fragment thereof. In some embodiments, a UL40 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of TSQCPDINHLRSLSILNRWLETELVFVGDEEDVSKLSEGELGFYRFLFAFLSAADDLVTENLGGLSGLFEQKDILHYYVEQECIEVVHS RVYNIIQLVLFHNNDQARRAYVARTINHPAIRVKVDWLEARVRECDSIPEKFILMILIEGVFFAASFAAIAYLRTNNLLR (SEQ ID NO: 572). [0396] In some embodiments, a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is a UL5 polypeptide or antigenic fragment thereof. In some embodiments, a UL5 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of HEFGNLMKVLEYGLPITEEHMQFVDRFVVPESYITNPANLPGWTRLFSSHKEVSAYMAKLHAYLKVTREGEFVVFTLPVLTFVSVKEF DEYRRL (SEQ ID NO: 573). In some embodiments, a UL5 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of ELFGEVFESAPFSTYVDNVIFRGCELLTGSPRGGLMSVALQTDNYTLMGYTYTRVFAFAEELRRRHATAGVAEFLEESPLPYIVLRDQ HGFMSVVNTNI (SEQ ID NO: 574). [0397] In some embodiments, a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is a UL52 polypeptide or antigenic fragment thereof. In some embodiments, a UL52 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of SVAAPVEVTALYATDGCVITSSLALLTNCLLGAEPLYIFSYDAYRSDAPNGPTGAPTEQERFEGSRALYRDAGGLNGDSFRVTFCLLGT EVGVTHHPKGRTRPMFVCRFERADDVAVLQDALGRGTPLLPAHVTATLDLEATFALHANIIMALTVAIVHNAPARIGSGSTAPLYEPG ESMRSVV (SEQ ID NO: 575). [0398] In some embodiments, a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is a UL1 polypeptide or antigenic fragment thereof. In some embodiments, a UL1 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of RTPADDVSWRYEAPSVIDYARIDGIFLRYHCPGLDTFLWDRHAQRAYLVNPFLFAAGFLEDLSHSVFPADTQETT (SEQ ID NO: 576). [0399] In some embodiments, a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is a UL19 polypeptide or antigenic fragment thereof. In some embodiments, a UL19 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of DGRLLHNTQARAADAADDRPHRPADWTVHHKIYYYVLVPAFSRGRCCTAGVRFDRVYATLQNMVVPEIAPGEECPSDPVTDPAHPL HPANLVANTVKRMFHN (SEQ ID NO: 577). [0400] In some embodiments, a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is a UL21 polypeptide or antigenic fragment thereof. In some embodiments, a UL21 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of SPTQKLAVYYYLIHRERRMSPFPALVRLVGRYIQRHGLYVPAPDEPTLADAMNGL (SEQ ID NO: 578). [0401] In some embodiments, a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is a UL27 polypeptide or antigenic fragment thereof. In some embodiments, a UL27 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of NYTEGIAVVFKENIAPYKFKATMYYKDVTVSQVWFGHRYSQFMGIFEDRAPVPFEEV (SEQ ID NO: 579). In some embodiments, a UL27 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of SVYPYDEFVLATGDFVYMSPFYGYREGSH (SEQ ID NO: 580). [0402] In some embodiments, a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is a UL46 polypeptide or antigenic fragment thereof. In some embodiments, a UL46 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of GLASDPHYDYIRHYASAAKQALGEVELSGGQLSRAILAQYWKYLQTVVPSGLDIPDDPAGDCDPSLHVLLRPTLLPKLLVRAPFKSGA AAAKYAAAVAGLRDAAHRLQQYMFFMRPADPSRPSTDTALRLSELLAYVSVLYHWASWMLWTADKYV (SEQ ID NO: 581). [0403] In some embodiments, a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is a UL47 polypeptide or antigenic fragment thereof. In some embodiments, a UL47 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of GPDAAVFRSSLGSLLYWPGVRALLGRDCRVAARYAGRMTYIATGALLARFNPGAVKCVLPREAAFAGRVL (SEQ ID NO: 582). [0404] In some embodiments, a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is a UL48 polypeptide or antigenic fragment thereof. In some embodiments, a UL48 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of ALFNRLLDDLGFSAGPALCTMLDTWNEDLFSGFPTNADMYRECKFLSTLPSDVIDWGDAHVPERSPIDIRAHGDVAFPTLPATRDEL PSYYEAMAQFFRGELRA (SEQ ID NO: 583). [0405] In some embodiments, a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is a UL25 polypeptide or antigenic fragment thereof. In some embodiments, a UL25 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of FLWEDQTLLRATANTITALAVLRRLLANGNVYADRLDNRLQLGMLIPGAVPAEAIARGASGLDSGAIKSGDNNLEALCVNYVLPLYQA DPTVELTQLFPGLAALCL (SEQ ID NO: 584). [0406] In some embodiments, a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is a US1 polypeptide or antigenic fragment thereof. In some embodiments, a US1 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of DDASDGWLVDTPPRKSKRPRINLRLTSSPDRRAGVVFPEV (SEQ ID NO: 585). In some embodiments, a US1 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of PASLPGIAHAHRRSARQAQMRSGAAWTLDLHYIRQCVNQL (SEQ ID NO: 586). [0407] In some embodiments, a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is a US8 polypeptide or antigenic fragment thereof. In some embodiments, an US8 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of ILSPTAPSVYPHSEGRKSRRPLTTFGSGSPGRRHSQASYPSVLW (SEQ ID NO: 587). In some embodiments, a US8 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of GLAWLASTVNLEFQHASPQHAGLYLCVVYVDDHIHAWGHMTISTAAQYRNAVVEQHLPQRQPEPVEPTRPHVRAPHPAPSARGPL RL (SEQ ID NO: 588). In some embodiments, a US8 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of KLLWAAEPLDACGPLRPSWVALWPPRRVLETVVDAACMRAPEPLAIAYSPPFPAGDEGLYSELAWRDRVAVVNESLVIYGALETDSG LYTLSVVGLSDEARQVASVVLVVEPAP (SEQ ID NO: 589). [0408] In some embodiments, a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is a US12 polypeptide or antigenic fragment thereof. In some embodiments, an US12 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of EDREAARTAVTDPELPLLCPPDV (SEQ ID NO: 590). [0409] In some embodiments, a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is an UL50 polypeptide or antigenic fragment thereof. In some embodiments, an UL50 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of ANGATVIQPSLRVLRAADGPEACYVLGRSSLNARGLLVMPTRWPSGHACAFVVCNLTGVPVTL (SEQ ID NO: 591). [0410] In In some embodiments, a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is an UL26 polypeptide or antigenic fragment thereof. In some embodiments, an UL26 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of APLPDRAVPIYVAGFLALYDSGDPGELALDPDTVRAALPPENPLPINVDHRARCEVGRVLAVVNDPRGPFFVGLIACVQLERVLETAAS AAIFERRGPALSREERLLYLITNYLPSVSLSTKRRGDEVPPDRTLFAHVALCAIGRRLGTIVTYDTSLDAA (SEQ ID NO: 592). In some embodiments, an UL26 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of HYPPPPAHPYPGMLFAGPSPLEAQIAALVGAIAADRQAGGLPAAAGDHGIRGSAKRRRHEVEQPEYDCG (SEQ ID NO: 593). [0411] In some embodiments, a polyribonucleotide provided herein encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof (e.g., HSV-1 and/or HSV- 2), wherein at least one antigen is an US10 polypeptide or antigenic fragment thereof. In some embodiments, an US10 polypeptide or antigenic fragment thereof comprises or consists of an amino acid sequence of SSPRQRTYVLPRVGIHNAPASDTRAPKRANSRHRADRPPESPGSELYPLNAQALAHLQMLPADHRAFFRTVIEVSRLCALNTHDPPPP LAGARVGQEAQLVHTQWLRANRESSPLWPWRTAAMNFIAAAAPCVQTHRHMHDLLMACAFWC (SEQ ID NO: 594). 1. Intermediate early [0412] In some embodiments, an HSV (e.g., HSV-1 and/or HSV-2) T-cell antigen for use in accordance with the present disclosure is an intermediate early protein or an antigenic fragment thereof. In some embodiments, a polyribonucleotide provided herein encodes one or more HSV (e.g., HSV-1 and/or HSV-2) antigens, wherein at least one HSV T-cell antigen comprises an intermediate early protein or an antigenic fragment thereof. In some embodiments, a polyribonucleotide provided herein encodes one or more HSV (e.g., HSV-1 and/or HSV-2) T-cell antigens, wherein each of the HSV T-cell antigens comprises an intermediate early protein or an antigenic fragment thereof. [0413] In some embodiments, an HSV-2 T-cell antigen for use in accordance with the present disclosure is an intermediate early protein or an antigenic fragment thereof. In some embodiments, a polyribonucleotide provided herein encodes one or more HSV-2 antigens, wherein at least one HSV-2 antigen comprises an intermediate early protein or an antigenic fragment thereof. In some embodiments, a polyribonucleotide provided herein encodes one or more HSV-2 antigens, wherein each of the HSV-2 antigens comprises an intermediate early protein or an antigenic fragment thereof. [0414] In some embodiments, an HSV-2 T-cell antigen for use in accordance with the present disclosure comprises an RL2 polypeptide or antigenic fragment thereof, an RS1 polypeptide or antigenic fragment thereof, a UL54 polypeptide or antigenic fragment thereof, or a combination thereof. [0415] In some embodiments, a polyribonucleotide provided herein encodes one or more of an RL2 polypeptide or antigenic fragment thereof, an RS1 protein or antigenic fragment thereof, and a UL54 protein or antigenic fragment thereof. 2. Early [0416] In some embodiments, an HSV (e.g., HSV-1 and/or HSV-2) T-cell antigen for use in accordance with the present disclosure is an early protein or an antigenic fragment thereof. In some embodiments, a polyribonucleotide provided herein encodes one or more HSV (e.g., HSV-1 and/or HSV-2) T-cell antigens, wherein at least one HSV T-cell antigen comprises an early protein or an antigenic fragment thereof. In some embodiments, a polyribonucleotide provided herein encodes one or more HSV (e.g., HSV-1 and/or HSV-2) T-cell antigens, wherein each of the HSV T-cell antigens comprises an early protein or an antigenic fragment thereof. [0417] In some embodiments, an HSV (e.g., HSV-1, HSV-2, or a combination) T-cell antigen for use in accordance with the present disclosure is an early protein or an antigenic fragment thereof. In some embodiments, a polyribonucleotide provided herein encodes one or more HSV T-cell antigens, wherein at least one HSV T-cell antigen comprises an early protein or an antigenic fragment thereof. In some embodiments, a polyribonucleotide provided herein encodes one or more HSV T-cell antigens, wherein each of the HSV T-cell antigens comprises an early protein or an antigenic fragment thereof. In some embodiments, the HSV T-cell antigen is a HSV-2 T-cell antigen. [0418] In some embodiments, an HSV (e.g., HSV-1, HSV-2, or a combination) T-cell antigen for use in accordance with the present disclosure comprises a UL29 polypeptide or antigenic fragment thereof, a UL39 polypeptide or antigenic fragment thereof, a UL49 polypeptide or antigenic fragment thereof, a UL9 polypeptide or antigenic fragment thereof, or a combination thereof. [0419] In some embodiments, a polyribonucleotide encoding a T-cell string provided herein encodes a UL29 polypeptide or antigenic fragment thereof, a UL39 polypeptide or antigenic fragment thereof, a UL49 polypeptide or antigenic fragment thereof, and a UL9 polypeptide or antigenic fragment thereof. [0420] In some embodiments, an HSV (e.g., HSV-1, HSV-2, or a combination) T-cell antigen for use in accordance with the present disclosure comprises a UL30 polypeptide or antigenic fragment thereof, a UL40 polypeptide or antigenic fragment thereof, a UL5 polypeptide or antigenic fragment thereof, a UL52 polypeptide or antigenic fragment thereof, or a combination thereof. [0421] In some embodiments, a polyribonucleotide encoding a T-cell string provided herein encodes one or more of a UL30 polypeptide or antigenic fragment thereof, a UL40 polypeptide or antigenic fragment thereof, a UL5 polypeptide or antigenic fragment thereof, and a UL52 polypeptide or antigenic fragment thereof. [0422] In some embodiments, an HSV (e.g., HSV-1, HSV-2, or a combination) T-cell antigen for use in accordance with the present disclosure comprises a UL29 polypeptide or antigenic fragment thereof, a UL39 polypeptide or antigenic fragment thereof, a UL49 polypeptide or antigenic fragment thereof, a UL9 polypeptide or antigenic fragment thereof, a UL30 polypeptide or antigenic fragment thereof, a UL40 polypeptide or antigenic fragment thereof, a UL5 polypeptide or antigenic fragment thereof, a UL52 polypeptide or antigenic fragment thereof, or a combination thereof. [0423] In some embodiments, a polyribonucleotide encoding a T-cell string provided herein encodes one or more of a UL29 polypeptide or antigenic fragment thereof, a UL39 polypeptide or antigenic fragment thereof, a UL49 polypeptide or antigenic fragment thereof, a UL9 polypeptide or antigenic fragment thereof, a UL30 polypeptide or antigenic fragment thereof, a UL40 polypeptide or antigenic fragment thereof, a UL5 polypeptide or antigenic fragment thereof, and a UL52 polypeptide or antigenic fragment thereof. 3. Late [0424] In some embodiments, an HSV (e.g., HSV-1 and/or HSV-2) antigen for use in accordance with the present disclosure is a late protein or an antigenic fragment thereof. In some embodiments, a polyribonucleotide provided herein encodes one or more HSV (e.g., HSV-1 and/or HSV-2) antigens, wherein at least one HSV antigen comprises a late protein or an antigenic fragment thereof. In some embodiments, a polyribonucleotide provided herein encodes one or more HSV (e.g., HSV-1 and/or HSV-2) antigens, wherein each of the HSV antigens comprises a late protein or an antigenic fragment thereof. [0425] In some embodiments, an HSV (e.g., HSV-1, HSV-2, or a combination) T-cell antigen for use in accordance with the present disclosure is a late protein or an antigenic fragment thereof. In some embodiments, a polyribonucleotide provided herein encodes one or more HSV antigens, wherein at least one HSV antigen comprises a late protein or an antigenic fragment thereof. In some embodiments, a polyribonucleotide provided herein encodes one or more HSV antigens, wherein each of the HSV antigens comprises a late protein or an antigenic fragment thereof. In some embodiments, an HSV T-cell antigen is an HSV-2 T-cell antigen. [0426] In some embodiments, an HSV (e.g., HSV-1, HSV-2, or a combination) T-cell antigen for use in accordance with the present disclosure comprises a UL1 polypeptide or antigenic fragment thereof, a UL19 polypeptide or antigenic fragment thereof, a UL21 polypeptide or antigenic fragment thereof, a UL27 polypeptide or antigenic fragment thereof, a UL46 polypeptide or antigenic fragment thereof, a UL47 polypeptide or antigenic fragment thereof, a UL48 polypeptide or antigenic fragment thereof, a UL25 polypeptide or antigenic fragment thereof, or a combination thereof. [0427] In some embodiments, a polyribonucleotide encoding a T-cell string provided herein encodes one or more of a UL1 polypeptide or antigenic fragment thereof, a UL19 polypeptide or antigenic fragment thereof, a UL21 polypeptide or antigenic fragment thereof, a UL27 polypeptide or antigenic fragment thereof, a UL46 polypeptide or antigenic fragment thereof, a UL47 polypeptide or antigenic fragment thereof, a UL48 polypeptide or antigenic fragment thereof, and a UL25 polypeptide or antigenic fragment thereof. Exemplary polyribonucleotide sequences encoding antigenic fragments. Table 9: Exemplary polyribonucleotide sequences encoding antigenic fragments SEQ ID Antigen Optimized Nucleotide Sequence NO c c g a a a g c gc c c c g a a a g c gc
Figure imgf000148_0001
uccaggcgauacuggaacuccugcuccugccucuggcgagaucgccccuccuaauucuacaagaagc gccagcgagagccggcacagaugauaa 611 US7 [HSV-2 Opt3 gguccuaccgugucucucgugucugacagccugguagacgccggugcugugggcccucagggcuuc a a cc cg c a ac gg g u a c c u a u ug c a u a c c u a u g c a a c g a cc g c c c a g c c a cc
Figure imgf000149_0001
acauccacgccuggggccacaugaccaucuccaccgccgcccaguaccgcaacgccgugguggagcag caccugccccagcgccagcccgagcccguggagcccacccgcccccacgugcgcgccuaa 615 US8 [HSV-2 Opt1 agcagaaccagcuggaaaagagugaccagcggcgaggauguggugcugcuuccugcuccugcuggc uc u u g gu ag ac u c ca a g u u uc ag g a a a cu c gg ca a gc uc cc g u u u u cg g g ag ac a ga g ac c u g ua
Figure imgf000150_0001
ugcuggguguucacggaccaccccaccuccacguugcuuugcugaagccagaauggaacccgugccu ggucuggcuuggcuggcaucaacugucaaccuggaguuccagcaugccucuccacagcacgcaggcc uguaucucugcgugguguacguugacgaucacauccaugcgugggggcauaugaccaucagcacag cc g ga ga cg aa gu u cc ac ca a cc cc ac g gu
Figure imgf000151_0001
the plurality of polyribonucleotides comprises (a) a first set of polyribonucleotides that comprises: (i) a polyribonucleotide that encoding an antigenic portion of HSV gC as provided herein, (ii) a polyribonucleotide that encoding an antigenic portion of HSV gD as provided herein, and (iii) a polyribonucleotide that encoding an antigenic portion of HSV gE as provided herein, and (b) a second set of polyribonucleotides that encode one or more T-cell string polypeptides, wherein the one or more T-cell string polypeptides each comprise one or more HSV T-cell antigens or antigenic portions thereof. [0429] In some embodiments, a first set of polyribonucleotides further comprises a polyribonucleotide encoding an HSV glycoprotein B (gB) , variant thereof, or an antigenic portion thereof, a polyribonucleotide encoding an HSV glycoprotein G (gG) or an antigenic portion thereof, a polyribonucleotide encoding an HSV glycoprotein H (gH) or an antigenic portion thereof, a polyribonucleotide encoding an HSV glycoprotein I (gI) or an antigenic portion thereof, a polyribonucleotide encoding an HSV glycoprotein L (gL) or an antigenic portion thereof, or a combination thereof. C. Additional Proteins [0430] In some embodiments, additional polyribonucleotides are used with the first and/or second set of polyribonucleotides. In some embodiments, an additional polyribonucleotide encodes a polypeptide comprising ICP0, ICP4, ICP22, VP16, ICP47, VHS, or US3. In some embodiments, an additional set of polyribonucleotides encodes a set of polypeptides, wherein the set of polypeptides comprises ICP0, ICP4, ICP22, VP16, ICP47, VHS, US3, or any combination thereof. In some embodiments, an additional set of polyribonucleotides encodes a set of polypeptides, wherein the set of polypeptides comprises ICP0, ICP4, or both. [0431] In some embodiments, a first set of polyribonucleotides comprises: a polyribonucleotide encoding an HSV glycoprotein B (gB), variant thereof, or an antigenic portion thereof; a polyribonucleotide encoding an HSV glycoprotein C (gC) , variant thereof, or an antigenic portion thereof; and a polyribonucleotide encoding an HSV glycoprotein D (gD) , variant thereof, or an antigenic portion thereof. In some embodiments, an additional set of polyribonucleotides encodes a set of polypeptides, wherein the set of polypeptides comprises ICP0, ICP4, or both. [0432] In some embodiments, a combination provided herein comprises a polyribonucleotide encoding an HSV glycoprotein B (gB), variant thereof, or an antigenic portion thereof; a polyribonucleotide encoding an HSV glycoprotein C (gC) , variant thereof, or an antigenic portion thereof; a polyribonucleotide encoding an HSV glycoprotein D (gD) , variant thereof, or an antigenic portion thereof; a polyribonucleotide encoding an HSV ICP0, variant thereof, or an antigenic portion thereof; and a polyribonucleotide encoding an HSV ICP4, variant thereof, or an antigenic portion thereof. In some embodiments, a combination provided herein comprises a polyribonucleotide encoding an antigenic portion HSV glycoprotein B (gB), variant thereof, or an antigenic portion thereof; a polyribonucleotide encoding an HSV glycoprotein C (gC) , variant thereof, or an antigenic portion thereof; a polyribonucleotide encoding an HSV glycoprotein D (gD) , variant thereof, or an antigenic portion thereof; a polyribonucleotide encoding an HSV ICP0, variant thereof, or an antigenic portion thereof; and a polyribonucleotide encoding an HSV ICP4, variant thereof, or an antigenic portion thereof. In some embodiments, a combination provided herein comprises a polyribonucleotide encoding an antigenic portion HSV glycoprotein B (gB); a polyribonucleotide encoding an antigenic portion of HSV glycoprotein C (gC); a polyribonucleotide encoding an HSV glycoprotein D (gD) or an antigenic portion thereof; a polyribonucleotide encoding an antigenic portion of HSV ICP0; and a polyribonucleotide encoding an antigenic portion of HSV ICP4. In some embodiments, a combination provided herein comprises a polyribonucleotide encoding an antigenic portion HSV-2 glycoprotein B (gB); a polyribonucleotide encoding an antigenic portion of HSV-2 glycoprotein C (gC); a polyribonucleotide encoding an HSV-2 glycoprotein D (gD) or an antigenic portion thereof; a polyribonucleotide encoding an antigenic portion of HSV-2 ICP0; and a polyribonucleotide encoding an antigenic portion of HSV-2 ICP4. D. Secretory Signals [0433] Provided herein are polyribonucleotides encoding polypeptides (e.g., GP polypeptide) comprising (i) an HSV glycoprotein or antigenic portion thereof and (ii) a secretory signal. Provided herein are also polyribonucleotides encoding polypeptides (e.g., T-cell string polypeptides) comprising (i) one or more HSV T-cell antigens or antigenic portions thereof and (ii) secretory signal. [0434] In some embodiments, a GP polypeptide comprises a secretory signal. In some embodiments, a GP polypeptide comprises (i) an HSV gC or antigenic portion thereof and (ii) a secretory signal. In some embodiments, a GP polypeptide comprises (i) an HSV gD or antigenic portion thereof and (ii) a secretory signal. In some embodiments, a GP polypeptides comprises (i) an HSV gD or antigenic portion thereof and (ii) a secretory signal. In some embodiments, a GP polypeptide comprises (i) an HSV gB, variant thereof, or one or more antigenic portions thereof and (ii) a secretory signal. [0435] In some embodiments, a T-cell string polypeptide comprises a secretory signal. In some embodiments, a T-cell string polypeptide comprises (i) one or more HSV T-cell antigens or antigenic portions thereof and (ii) a secretory signal. [0436] In some embodiments, a secretory signal is functional in mammalian cells. In some embodiments, a utilized secretory signal is a heterologous secretory signal. In some embodiments, a secretory signal comprises or consists of a human secretory signal. In some embodiments, a secretory signal comprises or consists of an IL2 secretory signal. [0437] In some embodiments, a secretory signal comprises or consists of a viral secretory signal. In some embodiments, a viral secretory signal comprises or consists of an HSV secretory signal (e.g., an HSV-1 or HSV-2 secretory signal). In some embodiments, a secretory signal comprises or consists of an HSV-1 secretory signal. In some embodiments, a secretory signal comprises or consists of an HSV-2 secretory signal. [0438] In some embodiments, an HSV secretory signal comprises or consists of an HSV glycoprotein D (gD) secretory signal (e.g., an HSV-1 or HSV-2 gD secretory signal). In some embodiments, an HSV secretory signal comprises or consists of an HSV-1 gD secretory signal. In some embodiments, an HSV-1 gD secretory signal comprises one or more additional amino acids. In some embodiments, an HSV-1 gD secretory signal comprises KY at the C terminus of the signal sequence. In some embodiments, an HSV secretory signal comprises or consists of an HSV-2 gD secretory signal. In some embodiments, an HSV-2 gD secretory signal comprises one or more additional amino acids. In some embodiments, an HSV-2 gD secretory signal comprises KY, KYA, KYAL (SEQ ID NO: 842), or KYALA (SEQ ID NO: 841) at the C terminus of the signal sequence. [0439] In some embodiments, an HSV secretory signal comprises or consists of an HSV glycoprotein C (gC) secretory signal (e.g., an HSV-1 or HSV-2 gC secretory signal). In some embodiments, an HSV secretory signal comprises or consists of an HSV-2 gC secretory signal. [0440] In some embodiments, an HSV secretory signal comprises or consists of an HSV glycoprotein E (gE) secretory signal (e.g., an HSV-1 or HSV-2 gE secretory signal). In some embodiments, an HSV secretory signal comprises or consists of an HSV-1 gE secretory signal. In some embodiments, an HSV secretory signal comprises or consists of an HSV-2 gE secretory signal. In some embodiments, an HSV-2 gE secretory signal comprises one or more additional amino acids. In some embodiments, an HSV-2 gE secretory signal comprises RTS. In some embodiments, an HSV-2 secretory signal comprises A20V, A21V, A22V substitutions. [0441] In some embodiments, an HSV secretory signal comprises or consists of an HSV glycoprotein B (gB) secretory signal (e.g., an HSV-1 or HSV-2 gB secretory signal). In some embodiments, an HSV secretory signal comprises or consists of an HSV-1 gB secretory signal. In some embodiments, an HSV-1 gB secretory signal comprises one or more additional amino acids. In some embodiments, an HSV-1 gB secretory signal comprises AP at the C terminus of the signal sequence. In some embodiments, an HSV secretory signal comprises or consists of an HSV-2 gB secretory signal. [0442] In some embodiments, an HSV secretory signal comprises or consists of an HSV glycoprotein I (gI) secretory signal (e.g., an HSV-1 or HSV-2 gI secretory signal). In some embodiments, an HSV secretory signal comprises or consists of an HSV-1 gI secretory signal. In some embodiments, an HSV-1 gI secretory signal comprises one or more additional amino acids. In some embodiments, an HSV secretory signal comprises or consists of an HSV- 2 gI secretory signal. In some embodiments, an HSV-2 gI secretory signal comprises an additional leucine residue at the C terminus of the signal sequence. [0443] In some embodiments, a secretory signal comprises or consists of an Ebola spike glycoprotein (EboZ). In some embodiments, an EboZ secretory signal comprises one or more additional amino acids. In some embodiments, an EboZ secretory signal comprises IP at the C terminus of the signal sequence. [0444] In some embodiments, a secretory signal is characterized by a length of about 15 to 30 amino acids. [0445] In some embodiments, a secretory signal is positioned at the N-terminus of a polyribonucleotide. In some embodiments, a secretory signal preferably allows transport of a polyribonucleotide with which it is associated into a defined cellular compartment, preferably a cell surface, endoplasmic reticulum (ER) or endosomal- lysosomal compartment. [0446] In some embodiments, polyribonucleotides comprising an HSV antigen do not comprise a secretory signal. In some embodiments, polyribonucleotides comprising an HSV antigen further comprise a codon initiation start site. [0447] In some embodiments, a secretory signal is one listed in Table 10, or a secretory signal having 1, 2, 3, 4, or 5 amino acid differences relative thereto. In some embodiments, a secretory signal is selected from those included in the Table 10 below and/or those encoded by the sequences in Table 11 and/or Table 12 below. Table 10: Example secretory signals SEQ ID Secretory signal Sequence (Amino Acid) NO:
Figure imgf000154_0001
33 HSV-2 gE MARGAGLVFFVGVWVVSCLAAAP 292 HSV-2 gE – LVVVP (SEQ ID NO: MARGAGLVFFVGVWVVSCLVVVP
Figure imgf000155_0001
399 human Ig heavy chain signal MDWTWRFLFVVAAATGVQS peptide 5
Figure imgf000156_0001
SEQ ID Secretory signal Versions Sequence (Nucleotide) NO: G G G G G G G C G
Figure imgf000156_0002
42 HSV-2 gD-KYA Wild type ATGGGCCGCCTGACCTCCGGCGTGGGCACCGCCGCCCTGCTGGTG GTGGCCGTGGGCCTGCGCGTGGTGTGCGCCAAGTACGCC G C G C G G C G C C C A G T G G G TG G T C T
Figure imgf000157_0001
410 HSV-1 gD ATGGGGGGGGCTGCCGCCAGGTTGGGGGCCGTGATTTTGTTTGTC GTCATAGTGGGCCTCCATGGGGTCCGCAGCAAATAT G G G G G G G G G G G G G G G G G
Figure imgf000158_0001
222 HSV-1 gD - KY Version 4.1 ATGGGCGGAGCTGCGGCTCGCCTCGGAGCCGTCATTCTGTTCGTG GTGATCGTGGGTCTGCATGGGGTCAGAGGAAAGTAC G T G A G T G T G T G T G T G A G T G T G G G G G G G
Figure imgf000159_0001
231 HSV-2 gE – RTS Version 1 ATGGCTAGAGGTGCCGGCCTGGTGTTCTTTGTTGGCGTGTGGGTC GTGTCCTGTCTGGCCGCTGCTCCTAGAACATCT TC G T G T G T A T G T G A G G T A A A A C
Figure imgf000160_0001
434 human Ig kappa chain ATGGACATGAGGGTCCCTGCTCAGCTCCTGGGGCTCCTGCTGCTC signal peptide 1 TGGCTCTCAGGTGCCAGATGT G G T G T G A C T G G G
Figure imgf000161_0001
SEQ ID Secretory signal Versions Sequence (Nucleotide) NO: U U U U U G U
Figure imgf000161_0002
57 HSV-2 gD Version 3 AUGGGCAGACUGACCUCCGGCGUGGGCACCGCCGCCCUGCUGGU GGUGGCCGUGGGCCUGAGAGUGGUGUGCGCC U G U U U G U G U G G U U G G U U G C C C C G G G
Figure imgf000162_0001
307 HSV-2 gE Version 2.3 AUGGCGAGAGGAGCCGGGCUCGUGUUCUUUGUGGGCGUAUGGG UCGUUUCCUGCCUGGCUGCCGCACCC G G G G G G U G G G G G G G G G
Figure imgf000163_0001
241 HSV-1 gD - KY Version 3 AUGGGAGGAGCUGCUGCUAGAUUGGGAGCUGUGAUUCUGUUUG UGGUGAUUGUGGGACUGCAUGGAGUGAGAGGAAAAUAC G G G C G G G C U U G G G G G C G G G C U U G U
Figure imgf000164_0001
248 HSV-2 gI +L Version 1 AUGCCUGGCAGAUCUCUGCAAGGACUGGCCAUCCUCGGACUGUG GGUUUGCGCAACAGGACUG G G G G G G G A A A A A A A A U A A G G
Figure imgf000165_0001
458 human Ig heavy AUGGACUGGACCUGGAGGAUCCUCUUCUUGGUGGCAGCAGCAAC chain signal peptide AGGUGCCCACUCG 4 A A A U U U G G G G C G C C C C C
Figure imgf000166_0001
E. Transmembrane Regions [0448] In some embodiments, a polypeptide (e.g., a GP polypeptide or a T-cell string polypeptide) described herein includes a transmembrane region. In some embodiments, a polyribonucleotide described herein encodes a polypeptide (e.g., a GP polypeptide or a T-cell string polypeptide) that comprises a transmembrane region. In some embodiments, a transmembrane region is located at the N-terminus of a polypeptide (e.g., a GP polypeptide or a T-cell string polypeptide). In some embodiments, a transmembrane region is located at the C-terminus of a polypeptide (e.g., a GP polypeptide or a T-cell string polypeptide). In some embodiments, a transmembrane region is not located at the N-terminus or C-terminus of a polypeptide (e.g., a GP polypeptide or a T-cell string polypeptide). In some embodiments, a polypeptide does not include a transmembrane region. [0449] Transmembrane regions are known in the art, any of which can be utilized in a polypeptide described herein. In some embodiments, a transmembrane region comprises or is a transmembrane region of Hemagglutinin (HA) of Influenza virus, Env of HIV-1, equine infectious anaemia virus (EIAV), murine leukaemia virus (MLV), mouse mammary tumor virus, G protein of vesicular stomatitis virus (VSV-G), Rabies virus, or a seven transmembrane domain receptor. [0450] In some embodiments, a polypeptide comprises an HSV transmembrane region. In some embodiments, an HSV transmembrane region is an HSV-1 or HSV-2 transmembrane region. In some embodiments, an HSV transmembrane region is an HSV-2 gD transmembrane region. In some embodiments, an HSV transmembrane region is an HSV-2 gC transmembrane region. In some embodiments, an HSV transmembrane region is an HSV-2 gE transmembrane region. In some embodiments, an HSV transmembrane region comprises or consists of an HSV gD transmembrane region, e.g., comprising or consisting of an amino acid sequence of GLIAGAVGGSLLAALVICGIVYWMRRHTQKAPKRIRLPHIR (SEQ ID NO: 468). In some embodiments, a utilized transmembrane region is a heterologous transmembrane region. In some embodiments, a heterologous transmembrane region comprises or consists of a human transmembrane region. In some embodiments, a human transmembrane region comprises or consists of a human decay accelerating factor glycosylphosphatidylinositol (hDAF-GPI) anchor region. In some embodiments, an hDAF-GPI anchor region comprises or consists of an amino acid sequence of PNKGSGTTSGTTRLLSGHTCFTLTGLLGTLVTMGLLT (SEQ ID NO: 469). In some embodiments, an HSV transmembrane region comprises or consists of an HSV gB transmembrane region, e.g., comprising or consisting of an amino acid sequence of MSNPFGALAVGLLVLAGLAAAFFAFRYVMRL (SEQ ID NO: 470) or MSNPFGALAVGLLVLAGLVAAFFAFRYVLQL (SEQ ID NO: 471). [0451] In some embodiments, a viral transmembrane region comprises or consists of a vesicular stomatitis virus G (VSV-G) transmembrane region. In some embodiments, a VSV-G transmembrane region comprises or consists of an amino acid sequence of IASFFFIIGLIIGLFLVLRVGIYLCIKLKHTKKRQIYTDIEMN (SEQ ID NO: 245). [0452] Example transmembrane regions are provided in the following Table 13: Table 13: Example transmembrane regions SEQ Transmembrane Region Sequence (Amino Acid) ID
Figure imgf000167_0001
468 HSV-1 gD GLIAGAVGGSLLAALVICGIVYWMRRHTQKAPKRIRLPHIR 469 hDAF-GPI anchor region PNKGSGTTSGTTRLLSGHTCFTLTGLLGTLVTMGLLT
Figure imgf000168_0001
[0453] Provided herein are polypeptides (e.g., GP polypeptides and/or T-cell string polypeptides) that comprise a trafficking signal. In some embodiments, a trafficking signal is an MHC Class I Trafficking Signal (MITD). [0454] In some embodiments, at least one of the one or more GP polypeptides comprise a MITD. In some embodiments, at least one of the one or more GP polypeptides comprise (i) an HSV gC or antigenic portion thereof and (ii) a MITD. In some embodiments, at least one of the one or more GP polypeptides comprise (i) an HSV gD or antigenic portion thereof and (ii) a MITD. In some embodiments, at least one of the one or more GP polypeptides comprise (i) an HSV gD or antigenic portion thereof and (ii) a MITD. In some embodiments, at least one of the one or more GP polypeptides comprise (i) an HSV gB, variant thereof, or one or more antigenic portions thereof and (ii) a MITD. [0455] In some embodiments, at least one of the one or more T-cell string polypeptides comprises a MITD. [0456] In some embodiments, a MITD comprises or consists of an amino acid sequence that is at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to IVGIVAGLAVLAVVVIGAVVATVMCRRKSSGGKGGSYSQAASSDSAQGSDVSLTA (SEQ ID NO: 473). In some embodiments, a MITD comprises or consists of an amino acid sequence of IVGIVAGLAVLAVVVIGAVVATVMCRRKSSGGKGGSYSQAASSDSAQGSDVSLTA (SEQ ID NO: 473). G. Multimerization Regions [0457] Provided herein are polypeptides (e.g., GP polypeptides and/or T-cell string polypeptides) that comprise one or more multimerization regions. In some embodiments, a multimerization region is a heterologous multimerization region. In some embodiments, a heterologous multimerization region comprises a dimerization, trimerization or tetramerization region. [0458] In some embodiments, at least one of the one or more GP polypeptides comprise one or more multimerization regions. In some embodiments, at least one of the one or more GP polypeptides comprise (i) an HSV gC or antigenic portion thereof and (ii) one or more multimerization regions. In some embodiments, at least one of the one or more GP polypeptides comprise (i) an HSV gD or antigenic portion thereof and (ii) one or more multimerization regions. In some embodiments, at least one of the one or more GP polypeptides comprise (i) an HSV gD or antigenic portion thereof and (ii) one or more multimerization regions. In some embodiments, at least one of the one or more GP polypeptides comprise (i) an HSV gB, variant thereof, or one or more antigenic portions thereof and (ii) one or more multimerization regions. [0459] In some embodiments, at least one of the one or more T-cell string polypeptides comprises one or more multimerization regions. [0460] In some embodiments, a multimerization region is one described in WO2017/081082, which is incorporated herein by reference in its entirety (e.g., SEQ ID NOs: 1116-1167, or fragments or variants thereof). Example trimerization and tetramerization regions include, but are not limited to, engineered leucine zippers, fibritin foldon domain from enterobacteria phage T4, GCN4pll, GCN4-pll, and p53. [0461] In some embodiments, a multimerization region comprising or consisting of the amino acid sequence GYIPEAPRDGQAYVRKDGEWVLLSTFL (SEQ ID NO: 474). [0462] In some embodiments, a GP polypeptide comprising an HSV-2 gB or antigenic portion thereof, as described herein, is able to form a trimeric complex. For example, a GP polypeptide comprising an HSV-2 gB or antigenic portion thereof, as described herein, may further comprise a multimerization region allowing formation of a multimeric complex, such as for example a trimeric complex of an HSV-2 gB described herein. In some embodiments, a multimerization region allowing formation of a multimeric complex comprises a trimerization region, for example, a trimerization region described herein. In some embodiments, a GP polypeptide comprising an HSV-2 gB or antigenic portion thereof, as described herein, further includes a T4-fibritin-derived “foldon” trimerization region, for example, to increase its immunogenicity. H. Linkers [0463] In some embodiments, a polypeptide (e.g., a GP polypeptide and/or T-cell string polypeptide) includes one or more linkers. In some embodiments, at least one of the one or more T-cell string polypeptides comprises a secretory signal. [0464] In some embodiments, a linker is or comprises 2, 3, 4, 5, 6, 7, 8, 9, 10 or more amino acids. In some embodiments, a linker is or comprises no more than about 30, 25, 20, 15, 10 or fewer amino acids. A linker can include any amino acid sequence and is not limited to any particular amino acids. In some embodiments, a linker comprises one or more glycine (G) amino acids. In some embodiments, a linker comprises one or more serine (S) amino acids. In some embodiments, a linker comprises a glycine-serine linker. A “glycine-serine linker” as used herein refers to a linker that comprises predominantly (e.g., 80% or more) glycine and serine amino acids. In some embodiments, a linker includes amino acids selected based on a cleavage predictor to generate highly-cleavable linkers. [0465] In some embodiments, a linker is or comprises SGGGGSGGGGS (SEQ ID NO: 475). In some embodiments, a linker is or comprises GSPGSGSGS (SEQ ID NO: 476). In some embodiments, a linker is or comprises GGSGGGGSGG (SEQ ID NO: 477). In some embodiments, a linker is one presented in Table 14. In some embodiments, a linker is or comprises a sequence as set forth in WO2017/081082, which is incorporated herein by reference in its entirety (see SEQ ID NOs: 1509-1565, or a fragment or variant thereof). [0466] In some embodiments, a GP polypeptide described herein comprises a linker between a C-terminal region or fragment thereof and a transmembrane region. In some embodiments, a GP polypeptide described herein comprises a linker after a minor repeat sequence. In some embodiments, a T-cell string polypeptide comprises one or more linkers. In some embodiments, a T-cell string polypeptide comprises a one or more linkers separating one or more HSV T-cell antigens or antigenic portions thereof. [0467] Example linkers are provided in the following Table 14: Table 14: Example linkers SEQ ID NO: Sequence (Amino Acid) 475 SGGGGSGGGGS
Figure imgf000170_0002
[0468] In some embodiments, a polyribonucleotide encodes a GP polypeptide, wherein the GP polypeptide comprises an HSV glycoprotein or antigenic portion thereof and a secretory signal. In some embodiments, a polyribonucleotide encodes a GP polypeptide, wherein the GP polypeptide comprises an antigenic portion of an HSV glycoprotein and a secretory signal. [0469] Example polyribonucleotide constructs encoding a gC, gD, or gE as described herein are provided in Table 15 below. Table 15: Example Polyribonucleotide Constructs Amino struct Secr RNA Con etory signal Glycoprotein Additional modifications acid SEQ SEQ ID ID NO NO 1597 IL2 gC - 65 104 1598 gD2 gD - 70 116 1599 IL2 gE - 73 121 1600 IL2 gC Codon optimized (Version 2)
Figure imgf000170_0001
65 106 1601 gD2 gD Codon optimized (Version 2) 70 118 1602 IL2 gE Codon optimized (Version 2) 73 123 1658 gD2 gC Codon optimized (Version 2); KYA 67 109 secretory signal extension; ∆EAM mutation 1659 gD2 gD Codon optimized (Version 2); ∆EAM 70 119 mutation 1660 gD2 gE Codon optimized (Version 2); KYA 75 127 secretory signal extension; ∆EAM mutation 1622 gD2 gC KYA secretory signal extension 67 108 1623 gD2 gE KYA secretory signal extension 75 125 1624 gD2 gC Codon optimized (Version 2); KYA 67 110 secretory signal extension; 4 point mutations relative to 1658 1625 gD2 gE Codon optimized (Version 2); KYA 75 128 secretory signal extension; 3 point mutations relative to 1660 1873 IL2 gC Codon optimized (Version 1) 65 105 1874 gD2 gD Codon optimized (Version 1) 70 117 1875 IL2 gE Codon optimized (Version 1) 73 122 1876 IL2 gC Codon optimized (Version 3) 65 107 1877 gD2 gD Codon optimized (Version 3) 70 120 1878 IL2 gE Codon optimized (Version 3) 73 124 1907 gD2 gC Codon optimized (Version 2); KYALA 68 113 secretory signal extension 1908 gD2 gC Codon optimized (Version 1); KYALA 68 112 (SEQ ID NO: 841) secretory signal extension 1909 gD2 gC Codon optimized (Version 3); KYALA 68 114 (SEQ ID NO: 841) secretory signal extension 1910 gC2 gC Codon optimized (Version 2); 131 190 1911 gD2 gE Codon optimized (Version 1); KYA 75 126 secretory signal extension 1912 gD2 gE Codon optimized (Version 3); KYA 75 129 secretory signal extension 1913 gE2 gE Codon optimized (Version 2); 132 130 2140 gD1 gC Codon optimized (Version 4); KY 159 192 secretory signal extension 2141 gB1 gC Codon optimized (Version 4); AP 160 196 secretory signal extension 2138 gE2 gC Codon optimized (Version 4); RTS 162 204 secretory signal extension 2143 gD1 gE Codon optimized (Version 4); KY 164 212 secretory signal extension; 3 point mutations relative to 1602 2538 gD1 gC Codon optimized (Version 2); KY 159 190 secretory signal extension 2541 gB1 gC Codon optimized (Version 2); AP 160 194 secretory signal extension 2544 gI2 gC Codon optimized (Version 2); TGL 161 198 secretory signal extension 2547 gE2 gC Codon optimized (Version 2); RTS 162 202 secretory signal extension 2550 EboZ gC Codon optimized (Version 2); IP 163 206 secretory signal extension 2553 gD1 gE Codon optimized (Version 2); KY 164 210 secretory signal extension; 3 point mutations relative to 1602 2537 gD1 gC Codon optimized (Version 1); KY 159 189 secretory signal extension 2540 gB1 gC Codon optimized (Version 1); AP 160 193 secretory signal extension 2543 gI2 gC Codon optimized (Version 1); TGL 161 197 secretory signal extension 2546 gE2 gC Codon optimized (Version 1); RTS 162 201 secretory signal extension 2549 EboZ gC Codon optimized (Version 1); IP 163 205 secretory signal extension 2552 gD1 gE Codon optimized (Version 1); KY 164 209 secretory signal extension; 3 point mutations relative to 1602 2539 gD1 gC Codon optimized (Version 3); KY 159 191 secretory signal extension 2542 gB1 gC Codon optimized (Version 3); AP 160 195 secretory signal extension 2545 gI2 gC Codon optimized (Version 3); TGL 161 199 secretory signal extension 2548 gE2 gC Codon optimized (Version 3); RTS 162 203 secretory signal extension 2551 EboZ gC Codon optimized (Version 3); IP 163 207 secretory signal extension 2554 gD1 gE Codon optimized (Version 3); KY 159 192 secretory signal extension; 3 point mutations relative to 1602 2790 gE2 gE Codon optimized (Version 2); 132 322 mutations to improve RNA secondary structure 2788 gE2 gE Codon optimized (Version 2) 132 323 2792 gE2 gE Codon optimized (Version 2) 132 324 2791 gE2 gE Codon optimized (Version 2); LVVV 327 325 secretory signal extension; artificial splicing site to increase secretion 2789 - gE Codon optimized (Version 2) 328 326 2786 gC2 gC Codon optimized (Version 2); 131 319 mutations to improve RNA secondary structure 2787 gD2 gC Codon optimized (Version 2); KYALA 68 318 secretory signal extension; mutations to improve RNA secondary structure 2785 gC2 gC Codon optimized (Version 2) 131 320 2784 gC2 gC Codon optimized (Version 2) 131 321 3215 gD gC Codon optimized (Version 2); KY 159 348 secretory signal extension; ∆EAM deletion 3216 gB gC Codon optimized (Version 2); AP 160 350 secretory signal extension; ∆EAM deletion 3217 gE gC Codon optimized (Version 2); RTS 162 351 secretory signal extension; ∆EAM deletion 3233 gD gC Codon optimized (Version 2); KY 159 349 secretory signal extension; ∆EAM deletion 3234 gD gD Codon optimized (Version 2); second 70 352 stop codon 3235 gD gE Codon optimized (Version 2); KYA 75 353 secretory signal extension; second stop codon 4068 gD2 gE Codon optimized (Version 3); KYA 700 703 secretory signal extension; second stop codon 4069 gE2 gE Codon optimized (Version 2); 701 704 +4AA(LVVV); 1 unpaired cysteine 4070 gE2 gE Codon optimized (Version 2); 702 705 +4AA(LVVV); 3 unpaired cysteines AGNT1 gB1 gB - 710 712 AGNT2 gB1 gB Codon optimized (Version 1) 710 713 AGNT3 gB1 gB Codon optimized (Version 2) 710 714 AGNT4 gB1 gB Codon optimized (Version 3) 710 715 4058 gB2 gB - 708 716 4059 gB2 gB Codon optimized (Version 1) 708 717 4060 gB2 gB Codon optimized (Version 2) 708 718 4061 gB2 gB Codon optimized (Version 3) 708 719 4066 gD1 gC Codon optimized (Version 2); 707 728 +2AA(KY) secretory signal extension; ∆EAM deletion 4067 gD2 gD Codon optimized (Version 2); ∆EAM 706 729 deletion; second stop codon 4068 gE2 gE Codon optimized (Version 2); 700 703 +3AA(KYA) secretor signal extension; ∆EAM deletion; second stop codon [0470] In some embodiments, a GP polypeptide as described herein (or encoded by a polyribonucleotide as described herein) comprises an antigenic portion of HSV-2 gC and a secretory signal. Example combinations of an antigenic portion of HSV-2 gC and a secretory signal are provided in Table 16 below, along with example corresponding amino acid sequences. [0471] In some embodiments, a GP polypeptide as described herein (or encoded by a polyribonucleotide as described herein) comprises an antigenic portion of HSV-2 gD and a secretory signal. Example combinations of an antigenic portion of HSV-2 gD and a secretory signal are provided in Table 16 below, along with example corresponding amino acid sequences. [0472] In some embodiments, a GP polypeptide as described herein (or encoded by a polyribonucleotide as described herein) comprises an antigenic portion of HSV-2 gE and a secretory signal. Example combinations of an antigenic portion of HSV-2 gE and a secretory signal are provided in Table 16 below, along with example corresponding amino acid sequences. Table 16: Example GP polypeptides comprising a secretory signal and an antigenic portions of an HSV-2 glycoprotein SEQ ID Secretory Antigenic NO: signal fragment Sequence (Amino acids) T IR P R W R SS PR RY Q Y G T A IP S DP PP Q T PP G YP P S LE A G PE G A PL P Q T
Figure imgf000175_0001
YYPGNRAEFVWFEDGRRVFDPAQIHTQTQENPDGFSTVSTVTSAAVGGQGPP RTFTCQLTWHRDSVSFSRRNASGTASVLPRPTITMEFTGDHAVCTAGCVPEG VTFAWFLGDDSSPAEKVAVASQTSCGRPGTATIRSTLPVSYEQTEYICRLAGYP S LA G G YY T TF GI S C V E T G G C P V D A L L E P A G G YY T TF GI SN N N L TC G C RL
Figure imgf000176_0001
69 IL2 HSV-2 gD MRMQLLLLIALSLALVTNSADPSLKMADPNRFRGKNLPVLDQLTDPPGVKR VYHIQPSLEDPFQPPSIPITVYYAVLERACRSVLLHAPSEAPQIVRGASDEARKH (30-331) TYNLTIAWYRMGDNCAIPITVMEYTECPYNKSLGVCPIRTQPRWSYYDSFSAV P TL H VL QI P R D V R A W H L D V S S R M E AY R T V GC HI G L D A SY D P E L P A R
Figure imgf000177_0001
SYAGCSRTTPPPRCFAEARMEPVPGLAWLASTVNLEFQHASPQHAGLYLCVVY VDDHIHAWGHMTISTAAQYRNAVVEQHLPQRQPEPVEPTRPHVRA T SP V PR R A T SP V PR R A L ES A G CL VN QP P AI EA PT V GC HI G L D A SY D G D G T SP V PR
Figure imgf000178_0001
VIPEVSHVRGVTVHMETPEAILFAPGETFGTNVSIHAIAHDDGPYAMDVVWM RFDVPSSCAEMRIYEACLYHPQLPECLSPADAPCAVSSWAYRLAVRSYAGCSR TTPPPRCFAEARMEPVPGLAWLASTVNLEFQHASPQHAGLYLSVVYVDDHIHA T P V PR R A VL QI P R P S LA G G YY T TF GI S RP A T VR G F Q M R W N CT V G A EH NI C RV T
Figure imgf000179_0001
TYAADRFKQVDGFYARDLTTKARATAPTTRNLLTTPKFTVAWDWVPKRPSVC TMTKWQEVDEMLRSEYGGSFRFSSDAISTTFTTNLTEYPLSRVDLGDCIGKDA RDAMDRIFARRYNATHIKVGQPQYYQANGGFLIAYQPLLSNTLAELYVREHLR RV V A F G A EH NI C RV T C A R RV V A F G LA E N
Figure imgf000180_0001
SEQ ID Secretory Antigen Version Sequences (Polynucleotides) NO: signal /OFR T G CG T C T T C T C C A T G A A C A G C C
Figure imgf000180_0002
CGTGCTGCCCCGCCCCACCATCACCATGGAGTTCACCGGCGACCACG CCGTGTGCACCGCCGGCTGCGTGCCCGAGGGCGTGACCTTCGCCTGG TTCCTGGGCGACGACTCCTCCCCCGCCGAGAAGGTGGCCGTGGCCTC C C T G TG A T T A C T C A A T G AC C A C G A C CC T C G A T G T G C C C C G G C C C G A A T A G C
Figure imgf000181_0001
AGAGACTCCGTAAGCTTCAGCCGTAGAAACGCCTCTGGAACCGCCAG TGTGTTGCCGAGGCCGACTATCACGATGGAATTCACAGGCGATCATG CCGTCTGTACTGCCGGCTGTGTGCCAGAAGGCGTAACCTTCGCTTGG C C C T G CG T C G C A G C G G A G C C A C G A C C T C T T GT C C C A G G C C C C TG A CA C A C CC C
Figure imgf000182_0001
CGTGTCCACCGTGACCTCCGCCGCCGTGGGCGGCCAGGGCCCCCCCC GCACCTTCACCTGCCAGCTGACCTGGCACCGCGACTCCGTGTCCTTC TCCCGCCGCAACGCCTCCGGCACCGCCTCCGTGCTGCCCCGCCCCAC T CC G C C T G A T A C A G A A A GT G AC C GC A C G A G A T G C G T T G A T A C A G A A A GT A AC C GC
Figure imgf000183_0001
CCTTTAAGGCCACATGCACTGCTGCGACCTACTACCCTGGCAACAGA GCCGAGTTTGTCTGGTTTGAGGATGGTCGGCGAGTATTCGATCCAGC CCAGATTCACACACAAACGCAGGAAAATCCGGACGGCTTCAGCACAG A G A T G C G T T G A T A C A G A A A GT G AC C GC A C G A G A T G C G T GT C C T C A C C G C T T G
Figure imgf000184_0001
AACTGACCCTGGAAACCCAGGGCATGTACTACTGGGTCTGGGGCAGA ACCGATAGACCAAGCGCCTATGGCACCTGGGTTCGAGTGCGAGTGTT CAGACCTCCTAGCCTGACCATCCATCCTCACGCCGTTCTGGAAGGCCA AG G C G A C G A A A G T C T C C G A T G C C C G C C C AC C A T T A C C C G C T C T C C G A T G C
Figure imgf000185_0001
CCAGGAGGACAGCTGGTCTATGACTCAGCGCCCAATAGGACAGACCC ACATGTTATCTGGGCAGAAGGAGCCGGTCCTGGGGCCTCTCCACGGC TGTACTCAGTTGTTGGCCCGCTTGGACGACAGAGACTCATCATCGAG C C C AC C A T T A C C C G C GT CC C C C A G AG A T A T A C A G A C GT A CA C T C C C C C A C C G T A
Figure imgf000186_0001
GATACGGTGTCGCTTCCCTAACAGTACAAGGACTGAGTTTCGGCTGC AGATCTGGCGTTATGCCACAGCTACTGACGCAGAGATTGGTACCGCC CCCAGTTTGGAGGAAGTGATGGTCAACGTGTCCGCACCCCCAGGAGG A A C CC C T A A C T A G C G A A G C A C C G T A C C G A A C CC C T A A C T A G C G A A G C A C C
Figure imgf000187_0001
TCAACCCAGGAAAGCGACAAAGTCCAAGGCCAGCACCGCAAAACCCG CTCCTCCTCCAAAGACTGGGCCCCCAAAGACAAGTAGCGAACCAGTT CGGTGCAACAGGCATGACCCACTTGCACGCTATGGGTCAAGAGTCCA C C G A A C CC C T A A C T A G C G A A G C G T C C T C G C G G C A AC C T G G C T T C C T G T
Figure imgf000188_0001
165 HSV-1 gD HSV-2 gC Version 1 ATGGGCGGAGCTGCTGCTAGACTGGGAGCCGTGATCCTGTTCGTGGT TATCGTGGGACTGCATGGCGTGCGGGGCAAGTATAGCCCTGGCAGAA CCATCACAGTGGGCCCTAGAGGCAACGCCTCTAATGCCGCTCCTAGC A C C G A C C T G C G C CA G T A T G A T C A G A T AA C T C C G A C A T G A C C T A A C T A G C G A A
Figure imgf000189_0001
GTACATCTGCAGGCTGGCCGGCTATCCCGATGGGATTCCAGTCCTGG AGCACCACTGATAA T A C C G T A C C G A A C CC C T A A C T A G C G A A G T AA C T C C G A C A T G A C C T A A C T A G C
Figure imgf000190_0001
AGAAGGCGTAACCTTCGCTTGGTTTCTCGGGGATGACTCAAGTCCTG CAGAGAAAGTGGCTGTGGCCTCTCAGACGAGCTGCGGTCGACCAGGA ACAGCTACCATTCGCAGCACTCTGCCCGTGTCCTACGAGCAGACGGA G T A T T A A T T T G G G G C T A TT A GT T T A G A C T C T G C C G GC G A C A TT G C G C CA G T A C
Figure imgf000191_0001
ACATGCCAATTGACCTGGCACCGCGACTCAGTTAGCTTTAGCCGCCG GAATGCCAGTGGGACCGCCAGTGTTCTCCCAAGGCCGACAATCACCA TGGAGTTCACTGGCGACCATGCAGTGTGCACAGCTGGGTGTGTCCCA C A G A G C T CC G G G C A G C C A G G A G TC T C G G G C G T C T CT C AC AG AG G T A G C C A G G A G
Figure imgf000192_0001
GCAACAGAGCCGAGTTTGTCTGGTTTGAGGATGGTCGGCGAGTATTC GATCCAGCCCAGATTCACACACAAACGCAGGAAAATCCGGACGGCTT CAGCACAGTGTCCACGGTGACCTCTGCTGCAGTTGGTGGACAAGGAC AA G C G C A A TG C C CT GC GT G G G C G A T G G A G C T C AA G C G C A A G C C A C G T A C A T TT T C
Figure imgf000193_0001
CCTTCCGCATATGGCACTTGGGTGAGAGTTCGCGTCTTTCGGCCCCC TTCTCTCACCATCCATCCTCATGCCGTGCTCGAAGGCCAGCCCTTTAA GGCCACATGCACTGCTGCGACCTACTACCCTGGCAACAGAGCCGAGT T C A A T G A AC T G G TC C C C C G GA A T A A T G G G AA T C C TT AC T T G A G TG C CC C A A T C G T C
Figure imgf000194_0001
AGATCCTCACGTGATTTGGGCAGAAGGTGCGGGGCCTGGGGCCTCCC CAAGGCTCTACTCAGTGGTTGGACCCCTTGGGAGACAGCGGCTGATC ATCGAGGAACTGACTCTCGAAACCCAAGGTATGTACTACTGGGTATG A G C AT G G G TC A GA T C G T A C C G C C C A T G A G C C C A G C G G CC T C A C GT C T A G C T C A
Figure imgf000195_0001
TGGTCAACGTGTCCGCACCCCCAGGAGGACAGCTGGTCTATGACTCA GCGCCCAATAGGACCGATCCCCACGTGATCTGGGCAGAAGGAGCCGG TCCTGGGGCCTCTCCACGGCTGTACTCAGTTGTTGGCCCGCTTGGAC G C CC G T A G G C C T C A C T A TT A C C T CA T C T A T G AT C A T G A TG T T C G A GT A T CC G C A C
Figure imgf000196_0001
TGCCACAGACGCAGAGATTGGTACCGCTCCCAGCCTGGAGGAGGTCA TGGTGAACGTGTCAGCGCCTCCGGGTGGCCAGCTGGTCTACGACTCT GCCCCAAATCGAACCGACCCTCACGTCATCTGGGCTGAAGGAGCGGG A G AC C C A G C G G T C G C C GT A C G C T C T A G G G G C A G T C A C A G A G C A C G C C A C G
Figure imgf000197_0001
TGCAACAGGCATGACCCACTTGCACGCTATGGGTCAAGAGTCCAGAT ACGGTGTCGCTTCCCTAACAGTACAAGGACTGAGTTTCGGCTGCAGA TCTGGCGTTATGCCACAGCTACTGACGCAGAGATTGGTACCGCCCCC A T TT T C C A T T C A A T G A AC T G A AC AC A G A A A A C C G C A G G C C A G C G C A A G CA T G T
Figure imgf000198_0001
CCTCCAAAAACAGGACCTCCAAAAACCTCCTCTGAACCTGTGAGATGC AACAGACATGATCCTCTGGCAAGATATGGATCAAGAGTGCAGATCAG ATGCAGATTTCCAAATTCCACCAGAACAGAATTCAGACTCCAGATCTG C G C G A C C A T A G G G AT A A A T C G T C CA C G T A A CA G G G CC T T C C C A G A G T AT A G A
Figure imgf000199_0001
GCATCCCCAGCCCTGGCAGAACCATCACAGTGGGCCCTAGAGGCAAC GCCTCTAATGCCGCTCCTAGCGCCTCTCCTAGAAACGCCTCTGCTCCC AGAACCACACCTACACCTCCACAGCCTAGAAAGGCCACCAAGAGCAAG AG G C T T C T T G A TT A G C A A G G G A T G G G G A G GA C G G G G CT C T G C C T G A C C T G A G
Figure imgf000200_0001
TCCTACGAGCAGACGGAGTACATCTGCAGGCTGGCCGGCTATCCCGA TGGGATTCCAGTCCTGGAGCACCACTGATAA G C G A G A A GA C T A C A G T CT C G GA G GT A A A T T G G C G C AA AG G C C GT C CA T G A T T G GA C T T T
Figure imgf000201_0001
GCACTGCTGGGTGTGTACCCGAAGGCGTGACATTTGCCTGGTTTCTG GGAGATGACTCCTCACCCGCAGAAAAGGTCGCCGTTGCATCTCAGAC CAGTTGTGGCAGGCCCGGGACTGCTACCATCCGCAGCACTCTGCCGG A GT C T T C G C C C A A C T C G A C GT C A CA GT C G A C C C G G G A T C T G G A A A A CA T C G A
Figure imgf000202_0001
CCACATACAGCCTAGTCTTGAGGACCCTTTTCAGCCACCGTCTATCCC CATTACCGTGTACTATGCCGTGCTGGAACGCGCGTGTAGGTCAGTTC TGCTGCATGCCCCATCCGAAGCCCCCCAGATCGTCAGAGGAGCTTCT GA A C T A A G G A T T A A C T C G A C C T GA A C T A A G G A T T A A C T C G A C C T GA A C T A A
Figure imgf000203_0001
TCCTGGAACATAGGGCCAGAGCCAGCTGCAAGTATGCCCTTCCCCTG CGGATTCCGCCTGCAGCATGTCTGACCTCAAAAGCCTACCAGCAAGG GGTGACTGTGGACAGCATTGGCATGCTGCCTCGTTTCATTCCCGAGA T T A A C GT TC G A C C C G G G AC A A G G A C C A A C T G CC T A C A G T G A G C A G C C G C C C C
Figure imgf000204_0001
CCCCAGCACGCCGGCCTGTACCTGTGCGTGGTGTACGTGGACGACCA CATCCACGCCTGGGGCCACATGACCATCTCCACCGCCGCCCAGTACC GCAACGCCGTGGTGGAGCAGCACCTGCCCCAGCGCCAGCCCGAGCCC T G C T A T A TA C GT G T C T A CA G G A T C C C A G T G C T A C A C T G CG CA G T A T A T T C G AC C
Figure imgf000205_0001
CATGCGTGGGGGCATATGACCATCAGCACAGCTGCCCAGTACCGCAA TGCCGTCGTGGAGCAGCACCTCCCCCAACGGCAGCCAGAACCAGTGG AGCCCACTCGGCCTCATGTGCGAGCCTGA T G C T A C A GA G G T C C A C C G AT C C G C T A G GT CT C C C CT C C A G C G C A G T T C C C C T C G
Figure imgf000206_0001
CCACATGACCATCTCCACCGCCGCCCAGTACCGCAACGCCGTGGTGG AGCAGCACCTGCCCCAGCGCCAGCCCGAGCCCGTGGAGCCCACCCGC CCCCACGTGCGCGCCTAA GT T T C C GC A GT G T C A AC C G T T C CT C A C G A G G T T C G C G C G C G A T A C T G G C G T A G T
Figure imgf000207_0001
CTCTGCGTGGTGTACGTTGACGATCACATCCATGCGTGGGGGCATAT GACCATCAGCACAGCTGCCCAGTACCGCAATGCCGTCGTGGAGCAGC ACCTCCCCCAACGGCAGCCAGAACCAGTGGAGCCCACTCGGCCTCAT T T C G C G C G C G A T A C T G G C G T A G T T C T T T C G C G C G C G A T A C T G G C G T A G
Figure imgf000208_0001
TCAACCTGGAGTTCCAGCATGCCTCTCCACAGCACGCAGGCCTGTAT CTCTGCGTGGTGTACGTTGACGATCACATCCATGCGTGGGGGCATAT GACCATCAGCACAGCTGCCCAGTACCGCAATGCCGTCGTGGAGCAGC T GT T C C C GC C T G C CC T AC C T A A G C C C A G C C G G T A G A GA G A T C C C A G G T C C G C C
Figure imgf000209_0001
CCGTGCCTGGTCTGGCTTGGCTGGCATCAACTGTCAACCTGGAGTTC CAGCATGCCTCTCCACAGCACGCAGGCCTGTATCTCTGCGTGGTGTA CGTTGACGATCACATCCATGCGTGGGGGCATATGACCATCAGCACAG G G T A G A GA G A T C C C A G G T C C G C C C A G G G T A G A GA G A T C C C A G G T C C G C C C
Figure imgf000210_0001
CAGCATGCCTCTCCACAGCACGCAGGCCTGTATCTCTGCGTGGTGTA CGTTGACGATCACATCCATGCGTGGGGGCATATGACCATCAGCACAG CTGCCCAGTACCGCAATGCCGTCGTGGAGCAGCACCTCCCCCAACGG G TT A C C AA G C A T C CC C C G G T C A A C C C C C G T A G A GA G A T C C C A G G T C C G C C C A
Figure imgf000211_0001
CGTTGACGATCACATCCATGCGTGGGGGCATATGACCATCAGCACAG CTGCCCAGTACCGCAATGCCGTCGTGGAGCAGCACCTCCCCCAACGG CAGCCAGAACCAGTGGAGCCCACTCGGCCTCATGTGCGAGCCTGA T C T T A C G C A G C T A T A A G T C G AC C G C GT A C A T TG C G C G A C CT C T G C T T C G C T T C
Figure imgf000212_0001
CCCAGAGACAGCCCGAGCCTGTGGAACCTACAAGACCTCATGTTCGG GCCTGATAA GT A A G T AT C G A C A G C C T A G G C T C A C C C G T A T A T TG T C T A C G AC T T GC G GT TG A TG A G C
Figure imgf000213_0001
ACAGCTGCTCAGTACAGAAATGCTGTGGTGGAACAGCACCTGCCACA GAGACAGCCAGAACCAGTGGAACCAACCAGACCACATGTGAGAGCTT GATAA T G C G G T G T G A C A CC AC C T GT C CT C A C G A G G T T C G C G C G C G A T A C T G G C G T A G T
Figure imgf000214_0001
CTCTGCGTGGTGTACGTTGACGATCACATCCATGCGTGGGGGCATAT GACCATCAGCACAGCTGCCCAGTACCGCAATGCCGTCGTGGAGCAGC ACCTCCCCCAACGGCAGCCAGAACCAGTGGAGCCCACTCGGCCTCAT T T C G AA C T G A G T A G A GA G A T C C C A G G T C C G C C C A G G A G T A G A G G A G C C A C
Figure imgf000215_0001
ACCTCCAACCCATCCGAGAGTGATTCCCGAGGTCAGCCATGTGCGCG GCGTAACTGTGCACATGGAGACGCCCGAAGCGATACTGTTTGCCCCT GGAGAGACATTCGGCACCAATGTGTCCATACACGCAATTGCGCACGA C C T TT A T T A G T T C G CC C C C T GA G G A A G A G G TA T G A C C T T C G C C G T G G A G GA G
Figure imgf000216_0001
CCTGCTACAGCCGCCCCCTGGTCAGCTTTCGGTACGAAGACCAGGGC CCGTTGGTCGAGGGGCAGCTGGGGGAGAACAACGAGCTGCGGCTGA CGCGCGATGCGATCGAGCCGTGCACCGTGGGACACCGGCGCTACTTC A C G A G G A G C C C G G G A C A G C T C A T A C C A G C A A G A A C G T C T C G G A A T T G CT G AC A
Figure imgf000217_0001
CGAGCTGACACTGTGGAACGAGGCCAGAAAGCTGAACCCCAACGCCA TTGCCAGCGTGACAGTTGGCCGTAGAGTGTCCGCTAGAATGCTGGGA GATGTGATGGCCGTGTCTACCTGTGTGCCTGTGGCCGCCGATAACGT T G G C T AT C G T T T C T C A A G A G A T CC C C A A G CC T AT T C A C C C C C C A C CA A G A C C A CT C
Figure imgf000218_0001
AGAGTAGGAAACCTCCCAACCCCACCCCACCTCCTCCTGGGGCTTCC GCTAACGCCAGCGTGGAGCGGATCAAAACAACAAGTAGCATTGAGTT TGCGCGCCTCCAGTTTACATATAACCATATCCAAAGGCATGTTAACGA G C GA A T T T G T A G G AT T T TT G T T G C A G G G TC A C C G T A T CA GT A G A T T AC T T T C AC G T C GA
Figure imgf000219_0001
GATTGCATTGGAAAAGATGCAAGAGATGCAATGGATAGAATCTTTGC AAGAAGATACAATGCAACCCACATCAAAGTGGGACAGCCTCAGTACTA CCAGGCAAATGGAGGATTTCTGATTGCTTACCAGCCTCTGCTGTCAA A T A G C GT G A CC C A A A A AA C T T G A G C A T T A A C G G C G C C A G G G G G C T G C A T A G G C
Figure imgf000220_0001
CAAGTTTACCGTGGCCTGGGACTGGGTGCCGAAGCGACCGGCGGTCT GCACCATGACCAAGTGGCAGGAGGTGGACGAGATGCTCCGCGCCGA GTACGGCGGCTCCTTCCGCTTCTCCTCCGACGCCATCTCGACCACCTT G G A A A A G A C C G C T G G T G A A TC C G T G T C CA A G G T C G G A GC A C C T A C C C A C C CC C A
Figure imgf000221_0001
TCGTGGAAGAAGTGGACGCCAGAAGCGTGTACCCCTACGACGAATTT GTGCTGGCCACCGGCGACTTCGTGTACATGAGCCCTTTCTACGGCTA CAGAGAGGGCAGCCACACCGAGCACACAAGCTATGCCGCCGACCGGT C C T C CC T T C T G AT G CT A T T C A G G C C T A C C G T A A G AC A AT G C A G A G CA G C G C T C C A C
Figure imgf000222_0001
CACTGCTAAATACGTCAGGAACAATATGGAAACAACAGCTTTTCACCG GGACGACCACGAAACGGATATGGAGCTGAAACCGGCCAAAGTGGCCA CCCGCACTAGCCGCGGCTGGCATACAACGGATCTGAAGTACAATCCA T G A T G C A G A G C C G C C C G T C A C C A T G G C A G A G T CT A T TC C A T G A GC A C C G G G
Figure imgf000223_0001
CTGTGGTGTTCAAAGAAAACATTGCTCCTTACAAATTCAAAGCAACCA TGTACTACAAAGATGTGACAGTGTCTCAGGTGTGGTTTGGACACAGA TACTCTCAGTTCATGGGAATCTTTGAAGATAGAGCACCTGTGCCTTTT C GA A AA G T G C A G A TT A A C G AC T TG C TT A G C A G G A C CC AT C G T A A TT G A G T G G G C G A GC A C
Figure imgf000224_0001
TGATGCAAATGCAACAGTGGCTGCTGGACATGCAACCCTGAGAGCAC ACCTGAGAGAAATCAAAGTGGAAAATGCTGATGCTCAGTTTTATGTG TGCCCTCCTCCAACAGGAGCAACAGTGGTGCAGTTTGAACAGCCAAG G A A TT C GA A AA G T G C A G A TT A A C G AC T TG C TT A G C A G G A C CC AT C G T A A T AA C T C C G A C A
Figure imgf000225_0001
CAGCTGGTCTATGACTCAGCGCCCAATAGGACCGATCCCCACGTGAT CTGGGCAGAAGGAGCCGGTCCTGGGGCCTCTCCACGGCTGTACTCAG TTGTTGGCCCGCTTGGACGACAGAGACTCATCATCGAGGAACTGACA C C T A A C T A G C G A A G A C C T C G A C C T GA A C T A A G G A T T A A C T G C T
Figure imgf000226_0001
SEQ ID Secretory Antigen Version Sequences (Polyribonucleotide) i l FR
Figure imgf000226_0002
104 IL2 HSV-2 gC AUGCGCAUGCAGCUGCUGCUGCUGAUCGCCCUGUCCCUGGCCCUGG UGACCAACUCCGCCUCCCCCGGCCGCACCAUCACCGUGGGCCCCCGC GGCAACGCCUCCAACGCCGCCCCCUCCGCCUCCCCCCGCAACGCCUCC C G C A G C G G A G C C C C C G G U C G A A C A C C C C G C C A U U C G U C C A U
Figure imgf000227_0001
ACACUGCCUGUCAGCUACGAGCAGACCGAGUACAUCUGUCGGCUGGC CGGCUAUCCUGAUGGCAUCCCUGUGCUGGAACACCACUGA C A C C C C C U U C G U A G G C C G G G C U G C G C G G A G C C A G A C C C
Figure imgf000228_0001
CCAGACCUCCUGCGGCAGACCCGGCACCGCCACCAUCAGAUCCACCCU GCCCGUGUCCUACGAACAGACCGAAUACAUCUGCAGACUGGCCGGCU ACCCCGAUGGCAUCCCCGUGCUGGAACACCACUGA A A C C G A C U G C C C G G C C U G C C G C G A A C A G A A A A U C A U C C U G
Figure imgf000229_0001
ACUAUCACGAUGGAAUUCACAGGCGAUCAUGCCGUCUGUACUGCCGG CUGUGUGCCAGAAGGCGUAACCUUCGCUUGGUUUCUCGGGGAUGAC UCAAGUCCUGCAGAGAAAGUGGCUGUGGCCUCUCAGACGAGCUGCGG G U C G A A C A G A A A A U C A U C C U G G C G G U C G A A C A G A A A A U C A U C
Figure imgf000230_0001
ACAGUGUCCACGGUGACCUCUGCUGCAGUUGGUGGACAAGGACCCCC UCGAACCUUCACCUGUCAGCUGACCUGGCACAGAGACUCCGUAAGCU UCAGCCGUAGAAACGCCUCUGGAACCGCCAGUGUGUUGCCGAGGCCG G C G G U G C G C G G C U G U C C A A G G A C C C U U G U G C G G C C C C U U C C G A C A
Figure imgf000231_0001
GGCCAGCCCUUUAAGGCCACAUGCACUGCUGCGACCUACUACCCUGG CAACAGAGCCGAGUUUGUCUGGUUUGAGGAUGGUCGGCGAGUAUUC GAUCCAGCCCAGAUUCACACACAAACGCAGGAAAAUCCGGACGGCUU AC A G G A U C G G G G C C C C U UU CC C GG A C G A G C U AC A G G A U C G G C GC C C A A C G C C U AG
Figure imgf000232_0001
AACUGACCCUGGAAACCCAGGGCAUGUACUACUGGGUGUGGGGCAGA ACCGAUAGACCCUCCGCCUACGGCACCUGGGUGAGAGUGAGAGUGUU CAGACCCCCUUCCCUGACCAUCCACCCCCACGCCGUGCUGGAAGGCCA G C C G U A C A G U C U C G A C C G A A C C C C C A C U G C G U C U C G A C C G
Figure imgf000233_0001
ACAGCUGGUCUAUGACUCAGCGCCCAAUAGGACCGAUCCCCACGUGA UCUGGGCAGAAGGAGCCGGUCCUGGGGCCUCUCCACGGCUGUACUCA GUUGUUGGCCCGCUUGGACGACAGAGACUCAUCAUCGAGGAACUGAC C C C C A C U G C G U C G U C G A C C G A A C C C C C A C U G C G U C G U C G A
Figure imgf000234_0001
GAUACGGUGUCGCUUCCCUAACAGUACAAGGACUGAGUUUCGGCUGC AGAUCUGGCGUUAUGCCACAGCUACUGACGCAGAGAUUGGUACCGCC CCCAGUUUGGAGGAAGUGAUGGUCAACGUGUCCGCACCCCCAGGAGG A A C C C C C A C U G C G U G G A C G A C C G A C G A C U U G C A C U C C A G A C C
Figure imgf000235_0001
UCAACCCAGGAAAGCGACAAAGUCCAAGGCCAGCACCGCAAAACCCGC UCCUCCUCCAAAGACUGGGCCCCCAAAGACAAGUAGCGAACCAGUUC GGUGCAACAGGCAUGACCCACUUGCACGCUAUGGGUCAAGAGUCCAG A C A U G A C C U C C C C C C G U G G U C G A C C G A A C C C C C A C U G C G U
Figure imgf000236_0001
349 HSV-1 gD HSV-2 gC Version AUGGGAGGAGCAGCUGCCAGACUCGGUGCCGUGAUCCUGUUCGUG 2.2 GUCAUUGUUGGCCUGCACGGGGUAAGGGGCAAGUACAGCCCCGGCA GAACCAUAACAGUAGGGCCACGGGGGAAUGCUUCCAAUGCUGCACC UC A C A C G C C G C G C U C A A A GA G G U C U G GG AG U UC UG GA AG A AC GG GA C G GA GA CC GC CU U AA CA AU UG UC AC AG
Figure imgf000237_0001
ACAGAAUACAUUUGCAGACUGGCUGGAUACCCUGAUGGAAUCCCUGU GCUGGAACACCACUGAUAA G G U C C G CA C A G A G G A C C G C G G A A G C G C A U C U A C A A A C C A C G U G U A G C A A G G
Figure imgf000238_0001
UACAGCUGGAUGUGUUCCUGAGGGCGUGACCUUCGCUUGGUUUCUG GGCGACGAUAGCAGCCCUGCCGAAAAAGUGGCUGUGGCCAGCCAGAC AAGCUGUGGCAGACCUGGAACCGCCACCAUCAGAAGCACACUGCCUG CU GU GC UU GA CC CA UA GA CA U GG UC CA UG G UG UA GA AC AC AC UU CU CU GA CC CC U G C A A G G U G C U AC U C C C U A C
Figure imgf000239_0001
UGCUGCAGUUGGUGGACAAGGACCCCCUCGAACCUUCACCUGUCAG CUGACCUGGCACAGAGACUCCGUAAGCUUCAGCCGUAGAAACGCCU CUGGAACCGCCAGUGUGUUGCCGAGGCCGACUAUCACGAUGGAAUU U C A A C A A A C A A G G A A A U U G A U C U G C C A A A U C U G U G G
Figure imgf000240_0001
CUGGAAGGACAGCCGUUCAAGGCAACAUGCACAGCAGCCACUUACUA UCCCGGAAACCGUGCUGAGUUUGUGUGGUUCGAGGAUGGGCGACGU GUAUUCGACCCUGCCCAGAUUCACACCCAGACACAGGAGAAUCCCGA A C U C A C C U C C G C C C A G G A C A G G A G C C G G C A A C C G C U C A A G A U
Figure imgf000241_0001
ACUUGGGUGAGAGUUCGCGUCUUUCGGCCCCCUUCUCUCACCAUCCA UCCUCAUGCCGUGCUCGAAGGCCAGCCCUUUAAGGCCACAUGCACUG CUGCGACCUACUACCCUGGCAACAGAGCCGAGUUUGUCUGGUUUGAG A G C C G C G C U A C C G C A U U G U A G A A G C C G U U G C A G C C C G C A C A G A
Figure imgf000242_0001
UGUACUACUGGGUGUGGGGCAGGACUGACAGGCCCAGUGCCUAUGG AACUUGGGUUAGGGUCCGCGUCUUUCGGCCACCCAGUCUGACCAUCC AUCCACAUGCCGUGCUGGAAGGCCAGCCCUUCAAAGCGACUUGCACU A C U C A G C A G C C A C U C U A G G U U C U A C A A G A G C C A C U A U A G G
Figure imgf000243_0001
GGCAGAAGGAGCCGGUCCUGGGGCCUCUCCACGGCUGUACUCAGUU GUUGGCCCGCUUGGACGACAGAGACUCAUCAUCGAAGAGCUGACACU GGAGACACAGGGGAUGUACUACUGGGUGUGGGGCCGUACUGACCGC U A G C U U C C G G U A A A A G A A G A U C C A G C A G A G A G C C A C G A C G
Figure imgf000244_0001
CUGGUGUAUGAUUCUGCUCCAAACAGAACAGAUCCUCAUGUGAUCUG GGCUGAAGGAGCUGGACCUGGAGCUUCUCCAAGACUGUACUCUGUG GUGGGACCUCUGGGAAGACAGAGACUGAUCAUUGAAGAACUGACCCU C A U U C G A C A U G U C A C U U A A U C U C A G A U U C A C G A C C A A A
Figure imgf000245_0001
GAUACGGCAGCCGGGUGCAGAUCAGAUGCAGAUUCCCCAACAGCACC CGGACCGAGUUCCGGCUCCAGAUUUGGAGAUACGCCACCGCCACAGA UGCCGAGAUUGGAACAGCCCCUAGCCUGGAAGAAGUGAUGGUCAACG C U U C U A C G G U U A C U G A U C A A C A C A G U U G C G A U C U G G A C
Figure imgf000246_0001
CAAGAACCACCCCAACCCCUCCUCAGCCAAGAAAAGCAACCAAAUCCA AAGCAUCCACAGCAAAACCUGCACCUCCUCCAAAAACAGGACCUCCAA AAACCUCCUCUGAACCUGUGAGAUGCAACAGACAUGAUCCUCUGGCA C U A A C U C A A U U U C A U C A C A A U C G A A U G U G G A C A G G A U U U
Figure imgf000247_0001
116 HSV-2 gD HSV-2 gD AUGGGCCGCCUGACCUCCGGCGUGGGCACCGCCGCCCUGCUGGUGG UGGCCGUGGGCCUGCGCGUGGUGUGCGCCAAGUACGCCCUGGCCGA CCCCUCCCUGAAGAUGGCCGACCCCAACCGCUUCCGCGGCAAGAACC G C U U C C A A C U C A C U G G C G A C C C C C G C A C G C U C C G C A C C U G U A
Figure imgf000248_0001
CGGAGUGCCCCUACAACAAGAGCCUCGGUGUUUGCCCUAUCAGGACA CAACCCAGGUGGAGCUAUUACGACAGUUUCAGCGCCGUGUCUGAGGA CAAUCUGGGGUUUCUGAUGCACGCACCCGCCUUCGAGACUGCCGGCA G C C U A A C A C C U G U A A A A G C C C U A A C
Figure imgf000249_0001
UUGCUUCCACCAGAACUCAGUGACACCACUAAUGCGACACAGCCAGA ACUUGUGCCUGAGGAUCCUGAAGAUAGCGCUCUGUUGGAGGAUCCA GCCGGUACUGUGUCCUCCCAGAUACCACCCAAUUGGCACAUUCCUU G A C G C U U A C A A C U C G A C U G G AC G C C C A A C G C C C U A C C AC G C C C C C C CC C
Figure imgf000250_0001
122 IL2 HSV-2 gE Version 1 AUGAGAAUGCAGCUGCUGCUCCUGAUCGCCCUGUCUCUGGCCCUGG UCACCAAUAGCAGAACCAGCUGGAAAAGAGUGACCAGCGGCGAGGAU GUGGUGCUGCUUCCUGCUCCUGCUGGCCCCGAGGAAAGAACAAGAGC C A U G U C C U C G U U C A U C G C C C A C G C C G G U A G C G G G C G C A U
Figure imgf000251_0001
124 IL2 HSV-2 gE Version 3 AUGAGAAUGCAGCUGCUGCUGCUGAUCGCCCUGUCCCUGGCCCUGG UGACCAACUCCAGAACCUCCUGGAAAAGAGUGACCUCCGGCGAAGAU GUGGUGCUGCUGCCCGCCCCCGCCGGCCCCGAAGAAAGAACCAGAGC C A U G G U G G C A G C C C G C A C G C C A G C C G C A A G A U G C A A G C
Figure imgf000252_0001
126 HSV-2 gD – HSV-2 gE Version 1 AUGGGCAGACUGACAUCUGGCGUGGGAACAGCUGCUCUGCUGGUGG KYA UUGCUGUGGGCCUGAGAGUCGUGUGUGCCAAAUACGCCAGAACCAGC UGGAAAAGAGUGACCAGCGGCGAGGAUGUGGUGCUGCUUCCUGCUC C C U C C U C G G U U U C U U C G U G C C C U G A C G A G A G C C C U G U
Figure imgf000253_0001
GGAGCAGCACCUCCCCCAACGGCAGCCAGAACCAGUGGAGCCCACUC GGCCUCAUGUGCGAGCCUGA C C C U G A C G A G A G C C C U G U C C C U G A C G A G A G C C C U G
Figure imgf000254_0001
GGGCAUAUGACCAUCAGCACAGCUGCCCAGUACCGCAAUGCCGUCGU GGAGCAGCACCUCCCCCAACGGCAGCCAGAACCAGUGGAGCCCACUC GGCCUCAUGUGCGAGCCUGAUAA C C G C C A C C U A C U G C C A A G C C G U C G U C A A C C A U G A U G
Figure imgf000255_0001
UGGUGUACGUUGACGAUCACAUCCAUGCGUGGGGGCAUAUGACCAU CAGCACAGCUGCCCAGUACCGCAAUGCCGUCGUGGAGCAGCACCUCC CCCAACGGCAGCCAGAACCAGUGGAGCCCACUCGGCCUCAUGUGCGA G C G U A C G U C U A A U C C A U U U G A U G U C A G U A G GA G G A G G A C G C C G U U C G U G
Figure imgf000256_0001
GAGUUCCAGCAUGCCUCUCCACAGCACGCAGGCCUGUAUCUCUGCGU GGUGUACGUUGACGAUCACAUCCAUGCGUGGGGGCAUAUGACCAUCA GCACAGCUGCCCAGUACCGCAAUGCCGUCGUGGAGCAGCACCUCCCC C G U A G A G G A G C C C G U C G U U A C C G U A G A G G A G C C C G U C G U
Figure imgf000257_0001
GGAACCCGUGCCUGGUCUGGCUUGGCUGGCAUCAACUGUCAACCUG GAGUUCCAGCAUGCCUCUCCACAGCACGCAGGCCUGUAUCUCUGCGU GGUGUACGUUGACGAUCACAUCCAUGCGUGGGGGCAUAUGACCAUCA C C G C U G U U G U A C U G G G C A C G G C A U A G C G U A G C G G U C C
Figure imgf000258_0001
CACAGUGAACCUGGAAUUUCAGCACGCCUCUCCACAGCACGCCGGCC UGUAUCUGUGUGUGGUGUACGUGGACGAUCACAUCCACGCCUGGGG CCACAUGACCAUCUCUACAGCCGCUCAGUACCGGAACGCCGUGGUUG G G C A G C A G U G G C A A G G G C C A C G G U C G C A G G A C C G U C G U
Figure imgf000259_0001
UGCUGAAGCAAGAAUGGAACCUGUGCCUGGACUGGCAUGGCUGGCA UCCACAGUGAAUCUGGAAUUUCAGCAUGCUUCUCCUCAGCAUGCUGG ACUGUACCUGUGUGUGGUGUAUGUGGACGAUCACAUCCAUGCUUGG U G G G C A G C A G U G G C A A G G G C C A C G C C A C U A U A G G U C U A G C U
Figure imgf000260_0001
UCACCUGUCAGCUGACCUGGCACAGAGACUCCGUAAGCUUCAGCCGU AGAAACGCCUCUGGAACCGCCAGUGUGUUGCCGAGGCCGACUAUCAC GAUGGAAUUCACAGGCGAUCAUGCCGUCUGUACUGCCGGCUGUGUG C G G U U C G C A G G A U G G C A A G A C G U C G U U C G C C G U A G G G U C U G C
Figure imgf000261_0001
GACACGGUCAUCCACGCCGACGCCAACGCCGCCAUGUUCGCGGGCCU GGGCGCGUUCUUCGAGGGGAUGGGCGACCUGGGGCGCGCGGUCGGC AAGGUGGUGAUGGGCAUCGUGGGCGGCGUGGUAUCGGCCGUGUCGG U U A C C A U C G U C A G A U A G C A A A A C G C G C A C A U C A A C C A U C A C
Figure imgf000262_0001
UUCGGCGGAGGCUACGUGUACUUCGAGGAAUACGCCUACAGCCACCA GCUGAGCAGAGCCGAUAUCACAACCGUGUCCACCUUCAUCGACCUGA ACAUCACCAUGCUGGAAGAUCACGAGUUCGUGCCCCUGGAAGUGUAC U A C G C C G A G U C G C A A G C A U G G C A A U G A C C A U A A A C G C A C A C
Figure imgf000263_0001
UGACAACGUUAUCGUACAGAAUAGCAUGAGGAUCAGCUCUCGUCCCG GAGCCUGCUAUAGCAGACCACUGGUGUCCUUCCGCUACGAGGAUCAG GGUCCCCUCGUGGAGGGACAGCUGGGCGAGAAUAAUGAGCUGAGGC C A C A A U G U U G C U G U G U C G U A A G U G G A A G A A U A G A G C G U G A
Figure imgf000264_0001
UUGAAUUUGCAAGACUCCAGUUUACCUACAAUCACAUCCAGAGACAU GUGAAUGACAUGCUGGGCAGAGUGGCAAUUGCUUGGUGUGAACUCC AGAAUCAUGAACUGACCCUGUGGAAUGAAGCAAGAAAACUGAAUCCA C G G U U C G A C A G G U C G A G C G C G C C C C C U G C C A U A C A A C U G U G G
Figure imgf000265_0001
CGCAAGUACAACGCCACGCACAUCAAGGUGGGCCAGCCGCAGUACUA CCUGGCCACGGGGGGCUUCCUCAUCGCGUACCAGCCCCUCCUCAGCA ACACGCUCGCCGAGCUGUACGUGCGGGAGUACAUGCGGGAGCAGGAC G U C G G C C U C C C C G C A C G A G U G A G A C G C A C C U C C C U G C U A A
Figure imgf000266_0001
CCGUGGCCUGGGAUUGGGUGCCCAAAAGACCUGCCGUGUGCACCAUG ACCAAGUGGCAAGAGGUGGACGAGAUGCUGAGAGCCGAGUACGGCG GCAGCUUCAGAUUCAGCUCUGACGCCAUCAGCACCACCUUCACCACCA C A C G C C C C G G G G G G A A A A A A C C G A A A G G A G C G C U A U A A G U G
Figure imgf000267_0001
UUCGUGCUCGCUACCGGAGAUUUCGUUUAUAUGAGUCCCUUCUAUG GAUAUAGGGAAGGUUCACACACAGAACACACCUCCUACGCGGCAGAC CGCUUUAAACAGGUCGAUGGAUUUUACGCUCGGGAUCUGACCACUAA A C A A C C C G G U A U U U G C C C C C G U A U G A A G G C G G G C G U C A G G
Figure imgf000268_0001
AUCAACAGCAAAAUAUGUGAGAAACAACAUGGAAACCACAGCUUUUCA CAGAGAUGAUCAUGAAACCGAUAUGGAACUGAAACCUGCAAAAGUGG CAACCAGAACCAGCAGAGGAUGGCACACCACAGAUCUGAAAUACAAUC C G A A A U C U A G C A C A G G A A U G U C G A G U U G C A A A C A C C C U
Figure imgf000269_0001
UAGGGUGGCGGUGGUGAACGAGAGCCUCGUCAUCUACGGUGCUCUG GAAACCGACUCAGGACUGUAUACGCUCAGCGUUGUUGGCCUCUCCGA UGAGGCUCGACAGGUUGCCUCCGUAGUGCUGGUCGUAGAACCAGCCC G A G A G C C C U G U G C A C C A A A G U A G A G G A G C C C G U C G U U A C
Figure imgf000270_0001
CAACGGCAGCCAGAACCAGUGGAGCCCACUCGGCCUCAUGUGCGAGC CUGAUAA G U A G A G A G C C C G U C G U U A C C G G C G U C A G G A G C C G A A A U
Figure imgf000271_0001
GGAGGAUCUUUCAGAUUUUCUUCUGAUGCAAUCUCAACCACCUUCAC CACCAAUCUGACAGAAUACUCUCUGUCUAGAGUGGAUCUGGGAGAUU GCAUUGGGAGAGAUGCAAGAGAAGCAAUUGAUAGAAUGUUUGCAAGA UG C AA A C A A G A G A GA U G GU C AG A UG U U C G
Figure imgf000272_0003
GA ACCAUAACAGUAGGGCCACGGGGGAAUGCUUCCAAUGCUGCACCUUC CC C C G CA CC GA U AG CA C C U C C G C C C G G AC G GU GA
Figure imgf000272_0002
Figure imgf000272_0001
Figure imgf000272_0004
GCGACAGGUGAUCAGAGCCGUGGAAGGAGCAGGCAUUGGGGUUGCU GUGCUGGUUGCGGUCGUACUCGCUGGUACAGCCGUGGUCUACCUGA CCCACGCCAGCUCUGUGCGGUAUCGUCGCCUUAGAUGAUAA A C C U G U A A A A G C C U A A C U C
Figure imgf000273_0002
[0473] In some embodiments, a polyribonucleotide encodes a T-cell string polypeptide, wherein the T-cell string polypeptide comprises (i) secretory signal, (ii) one or more HSV T-cell antigenic portions, (iii) linkers, optionally a MITD and/or transmembrane region. [0474] Select example payloads in accordance with this disclosure encode polypeptides illustrated in FIG. 18 For example in some embodiments a pol ribonucleotide encoding a T-cell string provide herein can include, in ). ng DP DQ PD FA AHA
Figure imgf000273_0001
RLYPDAPPLRLCRGANVRYRVRTRFGPDTLVPMSPREYRRAVLPALDGRAAASGGSGGGGSGGETLVAHGPSLYRTFAANPRAASTA KAMRDCVLRQENLIEALASADETLAWCKMCIHHNLPLRPQDPIIGTAAAVLENLATRLRPFLQCYLKARGLCGLDDLCSRRRLSDIKD IASFVLVILARLANRVERGVSEIDYTTVGVGAGETMHFYIPGACMAGLIEILDTHRQECSSRVCELTASHTIAPLYVHGKYFYCNSLFG GSGGGGSGGIVGIVAGLAVLAVVVIGAVVATVMCRRKSSGGKGGSYSQAASSDSAQGSDVSLTA (SEQ ID NO: 595). In some embodiments, a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 595. [0475] In some embodiments, a polyribonucleotide encodes a T-cell string that comprises amino acids 26- 693 of amino acid sequence SEQ ID NO: 595. [0476] In some embodiments, a polyribonucleotide encoding a T-cell string provide herein can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an UL29 polypeptide or antigenic ). ng GSR ASL
Figure imgf000274_0001
FKPL VRRSARLYRILGVLVHLRIRTREASFEEWMRSKEVDLDFGLTERLREHEAQLMILAQALNPYDCLIHSTPNTLVERGLQSALKYEEFYL KRFGGHYMESVFQMYTRIAGFLAGGSGGGGSGGKMTRGAPKASATPATDPARGRRPAQADSAVLLDAPAPTASGRTKTPAQGLAK KLHFSTAPPSPTAPWTPRVAGFNKRVFCAAVGGGSGGGGSGGLLNNYDVLVLDEVMSTLGQLYSPTMQQLGRVDALMLRLLRTCPR IIAMDATANAQLVDFLCSLRGEKNVHVVIGEYAMPGFSARRCLFLPRLGPEVLQAALRRRGPAGGAPPPDAPPDATFFGELEARLAGG DNVCIFSSTVSFAEVVARFCRQFTDRVLLLHSLTPPGDVTTWGRYRVVIYTTVVTVGLSFDPPHFDSMFAYVKPMNYGPDMVSVYQS LGRVRTLRKGELLIYMDGSGARSEPVGGSGGGGSGGIVGIVAGLAVLAVVVIGAVVATVMCRRKSSGGKGGSYSQAASSDSAQGSD VSLTA (SEQ ID NO: 596). In some embodiments, a polyribonucleotide encodes a polypeptide having an amino acid 26- can YG RAY FL
Figure imgf000274_0002
FAFLSAADDLVTENLGGLSGLFEQKDILHYYVEQECIEVVHSRVYNIIQLVLFHNNDQARRAYVARTINHPAIRVKVDWLEARVRECD SIPEKFILMILIEGVFFAASFAAIAYLRTNNLLRGGSGGGGSGGHEFGNLMKVLEYGLPITEEHMQFVDRFVVPESYITNPANLPGWTR LFSSHKEVSAYMAKLHAYLKVTREGEFVVFTLPVLTFVSVKEFDEYRRLGGSGGGGSGGELFGEVFESAPFSTYVDNVIFRGCELLTGS PRGGLMSVALQTDNYTLMGYTYTRVFAFAEELRRRHATAGVAEFLEESPLPYIVLRDQHGFMSVVNTNIGGSGGGGSGGSVAAPVE VTALYATDGCVITSSLALLTNCLLGAEPLYIFSYDAYRSDAPNGPTGAPTEQERFEGSRALYRDAGGLNGDSFRVTFCLLGTEVGVTH HPKGRTRPMFVCRFERADDVAVLQDALGRGTPLLPAHVTATLDLEATFALHANIIMALTVAIVHNAPARIGSGSTAPLYEPGESMRSV VGGSGGGGSGGIVGIVAGLAVLAVVVIGAVVATVMCRRKSSGGKGGSYSQAASSDSAQGSDVSLTA (SEQ ID NO: 597). In some embodiments, a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 597. [0479] In some embodiments, a polyribonucleotide encodes a T-cell string that comprises amino acids 26- 792 of amino acid sequence SEQ ID NO: 597. [0480] In some embodiments, a polyribonucleotide encoding a T-cell string polypeptide provide herein can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an UL1 polypeptide or antigenic fragment thereof, a linker, an UL19 polypeptide or antigenic fragment thereof, a linker, an UL21 polypeptide or antigenic fragment thereof, a linker, a UL27 polypeptide or fragment thereof, a linker, a UL27 polypeptide or fragment thereof, a linker, a UL46 polypeptide or fragment thereof, a linker, a UL47 polypeptide or fragment thereof, a linker, a UL25 polypeptide or fragment thereof, a linker, a UL48 polypeptide or fragment thereof, a linker, and a MITD (see FIG.18D). In some embodiments, such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGRTPADDVSWRYEAPSVIDYARIDGIFLRYHCPGLDTFLWDRHAQRAYLVNPFLFAAGFLEDL SHSVFPADTQETTGGSGGGGSGGDGRLLHNTQARAADAADDRPHRPADWTVHHKIYYYVLVPAFSRGRCCTAGVRFDRVYATLQ NMVVPEIAPGEECPSDPVTDPAHPLHPANLVANTVKRMFHNGGSGGGGSGGSPTQKLAVYYYLIHRERRMSPFPALVRLVGRYIQR HGLYVPAPDEPTLADAMNGLGGSGGGGSGGNYTEGIAVVFKENIAPYKFKATMYYKDVTVSQVWFGHRYSQFMGIFEDRAPVPFEE VGGSGGGGSGGSVYPYDEFVLATGDFVYMSPFYGYREGSHGGSGGGGSGGGLASDPHYDYIRHYASAAKQALGEVELSGGQLSRAI LAQYWKYLQTVVPSGLDIPDDPAGDCDPSLHVLLRPTLLPKLLVRAPFKSGAAAAKYAAAVAGLRDAAHRLQQYMFFMRPADPSRPS TDTALRLSELLAYVSVLYHWASWMLWTADKYVGGSGGGGSGGGPDAAVFRSSLGSLLYWPGVRALLGRDCRVAARYAGRMTYIAT GALLARFNPGAVKCVLPREAAFAGRVLGGSGGGGSGGFLWEDQTLLRATANTITALAVLRRLLANGNVYADRLDNRLQLGMLIPGAV PAEAIARGASGLDSGAIKSGDNNLEALCVNYVLPLYQADPTVELTQLFPGLAALCLGGSGGGGSGGALFNRLLDDLGFSAGPALCTML DTWNEDLFSGFPTNADMYRECKFLSTLPSDVIDWGDAHVPERSPIDIRAHGDVAFPTLPATRDELPSYYEAMAQFFRGELRAGGSG GGGSGGIVGIVAGLAVLAVVVIGAVVATVMCRRKSSGGKGGSYSQAASSDSAQGSDVSLTA (SEQ ID NO: 598). In some embodiments, a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 598. [0481] In some embodiments, a polyribonucleotide encodes a T-cell string that comprises amino acids 26- 858 of amino acid sequence SEQ ID NO: 598. [0482] In some embodiments, a polyribonucleotide encoding a T-cell string polypeptide provide herein can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an UL54 polypeptide or antigenic fragment thereof, a linker, an RS1 polypeptide or antigenic fragment thereof, a linker, an RL2 polypeptide or antigenic fragment thereof, a linker, a RL2 polypeptide or fragment thereof, a linker, and a MITD. In some embodiments, such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGETLVAHGPSLYRTFAANPRAASTAKAMRDCVLRQENLIEALASADETLAWCKMCIHHNLPLR PQDPIIGTAAAVLENLATRLRPFLQCYLKARGLCGLDDLCSRRRLSDIKDIASFVLVILARLANRVERGVSEIDYTTVGVGAGETMHFYI PGACMAGLIEILDTHRQECSSRVCELTASHTIAPLYVHGKYFYCNSLFGGSGGGGSGGRAAAWMRQVPDPEDVRVVILYSPLPGEDL AAGRAGGGPPPEWSAERGGLSCLLAALGNRLCGPATAAWAGNWTGAPDVSALGAQGVLLLSTRDLAFAGAVEFLGLLAGACDRRLI VVNAVRAADWPADGPVVSRQHAYLACEVLPAVQCAVRWPAARDLRRTVLASGRVFGPGVFARVEAAHARLYPDAPPLRLCRGANV RYRVRTRFGPDTLVPMSPREYRRAVLPALDGRAAASGGSGGGGSGGCTDEIAPPLRCQSFPCLHPFCIPCMKTWIPLRNTCPLCNTP VAYLIVGVTASGSFSTIPIVNDPRTRVEAEAAVRAGTAVDFIWTGNPRTAPRSLSGGSGGGGSGGLPIAGVSSVVALAPYVNKTVTGD CLPVLDMETGHIGAYVVLVDQTGNVADLLRAAAPAWSRRTLLPEHARNCVRPPDYPTPPASEWNSLWMTPVGNMLFDQGTLVGGG SGGGGSGGIVGIVAGLAVLAVVVIGAVVATVMCRRKSSGGKGGSYSQAASSDSAQGSDVSLTA (SEQ ID NO: 599). In some embodiments, a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 599. [0483] In some embodiments, a polyribonucleotide encodes a T-cell string that comprises amino acids 26- 692 of amino acid sequence SEQ ID NO: 599. [0484] In some embodiments, a polyribonucleotide encoding a T-cell string polypeptide provide herein can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an UL9 polypeptide or antigenic fragment thereof, a linker, an UL49 polypeptide or antigenic fragment thereof, a linker, an UL39 polypeptide or antigenic fragment thereof, a linker, a UL29 polypeptide or fragment thereof, a linker, and a MITD. In some embodiments, such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGLLNNYDVLVLDEVMSTLGQLYSPTMQQLGRVDALMLRLLRTCPRIIAMDATANAQLVDFLCS LRGEKNVHVVIGEYAMPGFSARRCLFLPRLGPEVLQAALRRRGPAGGAPPPDAPPDATFFGELEARLAGGDNVCIFSSTVSFAEVVAR FCRQFTDRVLLLHSLTPPGDVTTWGRYRVVIYTTVVTVGLSFDPPHFDSMFAYVKPMNYGPDMVSVYQSLGRVRTLRKGELLIYMD GSGARSEPVGGSGGGGSGGKMTRGAPKASATPATDPARGRRPAQADSAVLLDAPAPTASGRTKTPAQGLAKKLHFSTAPPSPTAPW TPRVAGFNKRVFCAAVGGGSGGGGSGGRTFGSAPRLTEDDFGLLNYALAEMRRLCLDLPPVPPNAYTPYHLREYATRLVNGFKPLVR RSARLYRILGVLVHLRIRTREASFEEWMRSKEVDLDFGLTERLREHEAQLMILAQALNPYDCLIHSTPNTLVERGLQSALKYEEFYLKR FGGHYMESVFQMYTRIAGFLAGGSGGGGSGGREDIETIAFIKRFSLDYGAINFINLAPNNVSELAMYYMANQILRYCDHSTYFINTLT AVIAGSRRPPSVQAAAAWAPQGGAGLEAGARALMDSLDAHPGAWTSMFASCNLLRPVMAARPMVVLGLSISKYYGMAGNDRVFQA GNWASLLGGKNACPLLIFDRTRKFVLGGSGGGGSGGIVGIVAGLAVLAVVVIGAVVATVMCRRKSSGGKGGSYSQAASSDSAQGSD VSLTA (SEQ ID NO: 600). In some embodiments, a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 600. [0485] In some embodiments, a polyribonucleotide encodes a T-cell string that comprises amino acids 26- 722 of amino acid sequence SEQ ID NO: 600. [0486] In some embodiments, a polyribonucleotide encoding a T-cell string polypeptide provide herein can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an UL52 polypeptide or antigenic fragment thereof, a linker, an UL5 polypeptide or antigenic fragment thereof, a linker, an UL5 polypeptide or antigenic fragment thereof, a linker, a UL40 polypeptide or fragment thereof, a linker, a UL30 polypeptide or fragment thereof, a linker, a UL30 polypeptide or fragment thereof, a linker, and a MITD. In some embodiments, such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGSVAAPVEVTALYATDGCVITSSLALLTNCLLGAEPLYIFSYDAYRSDAPNGPTGAPTEQERFEG SRALYRDAGGLNGDSFRVTFCLLGTEVGVTHHPKGRTRPMFVCRFERADDVAVLQDALGRGTPLLPAHVTATLDLEATFALHANIIM ALTVAIVHNAPARIGSGSTAPLYEPGESMRSVVGGSGGGGSGGHEFGNLMKVLEYGLPITEEHMQFVDRFVVPESYITNPANLPGWT RLFSSHKEVSAYMAKLHAYLKVTREGEFVVFTLPVLTFVSVKEFDEYRRLGGSGGGGSGGELFGEVFESAPFSTYVDNVIFRGCELLTG SPRGGLMSVALQTDNYTLMGYTYTRVFAFAEELRRRHATAGVAEFLEESPLPYIVLRDQHGFMSVVNTNIGGSGGGGSGGTSQCPDI NHLRSLSILNRWLETELVFVGDEEDVSKLSEGELGFYRFLFAFLSAADDLVTENLGGLSGLFEQKDILHYYVEQECIEVVHSRVYNIIQL VLFHNNDQARRAYVARTINHPAIRVKVDWLEARVRECDSIPEKFILMILIEGVFFAASFAAIAYLRTNNLLRGGSGGGGSGGISCLLYD LSTTALEHILLFSLGSCDLPESHLSDLASRGLPAPVVLEFDSEFEMLLAFMTFVKQYGPEFVTGYNIINFDWPFVLTKLTEIYKVPLDGY GRMNGRGVFRVWDIGQSHFGGSGGGGSGGGLLPCLHVAATVTTIGREMLLATRAYVHARWAEFDQLLADFPEAAGMRAPGPYSM GGSGGGGSGGIVGIVAGLAVLAVVVIGAVVATVMCRRKSSGGKGGSYSQAASSDSAQGSDVSLTA (SEQ ID NO: 601). In some embodiments, a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 601. [0487] In some embodiments, a polyribonucleotide encodes a T-cell string that comprises amino acids 26- 792 of amino acid sequence SEQ ID NO: 601. [0488] In some embodiments, a polyribonucleotide encoding a T-cell string polypeptide provide herein can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an UL48 polypeptide or antigenic fragment thereof, a linker, an UL25 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, a UL46 polypeptide or fragment thereof, a linker, a UL27 polypeptide or fragment thereof, a linker, a UL27 polypeptide or fragment thereof, a linker, a UL21 polypeptide or fragment thereof, a linker, a UL19 polypeptide or fragment thereof, a linker, a UL1 polypeptide or fragment thereof, a linker, and a MITD. In some embodiments, such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGALFNRLLDDLGFSAGPALCTMLDTWNEDLFSGFPTNADMYRECKFLSTLPSDVIDWGDAHV PERSPIDIRAHGDVAFPTLPATRDELPSYYEAMAQFFRGELRAGGSGGGGSGGFLWEDQTLLRATANTITALAVLRRLLANGNVYAD RLDNRLQLGMLIPGAVPAEAIARGASGLDSGAIKSGDNNLEALCVNYVLPLYQADPTVELTQLFPGLAALCLGGSGGGGSGGGPDAA VFRSSLGSLLYWPGVRALLGRDCRVAARYAGRMTYIATGALLARFNPGAVKCVLPREAAFAGRVLGGSGGGGSGGGLASDPHYDYIR HYASAAKQALGEVELSGGQLSRAILAQYWKYLQTVVPSGLDIPDDPAGDCDPSLHVLLRPTLLPKLLVRAPFKSGAAAAKYAAAVAGL RDAAHRLQQYMFFMRPADPSRPSTDTALRLSELLAYVSVLYHWASWMLWTADKYVGGSGGGGSGGNYTEGIAVVFKENIAPYKFK ATMYYKDVTVSQVWFGHRYSQFMGIFEDRAPVPFEEVGGSGGGGSGGSVYPYDEFVLATGDFVYMSPFYGYREGSHGGSGGGGSG GSPTQKLAVYYYLIHRERRMSPFPALVRLVGRYIQRHGLYVPAPDEPTLADAMNGLGGSGGGGSGGDGRLLHNTQARAADAADDRP HRPADWTVHHKIYYYVLVPAFSRGRCCTAGVRFDRVYATLQNMVVPEIAPGEECPSDPVTDPAHPLHPANLVANTVKRMFHNGGSG GGGSGGRTPADDVSWRYEAPSVIDYARIDGIFLRYHCPGLDTFLWDRHAQRAYLVNPFLFAAGFLEDLSHSVFPADTQETTGGSGG GGSGGIVGIVAGLAVLAVVVIGAVVATVMCRRKSSGGKGGSYSQAASSDSAQGSDVSLTA (SEQ ID NO: 602). In some embodiments, a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 602. [0489] In some embodiments, a polyribonucleotide encodes a T-cell string that comprises amino acids 26- 858 of amino acid sequence SEQ ID NO: 602. [0490] In some embodiments, a polyribonucleotide encoding a T-cell string polypeptide provide herein can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-2 gD secretory signal, an RL2 polypeptide or antigenic fragment thereof, a linker, an RL2 polypeptide or antigenic fragment thereof, a linker, an RS1 polypeptide or antigenic fragment thereof, a linker, a UL54 polypeptide or fragment thereof, a linker, and a MITD. In some embodiments, such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGRLTSGVGTAALLVVAVGLRVVCACTDEIAPPLRCQSFPCLHPFCIPCMKTWIPLRNTCPLCNTPVAYLIVGVTASGSFSTIPIVNDP RTRVEAEAAVRAGTAVDFIWTGNPRTAPRSLSGGSGGGGSGGLPIAGVSSVVALAPYVNKTVTGDCLPVLDMETGHIGAYVVLVDQ TGNVADLLRAAAPAWSRRTLLPEHARNCVRPPDYPTPPASEWNSLWMTPVGNMLFDQGTLVGGGSGGGGSGGRAAAWMRQVPD PEDVRVVILYSPLPGEDLAAGRAGGGPPPEWSAERGGLSCLLAALGNRLCGPATAAWAGNWTGAPDVSALGAQGVLLLSTRDLAFA GAVEFLGLLAGACDRRLIVVNAVRAADWPADGPVVSRQHAYLACEVLPAVQCAVRWPAARDLRRTVLASGRVFGPGVFARVEAAHA RLYPDAPPLRLCRGANVRYRVRTRFGPDTLVPMSPREYRRAVLPALDGRAAASGGSGGGGSGGETLVAHGPSLYRTFAANPRAASTA KAMRDCVLRQENLIEALASADETLAWCKMCIHHNLPLRPQDPIIGTAAAVLENLATRLRPFLQCYLKARGLCGLDDLCSRRRLSDIKD IASFVLVILARLANRVERGVSEIDYTTVGVGAGETMHFYIPGACMAGLIEILDTHRQECSSRVCELTASHTIAPLYVHGKYFYCNSLFG GSGGGGSGGIVGIVAGLAVLAVVVIGAVVATVMCRRKSSGGKGGSYSQAASSDSAQGSDVSLTA (SEQ ID NO: 603). In some embodiments, a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 603. [0491] In some embodiments, a polyribonucleotide encodes a T-cell string that comprises amino acids 26- 693 of amino acid sequence SEQ ID NO: 603. [0492] In some embodiments, a polyribonucleotide encoding a T-cell string polypeptide provide herein can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-2 gD secretory signal, an UL54 polypeptide or antigenic fragment thereof, a linker, an RS1 polypeptide or antigenic fragment thereof, a linker, an RL2 polypeptide or antigenic fragment thereof, a linker, a RL2 polypeptide or fragment thereof, a linker, and a MITD. In some embodiments, such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGRLTSGVGTAALLVVAVGLRVVCAETLVAHGPSLYRTFAANPRAASTAKAMRDCVLRQENLIEALASADETLAWCKMCIHHNLPLR PQDPIIGTAAAVLENLATRLRPFLQCYLKARGLCGLDDLCSRRRLSDIKDIASFVLVILARLANRVERGVSEIDYTTVGVGAGETMHFYI PGACMAGLIEILDTHRQECSSRVCELTASHTIAPLYVHGKYFYCNSLFGGSGGGGSGGRAAAWMRQVPDPEDVRVVILYSPLPGEDL AAGRAGGGPPPEWSAERGGLSCLLAALGNRLCGPATAAWAGNWTGAPDVSALGAQGVLLLSTRDLAFAGAVEFLGLLAGACDRRLI VVNAVRAADWPADGPVVSRQHAYLACEVLPAVQCAVRWPAARDLRRTVLASGRVFGPGVFARVEAAHARLYPDAPPLRLCRGANV RYRVRTRFGPDTLVPMSPREYRRAVLPALDGRAAASGGSGGGGSGGCTDEIAPPLRCQSFPCLHPFCIPCMKTWIPLRNTCPLCNTP VAYLIVGVTASGSFSTIPIVNDPRTRVEAEAAVRAGTAVDFIWTGNPRTAPRSLSGGSGGGGSGGLPIAGVSSVVALAPYVNKTVTGD CLPVLDMETGHIGAYVVLVDQTGNVADLLRAAAPAWSRRTLLPEHARNCVRPPDYPTPPASEWNSLWMTPVGNMLFDQGTLVGGG SGGGGSGGIVGIVAGLAVLAVVVIGAVVATVMCRRKSSGGKGGSYSQAASSDSAQGSDVSLTA (SEQ ID NO: 604). In some embodiments, a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 604. [0493] In some embodiments, a polyribonucleotide encodes a T-cell string that comprises amino acids 26- 692 of amino acid sequence SEQ ID NO: 604. [0494] In some embodiments, a polyribonucleotide encoding a T-cell string polypeptide provide herein can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an US1 polypeptide or antigenic fragment thereof, a linker, an US1 polypeptide or antigenic fragment thereof, a linker, an US8 polypeptide or antigenic fragment thereof, a linker, a US8 polypeptide or fragment thereof, a linker, a US8 polypeptide or fragment thereof, a linker, a US12 polypeptide or fragment thereof, a linker, a UL50 polypeptide or fragment thereof, a linker, a UL26 polypeptide or fragment thereof, a linker, a UL26 polypeptide or fragment thereof, a linker, a US10 polypeptide or fragment thereof, a linker, and a MITD. In some embodiments, such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGDDASDGWLVDTPPRKSKRPRINLRLTSSPDRRAGVVFPEVGGSGGGGSGGPASLPGIAHAH RRSARQAQMRSGAAWTLDLHYIRQCVNQLGGSGGGGSGGILSPTAPSVYPHSEGRKSRRPLTTFGSGSPGRRHSQASYPSVLWGG SGGGGSGGGLAWLASTVNLEFQHASPQHAGLYLCVVYVDDHIHAWGHMTISTAAQYRNAVVEQHLPQRQPEPVEPTRPHVRAPHP APSARGPLRLGGSGGGGSGGKLLWAAEPLDACGPLRPSWVALWPPRRVLETVVDAACMRAPEPLAIAYSPPFPAGDEGLYSELAWR DRVAVVNESLVIYGALETDSGLYTLSVVGLSDEARQVASVVLVVEPAPGGSGGGGSGGEDREAARTAVTDPELPLLCPPDVGGSGGG GSGGANGATVIQPSLRVLRAADGPEACYVLGRSSLNARGLLVMPTRWPSGHACAFVVCNLTGVPVTLGGSGGGGSGGAPLPDRAVP IYVAGFLALYDSGDPGELALDPDTVRAALPPENPLPINVDHRARCEVGRVLAVVNDPRGPFFVGLIACVQLERVLETAASAAIFERRGP ALSREERLLYLITNYLPSVSLSTKRRGDEVPPDRTLFAHVALCAIGRRLGTIVTYDTSLDAAGGSGGGGSGGHYPPPPAHPYPGMLFA GPSPLEAQIAALVGAIAADRQAGGLPAAAGDHGIRGSAKRRRHEVEQPEYDCGGGSGGGGSGGSSPRQRTYVLPRVGIHNAPASDT RAPKRANSRHRADRPPESPGSELYPLNAQALAHLQMLPADHRAFFRTVIEVSRLCALNTHDPPPPLAGARVGQEAQLVHTQWLRAN RESSPLWPWRTAAMNFIAAAAPCVQTHRHMHDLLMACAFWCGGSGGGGSGGIVGIVAGLAVLAVVVIGAVVATVMCRRKSSGGKG GSYSQAASSDSAQGSDVSLTA (SEQ ID NO: 605). In some embodiments, a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 605. [0495] In some embodiments, a polyribonucleotide encodes a T-cell string that comprises amino acids 26- 905 of amino acid sequence SEQ ID NO: 605. [0496] In some embodiments, a polyribonucleotide encoding a T-cell string polypeptide provide herein can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an UL26 polypeptide or antigenic fragment thereof, a linker, an UL26 polypeptide or antigenic fragment thereof, a linker, an US10 polypeptide or antigenic fragment thereof, a linker, a UL50 polypeptide or fragment thereof, a linker, a US12 polypeptide or fragment thereof, a linker, a US8 polypeptide or fragment thereof, a linker, a US8 polypeptide or fragment thereof, a linker, a US8 polypeptide or fragment thereof, a linker, a US1 polypeptide or fragment thereof, a linker, a US1 polypeptide or fragment thereof, a linker, and a MITD. In some embodiments, such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGHYPPPPAHPYPGMLFAGPSPLEAQIAALVGAIAADRQAGGLPAAAGDHGIRGSAKRRRHEVE QPEYDCGGGSGGGGSGGAPLPDRAVPIYVAGFLALYDSGDPGELALDPDTVRAALPPENPLPINVDHRARCEVGRVLAVVNDPRGPF FVGLIACVQLERVLETAASAAIFERRGPALSREERLLYLITNYLPSVSLSTKRRGDEVPPDRTLFAHVALCAIGRRLGTIVTYDTSLDAA GGSGGGGSGGSSPRQRTYVLPRVGIHNAPASDTRAPKRANSRHRADRPPESPGSELYPLNAQALAHLQMLPADHRAFFRTVIEVSRL CALNTHDPPPPLAGARVGQEAQLVHTQWLRANRESSPLWPWRTAAMNFIAAAAPCVQTHRHMHDLLMACAFWCGGSGGGGSGG ANGATVIQPSLRVLRAADGPEACYVLGRSSLNARGLLVMPTRWPSGHACAFVVCNLTGVPVTLGGSGGGGSGGEDREAARTAVTDP ELPLLCPPDVGGSGGGGSGGKLLWAAEPLDACGPLRPSWVALWPPRRVLETVVDAACMRAPEPLAIAYSPPFPAGDEGLYSELAWR DRVAVVNESLVIYGALETDSGLYTLSVVGLSDEARQVASVVLVVEPAPGGSGGGGSGGGLAWLASTVNLEFQHASPQHAGLYLCVVY VDDHIHAWGHMTISTAAQYRNAVVEQHLPQRQPEPVEPTRPHVRAPHPAPSARGPLRLGGSGGGGSGGILSPTAPSVYPHSEGRKS RRPLTTFGSGSPGRRHSQASYPSVLWGGSGGGGSGGPASLPGIAHAHRRSARQAQMRSGAAWTLDLHYIRQCVNQLGGSGGGGS GGDDASDGWLVDTPPRKSKRPRINLRLTSSPDRRAGVVFPEVGGSGGGGSGGIVGIVAGLAVLAVVVIGAVVATVMCRRKSSGGKG GSYSQAASSDSAQGSDVSLTA (SEQ ID NO: 606). In some embodiments, a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 606. [0497] In some embodiments, a polyribonucleotide encodes a T-cell string that comprises amino acids 26- 905 of amino acid sequence SEQ ID NO: 606. [0498] In some embodiments, a polyribonucleotide encoding a T-cell string provide herein can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an RL2 polypeptide or antigenic fragment thereof, a linker, an RL2 polypeptide or antigenic fragment thereof, a linker, an RS1 polypeptide or antigenic fragment thereof, a linker, a UL54 polypeptide or fragment thereof, a linker, and an HSV-1 transmembrane region. In some embodiments, such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGCTDEIAPPLRCQSFPCLHPFCIPCMKTWIPLRNTCPLCNTPVAYLIVGVTASGSFSTIPIVNDP RTRVEAEAAVRAGTAVDFIWTGNPRTAPRSLSGGSGGGGSGGLPIAGVSSVVALAPYVNKTVTGDCLPVLDMETGHIGAYVVLVDQ TGNVADLLRAAAPAWSRRTLLPEHARNCVRPPDYPTPPASEWNSLWMTPVGNMLFDQGTLVGGGSGGGGSGGRAAAWMRQVPD PEDVRVVILYSPLPGEDLAAGRAGGGPPPEWSAERGGLSCLLAALGNRLCGPATAAWAGNWTGAPDVSALGAQGVLLLSTRDLAFA GAVEFLGLLAGACDRRLIVVNAVRAADWPADGPVVSRQHAYLACEVLPAVQCAVRWPAARDLRRTVLASGRVFGPGVFARVEAAHA RLYPDAPPLRLCRGANVRYRVRTRFGPDTLVPMSPREYRRAVLPALDGRAAASGGSGGGGSGGETLVAHGPSLYRTFAANPRAASTA KAMRDCVLRQENLIEALASADETLAWCKMCIHHNLPLRPQDPIIGTAAAVLENLATRLRPFLQCYLKARGLCGLDDLCSRRRLSDIKD IASFVLVILARLANRVERGVSEIDYTTVGVGAGETMHFYIPGACMAGLIEILDTHRQECSSRVCELTASHTIAPLYVHGKYFYCNSLFG GSGGGGSGGGLIAGAVGGSLLAALVICGIVYWMRRHTQKAPKRIRLPHIR (SEQ ID NO: 607). In some embodiments, a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 607. [0499] In some embodiments, a polyribonucleotide encodes a T-cell string that comprises amino acids 26- 693 of amino acid sequence SEQ ID NO: 607. [0500] In some embodiments, a polyribonucleotide encoding a T-cell string polypeptide provide herein can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an RL2 polypeptide or antigenic fragment thereof, a linker, an RL2 polypeptide or antigenic fragment thereof, a linker, an RS1 polypeptide or antigenic fragment thereof, a linker, a UL54 polypeptide or fragment thereof, a linker, and a VSV-G transmembrane region. In some embodiments, such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGCTDEIAPPLRCQSFPCLHPFCIPCMKTWIPLRNTCPLCNTPVAYLIVGVTASGSFSTIPIVNDP RTRVEAEAAVRAGTAVDFIWTGNPRTAPRSLSGGSGGGGSGGLPIAGVSSVVALAPYVNKTVTGDCLPVLDMETGHIGAYVVLVDQ TGNVADLLRAAAPAWSRRTLLPEHARNCVRPPDYPTPPASEWNSLWMTPVGNMLFDQGTLVGGGSGGGGSGGRAAAWMRQVPD PEDVRVVILYSPLPGEDLAAGRAGGGPPPEWSAERGGLSCLLAALGNRLCGPATAAWAGNWTGAPDVSALGAQGVLLLSTRDLAFA GAVEFLGLLAGACDRRLIVVNAVRAADWPADGPVVSRQHAYLACEVLPAVQCAVRWPAARDLRRTVLASGRVFGPGVFARVEAAHA RLYPDAPPLRLCRGANVRYRVRTRFGPDTLVPMSPREYRRAVLPALDGRAAASGGSGGGGSGGETLVAHGPSLYRTFAANPRAASTA KAMRDCVLRQENLIEALASADETLAWCKMCIHHNLPLRPQDPIIGTAAAVLENLATRLRPFLQCYLKARGLCGLDDLCSRRRLSDIKD IASFVLVILARLANRVERGVSEIDYTTVGVGAGETMHFYIPGACMAGLIEILDTHRQECSSRVCELTASHTIAPLYVHGKYFYCNSLFG GSGGGGSGGIASFFFIIGLIIGLFLVLRVGIYLCIKLKHTKKRQIYTDIEMN (SEQ ID NO: 608). In some embodiments, a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 608. [0501] In some embodiments, a polyribonucleotide encodes a T-cell string that comprises amino acids 26- 693 of amino acid sequence SEQ ID NO: 608. 5’
Figure imgf000281_0001
to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an RL2 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, a UL46 polypeptide or antigenic fragment thereof, a linker, a UL21 polypeptide or antigenic fragment thereof, a linker, and a MITD. In some embodiments, such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGCTDEIAPPLRCQSFPCLHPFCIPCMKTWIPLRNTCPLCNTPVAYLIVGVTASGSFSTIPIVNDP RTRVEAEAAVRAGTAVDFIWTGNPRTAPRSLSGGSGGGGSGGETLVAHGPSLYRTFAANPRAASTAKAMRDCVLRQENLIEALASA DETLAWCKMCIHHNLPLRPQDPIIGTAAAVLENLATRLRPFLQCYLKARGLCGLDDLCSRRRLSDIKDIASFVLVILARLANRVERGVS EIDYTTVGVGAGETMHFYIPGACMAGLIEILDTHRQECSSRVCELTASHTIAPLYVHGKYFYCNSLFGGSGGGGSGGGPDAAVFRSSL GSLLYWPGVRALLGRDCRVAARYAGRMTYIATGALLARFNPGAVKCVLPREAAFAGRVLGGSGGGGSGGGLASDPHYDYIRHYASA AKQALGEVELSGGQLSRAILAQYWKYLQTVVPSGLDIPDDPAGDCDPSLHVLLRPTLLPKLLVRAPFKSGAAAAKYAAAVAGLRDAAH RLQQYMFFMRPADPSRPSTDTALRLSELLAYVSVLYHWASWMLWTADKYVGGSGGGGSGGSPTQKLAVYYYLIHRERRMSPFPAL VRLVGRYIQRHGLYVPAPDEPTLADAMNGLGGSGGGGSGGIVGIVAGLAVLAVVVIGAVVATVMCRRKSSGGKGGSYSQAASSDSA QGSDVSLTA (SEQ ID NO: 633). In some embodiments, a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 633. [0503] In some embodiments, a polyribonucleotide encodes a polypeptide having amino acids 26-651 of amino acid sequence SEQ ID NO: 633. [0504] In some embodiments, a polyribonucleotide encoding a T-cell string polypeptide can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an UL21 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, an RL2 polypeptide or antigenic fragment thereof, a linker, and a MITD. In some embodiments, such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGSPTQKLAVYYYLIHRERRMSPFPALVRLVGRYIQRHGLYVPAPDEPTLADAMNGLGGSGGGG SGGGLASDPHYDYIRHYASAAKQALGEVELSGGQLSRAILAQYWKYLQTVVPSGLDIPDDPAGDCDPSLHVLLRPTLLPKLLVRAPFK SGAAAAKYAAAVAGLRDAAHRLQQYMFFMRPADPSRPSTDTALRLSELLAYVSVLYHWASWMLWTADKYVGGSGGGGSGGGPDA AVFRSSLGSLLYWPGVRALLGRDCRVAARYAGRMTYIATGALLARFNPGAVKCVLPREAAFAGRVLGGSGGGGSGGETLVAHGPSLY RTFAANPRAASTAKAMRDCVLRQENLIEALASADETLAWCKMCIHHNLPLRPQDPIIGTAAAVLENLATRLRPFLQCYLKARGLCGLD DLCSRRRLSDIKDIASFVLVILARLANRVERGVSEIDYTTVGVGAGETMHFYIPGACMAGLIEILDTHRQECSSRVCELTASHTIAPLYV HGKYFYCNSLFGGSGGGGSGGCTDEIAPPLRCQSFPCLHPFCIPCMKTWIPLRNTCPLCNTPVAYLIVGVTASGSFSTIPIVNDPRTRV EAEAAVRAGTAVDFIWTGNPRTAPRSLSGGSGGGGSGGIVGIVAGLAVLAVVVIGAVVATVMCRRKSSGGKGGSYSQAASSDSAQG SDVSLTA (SEQ ID NO: 634). In some embodiments, a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 634. [0505] In some embodiments, a polyribonucleotide encodes a polypeptide having amino acids 26-651 of amino acid sequence SEQ ID NO: 634. [0506] In some embodiments, a polyribonucleotide encoding a T-cell string polypeptide can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an UL29 polypeptide or antigenic fragment thereof, a linker, an UL39 polypeptide or antigenic fragment thereof, a linker, an UL9 polypeptide or antigenic fragment thereof, a linker, an UL5.1 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL30.1 polypeptide or antigenic fragment thereof, a linker, and a MITD. In some embodiments, such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGREDIETIAFIKRFSLDYGAINFINLAPNNVSELAMYYMANQILRYCDHSTYFINTLTAVIAGSR RPPSVQAAAAWAPQGGAGLEAGARALMDSLDAHPGAWTSMFASCNLLRPVMAARPMVVLGLSISKYYGMAGNDRVFQAGNWASL LGGKNACPLLIFDRTRKFVLGGSGGGGSGGRTFGSAPRLTEDDFGLLNYALAEMRRLCLDLPPVPPNAYTPYHLREYATRLVNGFKPL VRRSARLYRILGVLVHLRIRTREASFEEWMRSKEVDLDFGLTERLREHEAQLMILAQALNPYDCLIHSTPNTLVERGLQSALKYEEFYL KRFGGHYMESVFQMYTRIAGFLAGGSGGGGSGGLLNNYDVLVLDEVMSTLGQLYSPTMQQLGRVDALMLRLLRTCPRIIAMDATAN AQLVDFLCSLRGEKNVHVVIGEYAMPGFSARRCLFLPRLGPEVLQAALRRRGPAGGAPPPDAPPDATFFGELEARLAGGDNVCIFSST VSFAEVVARFCRQFTDRVLLLHSLTPPGDVTTWGRYRVVIYTTVVTVGLSFDPPHFDSMFAYVKPMNYGPDMVSVYQSLGRVRTLR KGELLIYMDGSGARSEPVGGSGGGGSGGHEFGNLMKVLEYGLPITEEHMQFVDRFVVPESYITNPANLPGWTRLFSSHKEVSAYMA KLHAYLKVTREGEFVVFTLPVLTFVSVKEFDEYRRLGGSGGGGSGGELFGEVFESAPFSTYVDNVIFRGCELLTGSPRGGLMSVALQT DNYTLMGYTYTRVFAFAEELRRRHATAGVAEFLEESPLPYIVLRDQHGFMSVVNTNIGGSGGGGSGGTSQCPDINHLRSLSILNRWL ETELVFVGDEEDVSKLSEGELGFYRFLFAFLSAADDLVTENLGGLSGLFEQKDILHYYVEQECIEVVHSRVYNIIQLVLFHNNDQARRA YVARTINHPAIRVKVDWLEARVRECDSIPEKFILMILIEGVFFAASFAAIAYLRTNNLLRGGSGGGGSGGISCLLYDLSTTALEHILLFSL GSCDLPESHLSDLASRGLPAPVVLEFDSEFEMLLAFMTFVKQYGPEFVTGYNIINFDWPFVLTKLTEIYKVPLDGYGRMNGRGVFRV WDIGQSHFGGSGGGGSGGIVGIVAGLAVLAVVVIGAVVATVMCRRKSSGGKGGSYSQAASSDSAQGSDVSLTA (SEQ ID NO: 635). In some embodiments, a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 635. [0507] In some embodiments, a polyribonucleotide encodes a polypeptide having amino acids 26-1156 of amino acid sequence SEQ ID NO: 635. [0508] In some embodiments, a polyribonucleotide encoding a T-cell string polypeptide can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an UL30.1 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL5.1 polypeptide or antigenic fragment thereof, a linker, an UL9 polypeptide or antigenic fragment thereof, a linker, an UL39 polypeptide or antigenic fragment thereof, a linker, an UL29 polypeptide or antigenic fragment thereof, a linker, and a MITD. In some embodiments, such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGTSQCPDINHLRSLSILNRWLETELVFVGDEEDVSKLSEGELGFYRFLFAFLSAADDLVTENLG GLSGLFEQKDILHYYVEQECIEVVHSRVYNIIQLVLFHNNDQARRAYVARTINHPAIRVKVDWLEARVRECDSIPEKFILMILIEGVFFA ASFAAIAYLRTNNLLRGGSGGGGSGGISCLLYDLSTTALEHILLFSLGSCDLPESHLSDLASRGLPAPVVLEFDSEFEMLLAFMTFVKQY GPEFVTGYNIINFDWPFVLTKLTEIYKVPLDGYGRMNGRGVFRVWDIGQSHFGGSGGGGSGGELFGEVFESAPFSTYVDNVIFRGCE LLTGSPRGGLMSVALQTDNYTLMGYTYTRVFAFAEELRRRHATAGVAEFLEESPLPYIVLRDQHGFMSVVNTNIGGSGGGGSGGHEF GNLMKVLEYGLPITEEHMQFVDRFVVPESYITNPANLPGWTRLFSSHKEVSAYMAKLHAYLKVTREGEFVVFTLPVLTFVSVKEFDEY RRLGGSGGGGSGGLLNNYDVLVLDEVMSTLGQLYSPTMQQLGRVDALMLRLLRTCPRIIAMDATANAQLVDFLCSLRGEKNVHVVI GEYAMPGFSARRCLFLPRLGPEVLQAALRRRGPAGGAPPPDAPPDATFFGELEARLAGGDNVCIFSSTVSFAEVVARFCRQFTDRVLL LHSLTPPGDVTTWGRYRVVIYTTVVTVGLSFDPPHFDSMFAYVKPMNYGPDMVSVYQSLGRVRTLRKGELLIYMDGSGARSEPVGG SGGGGSGGRTFGSAPRLTEDDFGLLNYALAEMRRLCLDLPPVPPNAYTPYHLREYATRLVNGFKPLVRRSARLYRILGVLVHLRIRTR EASFEEWMRSKEVDLDFGLTERLREHEAQLMILAQALNPYDCLIHSTPNTLVERGLQSALKYEEFYLKRFGGHYMESVFQMYTRIAG FLAGGSGGGGSGGREDIETIAFIKRFSLDYGAINFINLAPNNVSELAMYYMANQILRYCDHSTYFINTLTAVIAGSRRPPSVQAAAAW APQGGAGLEAGARALMDSLDAHPGAWTSMFASCNLLRPVMAARPMVVLGLSISKYYGMAGNDRVFQAGNWASLLGGKNACPLLIF DRTRKFVLGGSGGGGSGGIVGIVAGLAVLAVVVIGAVVATVMCRRKSSGGKGGSYSQAASSDSAQGSDVSLTA (SEQ ID NO: 636). In some embodiments, a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 636. [0509] In some embodiments, a polyribonucleotide encodes a polypeptide having amino acids 26-1156 of amino acid sequence SEQ ID NO: 636. [0510] In some embodiments, a polyribonucleotide encoding a T-cell string polypeptide can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an RL2.1 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, an UL9 polypeptide or antigenic fragment thereof, a linker, a UL39 polypeptide or antigenic fragment thereof, a linker, a UL5.1 polypeptide or antigenic fragment thereof, a linker, and a MITD. In some embodiments, such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGCTDEIAPPLRCQSFPCLHPFCIPCMKTWIPLRNTCPLCNTPVAYLIVGVTASGSFSTIPIVNDP RTRVEAEAAVRAGTAVDFIWTGNPRTAPRSLSGGSGGGGSGGETLVAHGPSLYRTFAANPRAASTAKAMRDCVLRQENLIEALASA DETLAWCKMCIHHNLPLRPQDPIIGTAAAVLENLATRLRPFLQCYLKARGLCGLDDLCSRRRLSDIKDIASFVLVILARLANRVERGVS EIDYTTVGVGAGETMHFYIPGACMAGLIEILDTHRQECSSRVCELTASHTIAPLYVHGKYFYCNSLFGGSGGGGSGGLLNNYDVLVLD EVMSTLGQLYSPTMQQLGRVDALMLRLLRTCPRIIAMDATANAQLVDFLCSLRGEKNVHVVIGEYAMPGFSARRCLFLPRLGPEVLQ AALRRRGPAGGAPPPDAPPDATFFGELEARLAGGDNVCIFSSTVSFAEVVARFCRQFTDRVLLLHSLTPPGDVTTWGRYRVVIYTTVV TVGLSFDPPHFDSMFAYVKPMNYGPDMVSVYQSLGRVRTLRKGELLIYMDGSGARSEPVGGSGGGGSGGRTFGSAPRLTEDDFGLL NYALAEMRRLCLDLPPVPPNAYTPYHLREYATRLVNGFKPLVRRSARLYRILGVLVHLRIRTREASFEEWMRSKEVDLDFGLTERLRE HEAQLMILAQALNPYDCLIHSTPNTLVERGLQSALKYEEFYLKRFGGHYMESVFQMYTRIAGFLAGGSGGGGSGGHEFGNLMKVLEY GLPITEEHMQFVDRFVVPESYITNPANLPGWTRLFSSHKEVSAYMAKLHAYLKVTREGEFVVFTLPVLTFVSVKEFDEYRRLGGSGGG GSGGIVGIVAGLAVLAVVVIGAVVATVMCRRKSSGGKGGSYSQAASSDSAQGSDVSLTA (SEQ ID NO: 637). In some embodiments, a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 637. [0511] In some embodiments, a polyribonucleotide encodes a polypeptide having amino acids 26-880 of amino acid sequence SEQ ID NO: 637. [0512] In some embodiments, a polyribonucleotide encoding a T-cell string polypeptide can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an UL5.1 polypeptide or antigenic fragment thereof, a linker, an UL39 polypeptide or antigenic fragment thereof, a linker, an UL9 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, an RL2.1 polypeptide or antigenic fragment thereof, a linker, and a MITD. In some embodiments, such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGHEFGNLMKVLEYGLPITEEHMQFVDRFVVPESYITNPANLPGWTRLFSSHKEVSAYMAKLHA YLKVTREGEFVVFTLPVLTFVSVKEFDEYRRLGGSGGGGSGGRTFGSAPRLTEDDFGLLNYALAEMRRLCLDLPPVPPNAYTPYHLRE YATRLVNGFKPLVRRSARLYRILGVLVHLRIRTREASFEEWMRSKEVDLDFGLTERLREHEAQLMILAQALNPYDCLIHSTPNTLVER GLQSALKYEEFYLKRFGGHYMESVFQMYTRIAGFLAGGSGGGGSGGLLNNYDVLVLDEVMSTLGQLYSPTMQQLGRVDALMLRLLR TCPRIIAMDATANAQLVDFLCSLRGEKNVHVVIGEYAMPGFSARRCLFLPRLGPEVLQAALRRRGPAGGAPPPDAPPDATFFGELEAR LAGGDNVCIFSSTVSFAEVVARFCRQFTDRVLLLHSLTPPGDVTTWGRYRVVIYTTVVTVGLSFDPPHFDSMFAYVKPMNYGPDMVS VYQSLGRVRTLRKGELLIYMDGSGARSEPVGGSGGGGSGGETLVAHGPSLYRTFAANPRAASTAKAMRDCVLRQENLIEALASADET LAWCKMCIHHNLPLRPQDPIIGTAAAVLENLATRLRPFLQCYLKARGLCGLDDLCSRRRLSDIKDIASFVLVILARLANRVERGVSEID YTTVGVGAGETMHFYIPGACMAGLIEILDTHRQECSSRVCELTASHTIAPLYVHGKYFYCNSLFGGSGGGGSGGCTDEIAPPLRCQSF PCLHPFCIPCMKTWIPLRNTCPLCNTPVAYLIVGVTASGSFSTIPIVNDPRTRVEAEAAVRAGTAVDFIWTGNPRTAPRSLSGGSGGG GSGGIVGIVAGLAVLAVVVIGAVVATVMCRRKSSGGKGGSYSQAASSDSAQGSDVSLTA (SEQ ID NO: 638). In some embodiments, a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 638. [0513] In some embodiments, a polyribonucleotide encodes a polypeptide having amino acids 26-880 of amino acid sequence SEQ ID NO: 638. [0514] In some embodiments, a polyribonucleotide encoding a T-cell string polypeptide can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL21 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL30.1 polypeptide or antigenic fragment thereof, a linker, an UL29 polypeptide or antigenic fragment thereof, a linker, and a MITD. In some embodiments, such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGGPDAAVFRSSLGSLLYWPGVRALLGRDCRVAARYAGRMTYIATGALLARFNPGAVKCVLPRE AAFAGRVLGGSGGGGSGGGLASDPHYDYIRHYASAAKQALGEVELSGGQLSRAILAQYWKYLQTVVPSGLDIPDDPAGDCDPSLHV LLRPTLLPKLLVRAPFKSGAAAAKYAAAVAGLRDAAHRLQQYMFFMRPADPSRPSTDTALRLSELLAYVSVLYHWASWMLWTADKY VGGSGGGGSGGSPTQKLAVYYYLIHRERRMSPFPALVRLVGRYIQRHGLYVPAPDEPTLADAMNGLGGSGGGGSGGELFGEVFESA PFSTYVDNVIFRGCELLTGSPRGGLMSVALQTDNYTLMGYTYTRVFAFAEELRRRHATAGVAEFLEESPLPYIVLRDQHGFMSVVNT NIGGSGGGGSGGTSQCPDINHLRSLSILNRWLETELVFVGDEEDVSKLSEGELGFYRFLFAFLSAADDLVTENLGGLSGLFEQKDILH YYVEQECIEVVHSRVYNIIQLVLFHNNDQARRAYVARTINHPAIRVKVDWLEARVRECDSIPEKFILMILIEGVFFAASFAAIAYLRTNN LLRGGSGGGGSGGISCLLYDLSTTALEHILLFSLGSCDLPESHLSDLASRGLPAPVVLEFDSEFEMLLAFMTFVKQYGPEFVTGYNIINF DWPFVLTKLTEIYKVPLDGYGRMNGRGVFRVWDIGQSHFGGSGGGGSGGREDIETIAFIKRFSLDYGAINFINLAPNNVSELAMYYM ANQILRYCDHSTYFINTLTAVIAGSRRPPSVQAAAAWAPQGGAGLEAGARALMDSLDAHPGAWTSMFASCNLLRPVMAARPMVVL GLSISKYYGMAGNDRVFQAGNWASLLGGKNACPLLIFDRTRKFVLGGSGGGGSGGIVGIVAGLAVLAVVVIGAVVATVMCRRKSSGG KGGSYSQAASSDSAQGSDVSLTA (SEQ ID NO: 639). In some embodiments, a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 639. [0515] In some embodiments, a polyribonucleotide encodes a polypeptide having amino acids 26-927 of amino acid sequence SEQ ID NO: 639. [0516] In some embodiments, a polyribonucleotide encoding a T-cell string polypeptide can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an UL29 polypeptide or antigenic fragment thereof, a linker, an UL30.1 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL21 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, and a MITD. In some embodiments, such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGREDIETIAFIKRFSLDYGAINFINLAPNNVSELAMYYMANQILRYCDHSTYFINTLTAVIAGSR RPPSVQAAAAWAPQGGAGLEAGARALMDSLDAHPGAWTSMFASCNLLRPVMAARPMVVLGLSISKYYGMAGNDRVFQAGNWASL LGGKNACPLLIFDRTRKFVLGGSGGGGSGGISCLLYDLSTTALEHILLFSLGSCDLPESHLSDLASRGLPAPVVLEFDSEFEMLLAFMTF VKQYGPEFVTGYNIINFDWPFVLTKLTEIYKVPLDGYGRMNGRGVFRVWDIGQSHFGGSGGGGSGGTSQCPDINHLRSLSILNRWL ETELVFVGDEEDVSKLSEGELGFYRFLFAFLSAADDLVTENLGGLSGLFEQKDILHYYVEQECIEVVHSRVYNIIQLVLFHNNDQARRA YVARTINHPAIRVKVDWLEARVRECDSIPEKFILMILIEGVFFAASFAAIAYLRTNNLLRGGSGGGGSGGELFGEVFESAPFSTYVDNVI FRGCELLTGSPRGGLMSVALQTDNYTLMGYTYTRVFAFAEELRRRHATAGVAEFLEESPLPYIVLRDQHGFMSVVNTNIGGSGGGGS GGSPTQKLAVYYYLIHRERRMSPFPALVRLVGRYIQRHGLYVPAPDEPTLADAMNGLGGSGGGGSGGGLASDPHYDYIRHYASAAK QALGEVELSGGQLSRAILAQYWKYLQTVVPSGLDIPDDPAGDCDPSLHVLLRPTLLPKLLVRAPFKSGAAAAKYAAAVAGLRDAAHRL QQYMFFMRPADPSRPSTDTALRLSELLAYVSVLYHWASWMLWTADKYVGGSGGGGSGGGPDAAVFRSSLGSLLYWPGVRALLGR DCRVAARYAGRMTYIATGALLARFNPGAVKCVLPREAAFAGRVLGGSGGGGSGGIVGIVAGLAVLAVVVIGAVVATVMCRRKSSGGK GGSYSQAASSDSAQGSDVSLTA (SEQ ID NO: 640). In some embodiments, a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 640. [0517] In some embodiments, a polyribonucleotide encodes a polypeptide having amino acids 26-927 of amino acid sequence SEQ ID NO: 640. [0518] In some embodiments, a polyribonucleotide encoding a T-cell string polypeptide can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, a RL2.1 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, and a MITD. In some embodiments, such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGCTDEIAPPLRCQSFPCLHPFCIPCMKTWIPLRNTCPLCNTPVAYLIVGVTASGSFSTIPIVNDP RTRVEAEAAVRAGTAVDFIWTGNPRTAPRSLSGGSGGGGSGGETLVAHGPSLYRTFAANPRAASTAKAMRDCVLRQENLIEALASA DETLAWCKMCIHHNLPLRPQDPIIGTAAAVLENLATRLRPFLQCYLKARGLCGLDDLCSRRRLSDIKDIASFVLVILARLANRVERGVS EIDYTTVGVGAGETMHFYIPGACMAGLIEILDTHRQECSSRVCELTASHTIAPLYVHGKYFYCNSLFGGSGGGGSGGELFGEVFESAP FSTYVDNVIFRGCELLTGSPRGGLMSVALQTDNYTLMGYTYTRVFAFAEELRRRHATAGVAEFLEESPLPYIVLRDQHGFMSVVNTNI GGSGGGGSGGTSQCPDINHLRSLSILNRWLETELVFVGDEEDVSKLSEGELGFYRFLFAFLSAADDLVTENLGGLSGLFEQKDILHYY VEQECIEVVHSRVYNIIQLVLFHNNDQARRAYVARTINHPAIRVKVDWLEARVRECDSIPEKFILMILIEGVFFAASFAAIAYLRTNNLL RGGSGGGGSGGGPDAAVFRSSLGSLLYWPGVRALLGRDCRVAARYAGRMTYIATGALLARFNPGAVKCVLPREAAFAGRVLGGSGG GGSGGGLASDPHYDYIRHYASAAKQALGEVELSGGQLSRAILAQYWKYLQTVVPSGLDIPDDPAGDCDPSLHVLLRPTLLPKLLVRAP FKSGAAAAKYAAAVAGLRDAAHRLQQYMFFMRPADPSRPSTDTALRLSELLAYVSVLYHWASWMLWTADKYVGGSGGGGSGGIVG IVAGLAVLAVVVIGAVVATVMCRRKSSGGKGGSYSQAASSDSAQGSDVSLTA (SEQ ID NO: 641). In some embodiments, a
Figure imgf000286_0001
f amino acid sequence SEQ ID NO: 641. [0520] In some embodiments, a polyribonucleotide encoding a T-cell string polypeptide can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, a RL2.1 polypeptide or antigenic fragment thereof, a linker, and a MITD. In some embodiments, such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGGLASDPHYDYIRHYASAAKQALGEVELSGGQLSRAILAQYWKYLQTVVPSGLDIPDDPAGDC DPSLHVLLRPTLLPKLLVRAPFKSGAAAAKYAAAVAGLRDAAHRLQQYMFFMRPADPSRPSTDTALRLSELLAYVSVLYHWASWMLW TADKYVGGSGGGGSGGGPDAAVFRSSLGSLLYWPGVRALLGRDCRVAARYAGRMTYIATGALLARFNPGAVKCVLPREAAFAGRVL GGSGGGGSGGTSQCPDINHLRSLSILNRWLETELVFVGDEEDVSKLSEGELGFYRFLFAFLSAADDLVTENLGGLSGLFEQKDILHYY VEQECIEVVHSRVYNIIQLVLFHNNDQARRAYVARTINHPAIRVKVDWLEARVRECDSIPEKFILMILIEGVFFAASFAAIAYLRTNNLL RGGSGGGGSGGELFGEVFESAPFSTYVDNVIFRGCELLTGSPRGGLMSVALQTDNYTLMGYTYTRVFAFAEELRRRHATAGVAEFLE ESPLPYIVLRDQHGFMSVVNTNIGGSGGGGSGGETLVAHGPSLYRTFAANPRAASTAKAMRDCVLRQENLIEALASADETLAWCKM CIHHNLPLRPQDPIIGTAAAVLENLATRLRPFLQCYLKARGLCGLDDLCSRRRLSDIKDIASFVLVILARLANRVERGVSEIDYTTVGVG AGETMHFYIPGACMAGLIEILDTHRQECSSRVCELTASHTIAPLYVHGKYFYCNSLFGGSGGGGSGGCTDEIAPPLRCQSFPCLHPFCI PCMKTWIPLRNTCPLCNTPVAYLIVGVTASGSFSTIPIVNDPRTRVEAEAAVRAGTAVDFIWTGNPRTAPRSLSGGSGGGGSGGIVGI VAGLAVLAVVVIGAVVATVMCRRKSSGGKGGSYSQAASSDSAQGSDVSLTA (SEQ ID NO: 642). In some embodiments, a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 642. [0521] In some embodiments, a polyribonucleotide encodes a polypeptide having amino acids 26-874 of amino acid sequence SEQ ID NO: 642. [0522] In some embodiments, a polyribonucleotide encoding a T-cell string polypeptide can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, a RL2.1 polypeptide or antigenic fragment thereof, a linker, a RS1 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, and a MITD. In some embodiments, such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGCTDEIAPPLRCQSFPCLHPFCIPCMKTWIPLRNTCPLCNTPVAYLIVGVTASGSFSTIPIVNDP RTRVEAEAAVRAGTAVDFIWTGNPRTAPRSLSGGSGGGGSGGRAAAWMRQVPDPEDVRVVILYSPLPGEDLAAGRAGGGPPPEWS AERGGLSCLLAALGNRLCGPATAAWAGNWTGAPDVSALGAQGVLLLSTRDLAFAGAVEFLGLLAGACDRRLIVVNAVRAADWPADG PVVSRQHAYLACEVLPAVQCAVRWPAARDLRRTVLASGRVFGPGVFARVEAAHARLYPDAPPLRLCRGANVRYRVRTRFGPDTLVP MSPREYRRAVLPALDGRAAASGGSGGGGSGGETLVAHGPSLYRTFAANPRAASTAKAMRDCVLRQENLIEALASADETLAWCKMCI HHNLPLRPQDPIIGTAAAVLENLATRLRPFLQCYLKARGLCGLDDLCSRRRLSDIKDIASFVLVILARLANRVERGVSEIDYTTVGVGA GETMHFYIPGACMAGLIEILDTHRQECSSRVCELTASHTIAPLYVHGKYFYCNSLFGGSGGGGSGGIVGIVAGLAVLAVVVIGAVVAT VMCRRKSSGGKGGSYSQAASSDSAQGSDVSLTA (SEQ ID NO: 643). In some embodiments, a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 643. [0523] In some embodiments, a polyribonucleotide encodes a polypeptide having amino acids 26-587 of amino acid sequence SEQ ID NO: 643. [0524] In some embodiments, a polyribonucleotide encoding a T-cell string polypeptide can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an UL29 polypeptide or antigenic fragment thereof, a linker, an UL39 polypeptide or antigenic fragment thereof, a linker, an UL9 polypeptide or antigenic fragment thereof, a linker, and a MITD. In some embodiments, such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGREDIETIAFIKRFSLDYGAINFINLAPNNVSELAMYYMANQILRYCDHSTYFINTLTAVIAGSR RPPSVQAAAAWAPQGGAGLEAGARALMDSLDAHPGAWTSMFASCNLLRPVMAARPMVVLGLSISKYYGMAGNDRVFQAGNWASL LGGKNACPLLIFDRTRKFVLGGSGGGGSGGRTFGSAPRLTEDDFGLLNYALAEMRRLCLDLPPVPPNAYTPYHLREYATRLVNGFKPL VRRSARLYRILGVLVHLRIRTREASFEEWMRSKEVDLDFGLTERLREHEAQLMILAQALNPYDCLIHSTPNTLVERGLQSALKYEEFYL KRFGGHYMESVFQMYTRIAGFLAGGSGGGGSGGLLNNYDVLVLDEVMSTLGQLYSPTMQQLGRVDALMLRLLRTCPRIIAMDATAN AQLVDFLCSLRGEKNVHVVIGEYAMPGFSARRCLFLPRLGPEVLQAALRRRGPAGGAPPPDAPPDATFFGELEARLAGGDNVCIFSST VSFAEVVARFCRQFTDRVLLLHSLTPPGDVTTWGRYRVVIYTTVVTVGLSFDPPHFDSMFAYVKPMNYGPDMVSVYQSLGRVRTLR KGELLIYMDGSGARSEPVGGSGGGGSGGIVGIVAGLAVLAVVVIGAVVATVMCRRKSSGGKGGSYSQAASSDSAQGSDVSLTA (SEQ ID NO: 644). In some embodiments, a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 644. [0525] In some embodiments, a polyribonucleotide encodes a polypeptide having amino acids 26-638 of amino acid sequence SEQ ID NO: 644. [0526] In some embodiments, a polyribonucleotide encoding a T-cell string polypeptide can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an UL5.1 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL30.1 polypeptide or antigenic fragment thereof, a linker, and a MITD. In some embodiments, such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGHEFGNLMKVLEYGLPITEEHMQFVDRFVVPESYITNPANLPGWTRLFSSHKEVSAYMAKLHA YLKVTREGEFVVFTLPVLTFVSVKEFDEYRRLGGSGGGGSGGELFGEVFESAPFSTYVDNVIFRGCELLTGSPRGGLMSVALQTDNYT LMGYTYTRVFAFAEELRRRHATAGVAEFLEESPLPYIVLRDQHGFMSVVNTNIGGSGGGGSGGTSQCPDINHLRSLSILNRWLETEL VFVGDEEDVSKLSEGELGFYRFLFAFLSAADDLVTENLGGLSGLFEQKDILHYYVEQECIEVVHSRVYNIIQLVLFHNNDQARRAYVAR TINHPAIRVKVDWLEARVRECDSIPEKFILMILIEGVFFAASFAAIAYLRTNNLLRGGSGGGGSGGISCLLYDLSTTALEHILLFSLGSCD LPESHLSDLASRGLPAPVVLEFDSEFEMLLAFMTFVKQYGPEFVTGYNIINFDWPFVLTKLTEIYKVPLDGYGRMNGRGVFRVWDIGQ SHFGGSGGGGSGGIVGIVAGLAVLAVVVIGAVVATVMCRRKSSGGKGGSYSQAASSDSAQGSDVSLTA (SEQ ID NO: 645). In some embodiments, a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 645. [0527] In some embodiments, a polyribonucleotide encodes a polypeptide having amino acids 26-543 of amino acid sequence SEQ ID NO: 645. [0528] In some embodiments, a polyribonucleotide encoding a T-cell string polypeptide can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL21 polypeptide or antigenic fragment thereof, a linker, and a MITD. In some embodiments, such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGGPDAAVFRSSLGSLLYWPGVRALLGRDCRVAARYAGRMTYIATGALLARFNPGAVKCVLPRE AAFAGRVLGGSGGGGSGGGLASDPHYDYIRHYASAAKQALGEVELSGGQLSRAILAQYWKYLQTVVPSGLDIPDDPAGDCDPSLHV LLRPTLLPKLLVRAPFKSGAAAAKYAAAVAGLRDAAHRLQQYMFFMRPADPSRPSTDTALRLSELLAYVSVLYHWASWMLWTADKY VGGSGGGGSGGSPTQKLAVYYYLIHRERRMSPFPALVRLVGRYIQRHGLYVPAPDEPTLADAMNGLGGSGGGGSGGIVGIVAGLAVL AVVVIGAVVATVMCRRKSSGGKGGSYSQAASSDSAQGSDVSLTA (SEQ ID NO: 646). In some embodiments, a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 646. [0529] In some embodiments, a polyribonucleotide encodes a polypeptide having amino acids 26-335 of amino acid sequence SEQ ID NO: 646. [0530] In some embodiments, a polyribonucleotide encoding a T-cell string polypeptide can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an UL29 polypeptide or antigenic fragment thereof, a linker, an UL39 polypeptide or antigenic fragment thereof, a linker, an UL9 polypeptide or antigenic fragment thereof, a linker, an UL5.1 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL30.1 polypeptide or antigenic fragment thereof, a linker, and a MITD. In some embodiments, such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGREDIETIAFIKRFSLDYGAINFINLAPNNVSELAMYYMANQILRYCDHSTYFINTLTAVIAGSR RPPSVQAAAAWAPQGGAGLEAGARALMDSLDAHPGAWTSMFASCNLLRPVMAARPMVVLGLSISKYYGMAGNDRVFQAGNWASL LGGKNACPLLIFDRTRKFVLGGSGGGGSGGRTFGSAPRLTEDDFGLLNYALAEMRRLCLDLPPVPPNAYTPYHLREYATRLVNGFKPL VRRSARLYRILGVLVHLRIRTREASFEEWMRSKEVDLDFGLTERLREHEAQLMILAQALNPYDCLIHSTPNTLVERGLQSALKYEEFYL KRFGGHYMESVFQMYTRIAGFLAGGSGGGGSGGLLNNYDVLVLDEVMSTLGQLYSPTMQQLGRVDALMLRLLRTCPRIIAMDATAN AQLVDFLCSLRGEKNVHVVIGEYAMPGFSARRCLFLPRLGPEVLQAALRRRGPAGGAPPPDAPPDATFFGELEARLAGGDNVCIFSST VSFAEVVARFCRQFTDRVLLLHSLTPPGDVTTWGRYRVVIYTTVVTVGLSFDPPHFDSMFAYVKPMNYGPDMVSVYQSLGRVRTLR KGELLIYMDGSGARSEPVGGSGGGGSGGHEFGNLMKVLEYGLPITEEHMQFVDRFVVPESYITNPANLPGWTRLFSSHKEVSAYMA KLHAYLKVTREGEFVVFTLPVLTFVSVKEFDEYRRLGGSGGGGSGGELFGEVFESAPFSTYVDNVIFRGCELLTGSPRGGLMSVALQT DNYTLMGYTYTRVFAFAEELRRRHATAGVAEFLEESPLPYIVLRDQHGFMSVVNTNIGGSGGGGSGGTSQCPDINHLRSLSILNRWL ETELVFVGDEEDVSKLSEGELGFYRFLFAFLSAADDLVTENLGGLSGLFEQKDILHYYVEQECIEVVHSRVYNIIQLVLFHNNDQARRA YVARTINHPAIRVKVDWLEARVRECDSIPEKFILMILIEGVFFAASFAAIAYLRTNNLLRGGSGGGGSGGISCLLYDLSTTALEHILLFSL GSCDLPESHLSDLASRGLPAPVVLEFDSEFEMLLAFMTFVKQYGPEFVTGYNIINFDWPFVLTKLTEIYKVPLDGYGRMNGRGVFRV WDIGQSHFGGSGGGGSGGIVGIVAGLAVLAVVVIGAVVATVMCRRKSSGGKGGSYSQAASSDSAQGSDVSLTA (SEQ ID NO: 647). In some embodiments, a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 647. [0531] In some embodiments, a polyribonucleotide encodes a polypeptide having amino acids 26-1156 of amino acid sequence SEQ ID NO: 647. [0532] In some embodiments, a polyribonucleotide encoding a T-cell string polypeptide can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an UL30.1 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL5.1 polypeptide or antigenic fragment thereof, a linker, an UL9 polypeptide or antigenic fragment thereof, a linker, an UL39 polypeptide or antigenic fragment thereof, a linker, an UL29 polypeptide or antigenic fragment thereof, a linker, and a MITD. In some embodiments, such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGISCLLYDLSTTALEHILLFSLGSCDLPESHLSDLASRGLPAPVVLEFDSEFEMLLAFMTFVKQYG PEFVTGYNIINFDWPFVLTKLTEIYKVPLDGYGRMNGRGVFRVWDIGQSHFGGSGGGGSGGTSQCPDINHLRSLSILNRWLETELVF VGDEEDVSKLSEGELGFYRFLFAFLSAADDLVTENLGGLSGLFEQKDILHYYVEQECIEVVHSRVYNIIQLVLFHNNDQARRAYVARTI NHPAIRVKVDWLEARVRECDSIPEKFILMILIEGVFFAASFAAIAYLRTNNLLRGGSGGGGSGGELFGEVFESAPFSTYVDNVIFRGCE LLTGSPRGGLMSVALQTDNYTLMGYTYTRVFAFAEELRRRHATAGVAEFLEESPLPYIVLRDQHGFMSVVNTNIGGSGGGGSGGHEF GNLMKVLEYGLPITEEHMQFVDRFVVPESYITNPANLPGWTRLFSSHKEVSAYMAKLHAYLKVTREGEFVVFTLPVLTFVSVKEFDEY RRLGGSGGGGSGGLLNNYDVLVLDEVMSTLGQLYSPTMQQLGRVDALMLRLLRTCPRIIAMDATANAQLVDFLCSLRGEKNVHVVI GEYAMPGFSARRCLFLPRLGPEVLQAALRRRGPAGGAPPPDAPPDATFFGELEARLAGGDNVCIFSSTVSFAEVVARFCRQFTDRVLL LHSLTPPGDVTTWGRYRVVIYTTVVTVGLSFDPPHFDSMFAYVKPMNYGPDMVSVYQSLGRVRTLRKGELLIYMDGSGARSEPVGG SGGGGSGGRTFGSAPRLTEDDFGLLNYALAEMRRLCLDLPPVPPNAYTPYHLREYATRLVNGFKPLVRRSARLYRILGVLVHLRIRTR EASFEEWMRSKEVDLDFGLTERLREHEAQLMILAQALNPYDCLIHSTPNTLVERGLQSALKYEEFYLKRFGGHYMESVFQMYTRIAG FLAGGSGGGGSGGREDIETIAFIKRFSLDYGAINFINLAPNNVSELAMYYMANQILRYCDHSTYFINTLTAVIAGSRRPPSVQAAAAW APQGGAGLEAGARALMDSLDAHPGAWTSMFASCNLLRPVMAARPMVVLGLSISKYYGMAGNDRVFQAGNWASLLGGKNACPLLIF DRTRKFVLGGSGGGGSGGIVGIVAGLAVLAVVVIGAVVATVMCRRKSSGGKGGSYSQAASSDSAQGSDVSLTA (SEQ ID NO: 648). In some embodiments, a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 648. [0533] In some embodiments, a polyribonucleotide encodes a polypeptide having amino acids 26-1156 of amino acid sequence SEQ ID NO: 648. [0534] In some embodiments, a polyribonucleotide encoding a T-cell string polypeptide can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, a RL2.1 polypeptide or antigenic fragment thereof, a linker, a RS1 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL21 polypeptide or antigenic fragment thereof, a linker, and a MITD. In some embodiments, such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGCTDEIAPPLRCQSFPCLHPFCIPCMKTWIPLRNTCPLCNTPVAYLIVGVTASGSFSTIPIVNDP RTRVEAEAAVRAGTAVDFIWTGNPRTAPRSLSGGSGGGGSGGRAAAWMRQVPDPEDVRVVILYSPLPGEDLAAGRAGGGPPPEWS AERGGLSCLLAALGNRLCGPATAAWAGNWTGAPDVSALGAQGVLLLSTRDLAFAGAVEFLGLLAGACDRRLIVVNAVRAADWPADG PVVSRQHAYLACEVLPAVQCAVRWPAARDLRRTVLASGRVFGPGVFARVEAAHARLYPDAPPLRLCRGANVRYRVRTRFGPDTLVP MSPREYRRAVLPALDGRAAASGGSGGGGSGGETLVAHGPSLYRTFAANPRAASTAKAMRDCVLRQENLIEALASADETLAWCKMCI HHNLPLRPQDPIIGTAAAVLENLATRLRPFLQCYLKARGLCGLDDLCSRRRLSDIKDIASFVLVILARLANRVERGVSEIDYTTVGVGA GETMHFYIPGACMAGLIEILDTHRQECSSRVCELTASHTIAPLYVHGKYFYCNSLFGGSGGGGSGGGPDAAVFRSSLGSLLYWPGVR ALLGRDCRVAARYAGRMTYIATGALLARFNPGAVKCVLPREAAFAGRVLGGSGGGGSGGGLASDPHYDYIRHYASAAKQALGEVELS GGQLSRAILAQYWKYLQTVVPSGLDIPDDPAGDCDPSLHVLLRPTLLPKLLVRAPFKSGAAAAKYAAAVAGLRDAAHRLQQYMFFMR PADPSRPSTDTALRLSELLAYVSVLYHWASWMLWTADKYVGGSGGGGSGGSPTQKLAVYYYLIHRERRMSPFPALVRLVGRYIQRH GLYVPAPDEPTLADAMNGLGGSGGGGSGGIVGIVAGLAVLAVVVIGAVVATVMCRRKSSGGKGGSYSQAASSDSAQGSDVSLTA (SEQ ID NO: 649). In some embodiments, a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 649. [0535] In some embodiments, a polyribonucleotide encodes a polypeptide having amino acids 26-897 of amino acid sequence SEQ ID NO: 649. [0536] In some embodiments, a polyribonucleotide encoding a T-cell string polypeptide can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an UL21 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, a RS1 polypeptide or antigenic fragment thereof, a linker, a RL2.1 polypeptide or antigenic fragment thereof, a linker, and a MITD. In some embodiments, such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGSPTQKLAVYYYLIHRERRMSPFPALVRLVGRYIQRHGLYVPAPDEPTLADAMNGLGGSGGGG SGGGLASDPHYDYIRHYASAAKQALGEVELSGGQLSRAILAQYWKYLQTVVPSGLDIPDDPAGDCDPSLHVLLRPTLLPKLLVRAPFK SGAAAAKYAAAVAGLRDAAHRLQQYMFFMRPADPSRPSTDTALRLSELLAYVSVLYHWASWMLWTADKYVGGSGGGGSGGGPDA AVFRSSLGSLLYWPGVRALLGRDCRVAARYAGRMTYIATGALLARFNPGAVKCVLPREAAFAGRVLGGSGGGGSGGETLVAHGPSLY RTFAANPRAASTAKAMRDCVLRQENLIEALASADETLAWCKMCIHHNLPLRPQDPIIGTAAAVLENLATRLRPFLQCYLKARGLCGLD DLCSRRRLSDIKDIASFVLVILARLANRVERGVSEIDYTTVGVGAGETMHFYIPGACMAGLIEILDTHRQECSSRVCELTASHTIAPLYV HGKYFYCNSLFGGSGGGGSGGRAAAWMRQVPDPEDVRVVILYSPLPGEDLAAGRAGGGPPPEWSAERGGLSCLLAALGNRLCGPA TAAWAGNWTGAPDVSALGAQGVLLLSTRDLAFAGAVEFLGLLAGACDRRLIVVNAVRAADWPADGPVVSRQHAYLACEVLPAVQC AVRWPAARDLRRTVLASGRVFGPGVFARVEAAHARLYPDAPPLRLCRGANVRYRVRTRFGPDTLVPMSPREYRRAVLPALDGRAAA SGGSGGGGSGGCTDEIAPPLRCQSFPCLHPFCIPCMKTWIPLRNTCPLCNTPVAYLIVGVTASGSFSTIPIVNDPRTRVEAEAAVRAG TAVDFIWTGNPRTAPRSLSGGSGGGGSGGIVGIVAGLAVLAVVVIGAVVATVMCRRKSSGGKGGSYSQAASSDSAQGSDVSLTA (SEQ ID NO: 650). In some embodiments, a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 650. [0537] In some embodiments, a polyribonucleotide encodes a polypeptide having amino acids 26-897 of amino acid sequence SEQ ID NO: 650. [0538] In some embodiments, a polyribonucleotide encoding a T-cell string polypeptide can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an UL5.1 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL30.1 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL21 polypeptide or antigenic fragment thereof, a linker, and a MITD. In some embodiments, such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGHEFGNLMKVLEYGLPITEEHMQFVDRFVVPESYITNPANLPGWTRLFSSHKEVSAYMAKLHA YLKVTREGEFVVFTLPVLTFVSVKEFDEYRRLGGSGGGGSGGELFGEVFESAPFSTYVDNVIFRGCELLTGSPRGGLMSVALQTDNYT LMGYTYTRVFAFAEELRRRHATAGVAEFLEESPLPYIVLRDQHGFMSVVNTNIGGSGGGGSGGTSQCPDINHLRSLSILNRWLETEL VFVGDEEDVSKLSEGELGFYRFLFAFLSAADDLVTENLGGLSGLFEQKDILHYYVEQECIEVVHSRVYNIIQLVLFHNNDQARRAYVAR TINHPAIRVKVDWLEARVRECDSIPEKFILMILIEGVFFAASFAAIAYLRTNNLLRGGSGGGGSGGISCLLYDLSTTALEHILLFSLGSCD LPESHLSDLASRGLPAPVVLEFDSEFEMLLAFMTFVKQYGPEFVTGYNIINFDWPFVLTKLTEIYKVPLDGYGRMNGRGVFRVWDIGQ SHFGGSGGGGSGGGPDAAVFRSSLGSLLYWPGVRALLGRDCRVAARYAGRMTYIATGALLARFNPGAVKCVLPREAAFAGRVLGGS GGGGSGGGLASDPHYDYIRHYASAAKQALGEVELSGGQLSRAILAQYWKYLQTVVPSGLDIPDDPAGDCDPSLHVLLRPTLLPKLLV RAPFKSGAAAAKYAAAVAGLRDAAHRLQQYMFFMRPADPSRPSTDTALRLSELLAYVSVLYHWASWMLWTADKYVGGSGGGGSG GSPTQKLAVYYYLIHRERRMSPFPALVRLVGRYIQRHGLYVPAPDEPTLADAMNGLGGSGGGGSGGIVGIVAGLAVLAVVVIGAVVAT VMCRRKSSGGKGGSYSQAASSDSAQGSDVSLTA (SEQ ID NO: 651). In some embodiments, a polyribonucleotide
Figure imgf000291_0001
, p y u p yp p v g - of amino acid sequence SEQ ID NO: 651. [0540] In some embodiments, a polyribonucleotide encoding a T-cell string polypeptide can include, in 5’ to 3’ order, nucleotide sequences that encode an HSV-1 gD secretory signal, an UL21 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL30.1 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL5.1 polypeptide or antigenic fragment thereof, a linker, and a MITD. In some embodiments, such a polyribonucleotide encodes a polypeptide having an amino acid sequence comprising or consisting of MGGAAARLGAVILFVVIVGLHGVRGSPTQKLAVYYYLIHRERRMSPFPALVRLVGRYIQRHGLYVPAPDEPTLADAMNGLGGSGGGG SGGGLASDPHYDYIRHYASAAKQALGEVELSGGQLSRAILAQYWKYLQTVVPSGLDIPDDPAGDCDPSLHVLLRPTLLPKLLVRAPFK SGAAAAKYAAAVAGLRDAAHRLQQYMFFMRPADPSRPSTDTALRLSELLAYVSVLYHWASWMLWTADKYVGGSGGGGSGGGPDA AVFRSSLGSLLYWPGVRALLGRDCRVAARYAGRMTYIATGALLARFNPGAVKCVLPREAAFAGRVLGGSGGGGSGGISCLLYDLSTT ALEHILLFSLGSCDLPESHLSDLASRGLPAPVVLEFDSEFEMLLAFMTFVKQYGPEFVTGYNIINFDWPFVLTKLTEIYKVPLDGYGRM NGRGVFRVWDIGQSHFGGSGGGGSGGTSQCPDINHLRSLSILNRWLETELVFVGDEEDVSKLSEGELGFYRFLFAFLSAADDLVTEN LGGLSGLFEQKDILHYYVEQECIEVVHSRVYNIIQLVLFHNNDQARRAYVARTINHPAIRVKVDWLEARVRECDSIPEKFILMILIEGVF FAASFAAIAYLRTNNLLRGGSGGGGSGGELFGEVFESAPFSTYVDNVIFRGCELLTGSPRGGLMSVALQTDNYTLMGYTYTRVFAFAE ELRRRHATAGVAEFLEESPLPYIVLRDQHGFMSVVNTNIGGSGGGGSGGHEFGNLMKVLEYGLPITEEHMQFVDRFVVPESYITNPA NLPGWTRLFSSHKEVSAYMAKLHAYLKVTREGEFVVFTLPVLTFVSVKEFDEYRRLGGSGGGGSGGIVGIVAGLAVLAVVVIGAVVAT VMCRRKSSGGKGGSYSQAASSDSAQGSDVSLTA (SEQ ID NO: 692). In some embodiments, a polyribonucleotide encodes a polypeptide having an amino acid sequence having at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 99%, or 100% sequence identity with SEQ ID NO: 692. [0541] In some embodiments, a polyribonucleotide encodes a polypeptide having amino acids 26-853 of amino acid sequence SEQ ID NO: 692. Table 19: Example ribonucleotide sequences encoding T-cell string polypeptides SEQ Ribonucleic acid ID c u a g cc ca u gc g cc c au ga c ga ga
Figure imgf000292_0001
ccuagccuguaccggaccuuugccgccaauccuagagccgccucuacagccaaggccaugagagacugcgugcugcggcaagagaaucugauuga ggcccuggccagcgccgaugagacacuggcuuggugcaagaugugcauccaccacaaccugccucugcggccucaggacccuaucauuggaacag cugccgccgugcuggaaaaccuggcuacaagacugcggccauuccuccagugcuaccugaaggccagaggacugugcggccuggaugaucugug c g g cg g au au aa u u u a g g u c cc gc a gc c a u c c ca a g c ca au ca gg ca g cu ca g u g g ag ag u
Figure imgf000293_0001
ggcggaggaagcgguggacugcugaacaacuacgacgugcugguccuggacgaagugaugagcacacugggacagcuguacuccccaaccaugc agcagcugggcagaguggaugcccugaugcugagacugcugcggacaugccccagaauuaucgccauggacgccaccgccaaugcucagcuggu ggauuuccugugcagccugagaggcgagaagaaugugcacguggucaucggcgaguacgcuaugccuggcuucagcgccagaagaugccuguu ca u a gg g u g ac gc u u g ca uc u au g cc u u u u u cc ca uc c uc cg c g u ac g gu ga u c ca cu ga cu a
Figure imgf000294_0001
gccagagugcgcgagugugauagcauccccgagaaguucauccugaugauccugaucgaaggcguguucuucgccgccagcuuugccgccauug ccuaccugaggaccaacaaccugcugagagguggaaguggcggagguggaagcggaggacacgaguucggcaaccugaugaaggugcucgaau acggccugccuaucaccgaggaacacaugcaguucguggacagauucguggugcccgagagcuacaucacaaaccccgccaaucugcccggcugg gu c g g uc c cc c g u u u gc u c a a uc c a g ac c uc a u ac u gu u ug c ca a cg ug ca g g cu au c
Figure imgf000295_0001
ugugggauagacaugcccagagagccuaccuggucaaccccuuccuguuugccgccggauuccuggaagaucugagccacucuguguuccccgcc gacacacaagagacaacaggcggaucugguggcggaggaucuggcggagaugguagacugcugcacaacacacaggccagagccgccgaugccg cugacgauagaccucauagaccugccgauuggaccgugcaccacaagaucuacuacuacgugcuggugcccgccuucagcagaggcagauguug u a u gc g u uc cu a ua a g cu gc ca cu uu ug gg ac c ac u ca 626 ug ga u ga cc g a gc au gc cu g c gc cu u gu gg cu cc u
Figure imgf000296_0001
gaauggccucggugguagcgguggugguggaucuggcggagaugguagacugcugcacaacacacaggccagagcugccgaugccgccgacga uagaccucauagaccugccgauuggaccgugcaccacaagaucuacuacuaugugcuggugcccgccuucagcagaggcagauguuguacagcc ggcgucagauucgaccggguguacgccacacuccagaacaugguggugccugagauugccccuggcgaggaaugcccuagcgacccuguuaccg g u u g g cc ug g g cu u u gc g cc c au ga cc ga ga ga ag g c g g g g ac au aa u u u a cg g u c cc gc a gc
Figure imgf000297_0001
ugcuucugguggaucuggcggaggugguaguggcggauguaccgaugagauugccccaccucugcggugccagagcuuuccuugucugcaccc cuucugcaucccuugcaugaagaccuggauuccucugcggaacaccuguccucugugcaacacaccaguggccuaccugauugugggcgugaca gccucuggcagcuucagcacaauccccaucgugaacgaccccagaaccagaguugaagccgaggccgcuguuagagcuggaaccgccguggacuu g c a a a a g u c g gc c au g g ug a g cu u ac g g ug c c u u g g c c u g g cg c a gc g c a
Figure imgf000298_0001
gaugugaagugggcagagugcuggccguggucaacgauccuagaggcccuuucuucgugggccugaucgccugugugcagcuggaaagagucc uggaaacagccgccagcgccgccaucuuugaaagaaggggaccugcucugagcagagaggaacggcugcuguaccugaucaccaacuaccugccu agcgugucccugagcaccaagagaaggggagaugaggucccaccugacagaacccuguuugcccauguggcccugugugccauugguagaaggc ua ga a a a g cu gc ug cc c c u u ug g ca c ac cu cg gc u c u a g cc ca u gc g cc c au ga c ga ga ga ag g c g g
Figure imgf000299_0001
caacagccuguuuggcggcaguggugguggcggcucaggcggaggacuuauugcaggcgcuguuggagguucucugcuggcugcccuggucau cuguggcaucguguacuggaugcggcggcacacacagaaggccccuaagagaaucagacugccccacaucagaugauga SECgD1 UL2 GS linkerRL2 Gslinker RS1 GS linker UL54 GS linker VSV G TMD: Version 1 c u a g cc ca u gc g cc c u ga c ga ga ga ag g c g g u U A CG G U C G A G G G C G G U G C A U C C G
Figure imgf000300_0001
AUGCUGUGGACGGCGGACAAGUACGUGGGAGGUUCAGGCGGAGGCGGAUCUGGCGGAAGCCCGACCCAAAAGCUCGC CGUCUACUACUAUCUCAUCCACCGGGAGCGGCGCAUGUCCCCCUUCCCCGCGCUCGUGCGGCUCGUCGGUCGGUACAU CCAGCGCCACGGCCUGUACGUUCCCGCGCCCGACGAACCGACGUUGGCCGAUGCCAUGAACGGGCUGGGCGGUAGCGG G G U A G G U C G A G G G G GA G G A CC G CU C U U GA C G C A C A C A G C GC G G C C G AU
Figure imgf000301_0001
GCGCGACUGCGUGCUGCGCCAGGAAAAUCUCAUCGAGGCCCUGGCGUCCGCGGAUGAGACGCUGGCGUGGUGCAAGA UGUGCAUUCACCACAAUCUGCCGCUCCGCCCCCAGGACCCUAUCAUCGGAACGGCGGCCGCCGUGCUGGAAAACCUCG CCACGCGCCUGCGCCCCUUUCUGCAGUGCUACCUGAAGGCCCGAGGCCUGUGCGGGCUGGACGACCUGUGCUCGCGG GC G C UA U G G G G C A C A C G CU C GC G U G U G A G G G A U C U CU G A A G C C C A G G AC
Figure imgf000302_0001
UACGGCAUGGCCGGCAACGACCGCGUGUUUCAGGCCGGGAACUGGGCCAGCCUGCUGGGCGGCAAAAACGCGUGCCC GCUGCUGAUCUUUGACCGCACCCGCAAGUUUGUCCUGGGCGGUUCAGGCGGCGGUGGUAGCGGCGGACGAACCUUCG GCAGCGCCCCCCGCCUCACGGAGGACGACUUUGGGCUCCUGAACUACGCGCUCGCUGAGAUGCGACGCCUGUGCCUGG A G A A U AC G U U G A G G GC C U U A G GU C G U G U G C C G C U G C C G G GC 657 AG C U G U C C G
Figure imgf000303_0001
GAAGCGCCCCUAGACUGACCGAGGACGAUUUCGGCCUGCUGAAUUAUGCCCUGGCCGAGAUGAGAAGGCUGUGCCUG GAUCUGCCUCCUGUGCCUCCUAACGCCUACACACCCUACCACCUGAGAGAGUACGCCACCAGACUGGUCAACGGCUUCA AACCCCUCGUGCGGAGAAGCGCCAGACUGUACAGAAUCCUGGGCGUGCUGGUGCACCUGAGAAUCAGAACAAGAGAGG G AA C C UG G C G G G C U G G CA C A GA G G GC A GC C GA C G G G U GC U G G CA 658 A G G C C G U UG C CU
Figure imgf000304_0001
GGCCUUCAUGACCUUCGUCAAGCAGUACGGCCCCGAGUUCGUGACCGGGUACAACAUCAUCAACUUCGACUGGCCCUU CGUCCUGACCAAGCUGACGGAGAUCUACAAGGUCCCGCUCGACGGGUACGGGCGCAUGAACGGCCGGGGUGUGUUCC GCGUGUGGGACAUCGGCCAGAGCCACUUUGGCGGCUCCGGUGGCGGAGGCUCUGGUGGUGAGCUCUUCGGGGAGGU C G U G CC G C G AU A C G C G UG AA U AA U G C A C C G U C C C GG C UG GG 659 A G U C AC A U U GA G C U
Figure imgf000305_0001
CAGAGUGUGGGACAUCGGCCAGUCUCACUUUGGAGGAAGCGGAGGCGGAGGUAGUGGCGGAGAACUGUUUGGAGAGG UGUUCGAGAGCGCCCCUUUCAGCACCUACGUGGACAACGUGAUCUUCCGGGGCUGCGAACUGCUGACAGGAUCUCCUA GAGGCGGCCUGAUGUCUGUGGCCCUCCAGACCGAUAACUACACCCUGAUGGGCUACACCUACACCAGAGUGUUUGCCU CG A A G C U G CC C G CU U A A G U C U G C G G AU U U G U U A A G 660 U A CG GG U C G A UG G G C G U C
Figure imgf000306_0001
GCCAUGGACGCCACCGCCAACGCGCAGCUGGUGGACUUUCUGUGCAGCCUCCGGGGCGAAAAGAACGUUCACGUGGUC AUCGGGGAGUACGCCAUGCCCGGAUUUUCGGCGCGCCGUUGUCUGUUUCUCCCGCGCCUGGGGCCCGAGGUCCUGCA GGCGGCCCUGCGCCGCCGGGGGCCGGCGGGCGGGGCGCCCCCCCCGGACGCCCCCCCGGACGCCACCUUCUUCGGGGA A CC C G UA A C G A CC A G G C C U U AG U A G G U C G A U AG UG G G C C U G C A A CC A G GA C G GA U
Figure imgf000307_0001
AACACACUGGUGGAAAGGGGACUCCAGAGCGCCCUGAAGUACGAGGAAUUCUACCUGAAAAGAUUCGGCGGCCACUAC AUGGAAAGCGUGUUCCAGAUGUAUACCCGGAUCGCCGGCUUUCUGGCAGGCGGUAGCGGAGGUGGUGGCUCUGGUG GACACGAGUUCGGCAACCUGAUGAAGGUGCUGGAAUACGGCCUGCCUAUCACCGAGGAACACAUGCAGUUCGUGGACA G G G G C C C G C U A G U C AA A A AG CC U C G U U U A G C U GC U U G C G A AG AC U U G
Figure imgf000308_0001
663 AUGGGCGGAGCUGCUGCUAGACUGGGAGCCGUGAUCCUGUUCGUGGUUAUCGUGGGACUGCAUGGCGUGCGGGGAC ACGAGUUUGGCAACCUGAUGAAGGUGCUGGAAUACGGCCUGCCUAUCACCGAGGAACACAUGCAGUUCGUGGACAGAU UCGUGGUGCCCGAGAGCUACAUCACAAACCCCGCCAAUCUGCCCGGCUGGACCAGACUGUUUAGCAGCCACAAAGAGG G A G U G A AC A C C UG U GA C A C A A CC G AC G A A A C A G CC G G C G 664 G C C U G C C G C A C AC
Figure imgf000309_0001
GUUCCCGCGCCCGACGAACCGACGUUGGCCGAUGCCAUGAACGGGCUGGGCGGUUCAGGCGGCGGUGGUAGCGGCGG AGAGCUCUUCGGGGAGGUGUUUGAGAGCGCCCCCUUCAGCACCUACGUGGACAAUGUCAUUUUCCGGGGCUGCGAGC UGCUGACCGGCUCGCCGCGCGGGGGGCUGAUGUCCGUGGCCCUGCAGACGGACAACUACACGCUGAUGGGGUACACG A G U U G G G G G CG G A A GA G C G C A GU G A A G A U U G CA G G C A CC CG A U A AG GG A C G UC G
Figure imgf000310_0001
GACUGGCUGGAAGCCAGAGUGCGCGAGUGUGAUAGCAUCCCUGAGAAGUUCAUCCUCAUGAUUCUGAUCGAGGGCGU GUUCUUCGCCGCCAGCUUUGCCGCCAUUGCCUACCUGAGGACCAACAACCUUCUGAGAGGCGGCAGUGGUGGUGGCG GCUCAGGCGGAAUUAGCUGUCUGCUGUACGACCUGAGCACCACCGCUCUGGAACACAUCCUGCUGUUUAGCCUGGGCA C U U A G C CU G A C U G G C C A G G AC C C C G C G A CG G C U C CU GA G A C G C G C U C GU CC CU
Figure imgf000311_0001
CGUGGAUGCUGUGGACGGCGGACAAGUACGUGGGAGGUUCAGGUGGUGGCGGUAGUGGUGGAGGCCCCGACGCCGC GGUCUUUCGCAGUUCGCUGGGGUCCCUGCUGUACUGGCCCGGGGUGCGCGCGCUCCUGGGGCGCGACUGUCGCGUG GCCGCCCGCUACGCGGGGCGCAUGACGUACAUCGCCACCGGGGCUCUGCUCGCCCGCUUCAACCCCGGCGCCGUCAAA G G U AG C U G U C C C C G C G A U U U A G U GA CC G G C U U U GA G U CU U U G G G G A
Figure imgf000312_0001
668 AUGGGGGGGGCUGCCGCCAGGUUGGGGGCCGUGAUUUUGUUUGUCGUCAUAGUCGGCCUCCAUGGGGUCCGCGGCU GCACGGACGAGAUCGCCCCGCCCCUGCGCUGCCAGAGUUUUCCCUGCCUGCACCCCUUCUGCAUCCCGUGCAUGAAGA CCUGGAUUCCGUUGCGCAACACGUGUCCCCUGUGCAACACCCCGGUGGCGUACCUGAUAGUGGGCGUGACCGCCAGCG GG U C G A G G G C G U G C A U C C C G G A G A C A U C CA A G A C C 669 U A G G U C G A U AG UG G
Figure imgf000313_0001
CCUCUCACACAAUCGCCCCUCUGUAUGUGCACGGCAAGUACUUCUACUGCAACAGCCUGUUUGGCGGCAGCGGCGGAG GUGGAAGCGGAGGCGAACUUUUUGGAGAGGUGUUCGAGAGCGCCCCUUUCAGCACCUACGUGGACAACGUGAUCUUC CGGGGAUGCGAGCUGCUGACAGGAUCUCCUAGAGGCGGACUGAUGUCUGUGGCCCUCCAGACCGACAACUACACCCUG AG C UG G C CA AU C UU CU G C C G A UA CC UC G GA A S 670 G C G G C U G AC C C CA C U U AC U G G G C U C A UC G
Figure imgf000314_0001
UGCUACCUGAAGGCCCGAGGCCUGUGCGGGCUGGACGACCUGUGCUCGCGGCGACGCCUGUCGGACAUUAAGGAUAU UGCCUCCUUUGUGUUGGUCAUCCUGGCCCGCCUCGCCAACCGCGUCGAGCGCGGCGUGUCGGAGAUCGACUACACGA CCGUGGGGGUUGGGGCCGGCGAGACGAUGCACUUUUACAUCCCGGGGGCCUGCAUGGCGGGUCUCAUUGAAAUACUG C C G U U A U G 671 G C G UG C U G AC C A GA C U G CA C A AC A C A C G G G U AC G G C G C G U C C
Figure imgf000315_0001
SEC_RL2.1_GS linker_RS1_GS linker_UL54_GS linker_MITD: Wild type 672 AUGGGGGGGGCUGCCGCCAGGUUGGGGGCCGUGAUUUUGUUUGUCGUCAUAGUCGGCCUCCAUGGGGUCCGCGGCU GCACGGACGAGAUCGCCCCGCCCCUGCGCUGCCAGAGUUUUCCCUGCCUGCACCCCUUCUGCAUCCCGUGCAUGAAGA G G U U U CC G C U G G C U A C U U CC A G G GC U A G G U U U U A G A U U G C G G G U A G C C
Figure imgf000316_0001
AGGACUGGCAGUGCUGGCCGUGGUGGUGAUCGGAGCCGUGGUGGCUACCGUGAUGUGCAGACGGAAGUCCAGCGGAG GCAAGGGCGGCAGCUACAGCCAGGCCGCCAGCUCUGAUAGCGCCCAGGGCAGCGACGUGUCACUGACAGCCUGAUAA SEC-gD1 UL29 GS linker UL39 GS linker UL9 GS linker MITD: Wild tpe C C A G G AC C G G A G A A U C G U U G A G G C C G GC G C U G U C C G G CA G G AA C C G G C G
Figure imgf000317_0001
UGUGCAUCUUCAGCAGCACCGUGUCCUUCGCCGAAGUGGUGGCCAGAUUCUGUCGGCAGUUCACCGAUAGAGUGCUG CUGCUCCACAGCCUGACACCACCUGGGGAUGUGACAACCUGGGGCAGAUACCGGGUCGUGAUCUACACCACCGUGGUC ACAGUGGGCCUGUCCUUCGAUCCUCCUCACUUCGACAGCAUGUUUGCCUACGUGAAGCCCAUGAACUACGGCCCCGAU G G G C C G C CG G C G G C C A U A G C C U C C G G C C U G G A U A AG G A C A G G U G C A
Figure imgf000318_0001
CAGCGAGUUCGAGAUGCUGCUGGCCUUCAUGACCUUUGUGAAGCAGUACGGCCCUGAGUUCGUGACCGGCUACAAUA UCAUCAACUUCGACUGGCCCUUCGUGCUGACCAAGCUGACCGAGAUCUACAAGGUGCCCCUGGACGGCUACGGCAGAA UGAAUGGACGGGGAGUGUUCAGAGUGUGGGACAUCGGCCAGUCUCACUUUGGAGGUUCAGGUGGCGGUGGUUCAGG G C G C C U G C C G C A C C G C G G A U U G CA G G C A CC G A A U C C A G G AC C G G
Figure imgf000319_0001
ACCUUCCCCCGGUCCCCCCCAACGCAUACACGCCCUAUCAUCUGAGGGAGUAUGCGACGCGGCUGGUUAACGGGUUCA AACCCCUGGUGCGGCGGUCCGCCCGCCUGUAUCGCAUCCUGGGGGUUCUGGUCCACCUGCGCAUCCGUACCCGGGAG GCCUCCUUUGAGGAAUGGAUGCGCUCCAAGGAGGUGGACCUGGACUUCGGGCUGACGGAAAGGCUUCGCGAACACGA A U AC G U U G A G G GC C U U A G GU C G U G U G C C G C U G C C G G GC 681 AG C U G U C C G G CA G
Figure imgf000320_0001
CCAGCUUCGAGGAAUGGAUGCGGAGCAAAGAGGUGGACCUGGACUUUGGCCUGACCGAGAGACUGAGAGAGCACGAG GCCCAGCUGAUGAUUCUGGCCCAGGCUCUGAACCCCUACGACUGCCUGAUCCACAGCACACCCAACACACUGGUGGAAA GAGGCCUCCAGAGCGCCCUGAAGUACGAGGAAUUCUACCUGAAAAGAUUCGGCGGCCACUACAUGGAAAGCGUGUUCC C UG G C G G G C U G G CA C A GA G G GC A GC C GA C G G G U GC U G G CA 682 A C A U U G A G G C A UC U
Figure imgf000321_0001
GUUUGAGAGCGCCCCCUUCAGCACCUACGUGGACAAUGUCAUUUUCCGGGGCUGCGAGCUGCUGACCGGCUCGCCGC GCGGGGGGCUGAUGUCCGUGGCCCUGCAGACGGACAACUACACGCUGAUGGGGUACACGUACACCCGGGUGUUCGCG UUCGCGGAGGAGCUGCGGCGGCGGCACGCGACGGCCGGCGUGGCCGAGUUCUUGGAGGAGUCCCCCCUGCCCUACAU G CC G C G AU A C G C G UG AA U AA U G C A C C G U C C C GG C UG GG 683 A GC G C GC UG A G G C U U U AG U
Figure imgf000322_0001
CGCCGAGGAACUGAGAAGGCGGCAUGCUACAGCUGGCGUGGCCGAGUUCCUGGAAGAAAGCCCUCUGCCUUACAUCGU GCUGAGGGACCAGCACGGCUUCAUGAGCGUGGUCAACACCAAUAUCGGAGGCAGCGGAGGUGGCGGAAGCGGAGGAC ACGAAUUCGGCAACCUGAUGAAGGUCCUGGAAUACGGCCUGCCUAUCACAGAGGAACACAUGCAGUUCGUGGACAGAU G U G C CG C GA UG U U U A U U U A G G U UC G G GA G G C GG CA 684 U A CG GG U U U CC G C U G G C U A C
Figure imgf000323_0001
GCCCCCAGGACCCUAUCAUCGGAACGGCGGCCGCCGUGCUGGAAAACCUCGCCACGCGCCUGCGCCCCUUUCUGCAGU GCUACCUGAAGGCCCGAGGCCUGUGCGGGCUGGACGACCUGUGCUCGCGGCGACGCCUGUCGGACAUUAAGGAUAUU GCCUCCUUUGUGUUGGUCAUCCUGGCCCGCCUCGCCAACCGCGUCGAGCGCGGCGUGUCGGAGAUCGACUACACGACC GA G U CC G A CG G G C C GG UC C A G U A G G U U U U A G A U U G C G G G U A G C U G G U A U G
Figure imgf000324_0001
GGCUCCUUUUAAGUCUGGCGCCGCAGCCGCUAAAUAUGCCGCUGCUGUUGCCGGACUGAGAGAUGCCGCUCAUAGAC UCCAGCAGUACAUGUUCUUCAUGCGGCCUGCCGAUCCUAGCAGACCCAGCACAGAUACAGCCCUGAGACUGAGCGAAC UGCUGGCCUAUGUGUCCGUGCUGUACCACUGGGCCUCUUGGAUGCUGUGGACCGCCGACAAAUACGUUGGUGGAUCU G G G A A A C A C A G C GC G C G U A CA G C G C C G G C G G G G U C CC C C G A G G A A C
Figure imgf000325_0001
GGCUCGUGGGCAGAUAUAUCCAGAGACACGGCCUGUACGUGCCCGCUCCUGAUGAACCUACACUGGCCGAUGCCAUGA AUGGCCUCGGAGGAUCUGGCGGCGGAGGAAGUGGCGGAGGACUUGCUUCUGACCCUCACUACGACUACAUCCGGCAC UAUGCCUCUGCCGCCAAACAGGCCCUGGGAGAAGUUGAACUGUCUGGCGGCCAGCUGAGCAGAGCCAUUCUGGCCCAG CU C GC G U G U AG A G G G A CU C U C C U U C C G AG C CU CC C C C AG 688 C C G C CG G C G G C C A U A
Figure imgf000326_0001
GGUGGACUGGCUGGAGGCGCGGGUGCGGGAAUGCGACUCGAUCCCGGAGAAGUUCAUCCUCAUGAUCCUCAUCGAGG GCGUCUUUUUUGCCGCCUCGUUCGCCGCCAUCGCGUACCUGCGCACCAACAACCUCCUGCGGGGCGGUUCAGGCGGC GGUGGUAGCGGCGGAAUCUCCUGUCUGCUCUACGACCUGUCCACCACCGCCCUCGAGCACAUCCUCCUGUUUUCGCUC U AC C GG G C G U G C G C C C C U A CG 689 C AU G G A U A AG G A C A UG G U G C A A A G G CA U G U U
Figure imgf000327_0001
GCUGCCUAAGCUGCUCGUGCGGGCUCCUUUUAAAUCUGGCGCCGCAGCCGCCAAAUAUGCCGCUGCUGUUGCUGGCC UGAGAGAUGCCGCUCACAGACUCCAGCAGUACAUGUUCUUCAUGCGGCCUGCCGAUCCUAGCAGACCCAGCACAGAUA CAGCCCUGAGACUGAGCGAACUGCUGGCUUAUGUGUCCGUGCUGUACCACUGGGCCUCUUGGAUGCUGUGGACCGCC U U G U A A C A C A G C GC G G C C A C A U U G A G G C A UC U C G U G CC G C G G C A C
Figure imgf000328_0001
GGCUCGUGGGCAGAUAUAUCCAGAGACACGGCCUGUACGUGCCCGCUCCUGAUGAACCUACACUGGCCGAUGCCAUGA AUGGCCUCGGAGGAUCUGGCGGCGGAGGAAGUGGCGGAGGACUUGCUUCUGACCCUCACUACGACUACAUCCGGCAC UAUGCCUCUGCCGCCAAACAGGCCCUGGGAGAAGUUGAACUGUCUGGCGGCCAGCUGAGCAGAGCCAUUCUGGCCCAG CU C GC G U G U A GC G C G U G C A A G U G UA U CG A A G C G G U
Figure imgf000329_0001
[ ] n some em o ments, a poyr onuceot e enco ng a -ce strng poypept e comprses a ribonucleic acid sequence that is at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to the ribonucleic acid sequence of any one of SEQ ID NO: 619-632, or a portion thereof. III. Polyribonucleotides A. Example Polyribonucleotides Features [0543] The present disclosure also provides combinations comprising RNA constructs comprising polyribonucleotides described herein. [0544] In some embodiments, an RNA construct provided herein comprises a polyribonucleotide that encodes one or more GP polypeptides. In some embodiments, an RNA construct provided herein comprises a polyribonucleotide that encodes an HSV-2 gC or antigenic portion thereof. In some embodiments, an RNA construct provided herein comprises a polyribonucleotide that encodes an HSV-2 gD or antigenic portion thereof. In some embodiments, an RNA construct provided herein comprises a polyribonucleotide that encodes an HSV-2 gE or antigenic portion thereof. In some embodiments, an RNA construct provided herein comprises a polyribonucleotide that encodes an HSV-2 gB, variant, or antigenic portion thereof. In some embodiments, an RNA construct provided herein comprise a polyribonucleotide that encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof. [0545] In some embodiments, an RNA construct provided herein comprises a polyribonucleotide that encodes an antigenic portion of HSV-2 gC. In some embodiments, an RNA construct provided herein comprises a polyribonucleotide that encodes an antigenic portion of HSV-2 gD. In some embodiments, an RNA construct provided herein comprises a polyribonucleotide that encodes an antigenic portion of HSV-2 gE. In some embodiments, an RNA construct provided herein comprises a polyribonucleotide that encodes an antigenic portion of HSV-2 gB. In some embodiments, an RNA construct provided herein comprise a polyribonucleotide that encodes at least one T-cell string polypeptide that comprises one or more antigenic portions of HSV T-cell antigens. [0546] In some embodiments, an RNA construct provided herein comprises a polyribonucleotide that encodes an antigenic portion of HSV-2 gC and a secretory signal. In some embodiments, an RNA construct provided herein comprises a polyribonucleotide that encodes an antigenic portion of HSV-2 gD and a secretory signal. In some embodiments, an RNA construct provided herein comprises a polyribonucleotide that encodes an antigenic portion of HSV-2 gE and a secretory signal. In some embodiments, an RNA construct provided herein comprises a polyribonucleotide that encodes an antigenic portion of HSV-2 gB and a secretory signal. In some embodiments, an RNA construct provided herein comprise a polyribonucleotide that encodes at least one T-cell string polypeptide that comprises one or more antigenic portions of HSV T-cell antigens and a secretory signal. [0547] In some embodiments, polyribonucleotides described herein can comprise a nucleotide sequence that encodes a 5’UTR and/or a 3’ UTR. In some embodiments, polynucleotides described herein can comprise a nucleotide sequence that encodes a polyA tail. In some embodiments, polyribonucleotides described herein may comprise a 5’ cap, which may be incorporated during transcription, or joined to a polyribonucleotide post- transcription. 1. 5' Cap [0548] A structural feature of RNAs is cap structure at five-prime end (5’). Natural eukaryotic RNA comprises a 7-methylguanosine cap linked to the RNA via a 5´ to 5´-triphosphate bridge resulting in cap0 structure (m7GpppN). In most eukaryotic RNA and some viral RNA, further modifications can occur at the 2'-hydroxy-group (2’-OH) (e.g., the 2'-hydroxyl group may be methylated to form 2'-O-Me) of the first and subsequent nucleotides producing “cap1” and “cap2” five-prime ends, respectively). Diamond, et al., (2014) Cytokine & growth Factor Reviews, 25:543–550, which is incorporated herein by reference in its entirety, reported that cap0-RNA cannot be translated as efficiently as cap1-RNA in which the role of 2'-O-Me in the penultimate position at the RNA 5’ end is determinant. Lack of the 2'-O-met has been shown to trigger innate immunity and activate IFN response. Daffis, et al. (2010) Nature, 468:452-456; and Züst et al. (2011) Nature Immunology, 12:137-143, each of which is incorporated herein by reference in its entirety. [0549] RNA capping is well researched and is described, e.g., in Decroly E et al. (2012) Nature Reviews 10: 51-65; and in Ramanathan A. et al., (2016) Nucleic Acids Res; 44(16): 7511–7526, the entire contents of each of which is hereby incorporated by reference. For example, in some embodiments, a 5’-cap structure which may be suitable in the context of the present invention is a cap0 (methylation of the first nucleobase, e.g., m7GpppN), cap1 (additional methylation of the ribose of the adjacent nucleotide of m7GpppN), cap2 (additional methylation of the ribose of the 2nd nucleotide downstream of the m7GpppN), cap3 (additional methylation of the ribose of the 3rd nucleotide downstream of the m7GpppN), cap4 (additional methylation of the ribose of the 4th nucleotide downstream of the m7GpppN), ARCA (“anti-reverse cap analogue”), modified ARCA (e.g. phosphothioate modified ARCA), inosine, N1 -methyl-guanosine, 2’-fluoro-guanosine, 7-deaza-guanosine, 8-oxo-guanosine, 2-amino- guanosine, LNA-guanosine, and 2-azido-guanosine. [0550] The term “5'-cap” as used herein refers to a structure found on the 5'-end of an RNA, e.g., mRNA, and generally includes a guanosine nucleotide connected to an RNA, e.g., mRNA, via a 5'- to 5'-triphosphate linkage (also referred to as Gppp or G(5')ppp(5')). In some embodiments, a guanosine nucleoside included in a 5’ cap may be modified, for example, by methylation at one or more positions (e.g., at the 7-position) on a base (guanine), and/or by methylation at one or more positions of a ribose. In some embodiments, a guanosine nucleoside included in a 5’ cap comprises a 3’O methylation at a ribose (3’OMeG). In some embodiments, a guanosine nucleoside included in a 5’ cap comprises methylation at the 7-position of guanine (m7G). In some embodiments, a guanosine nucleoside included in a 5’ cap comprises methylation at the 7-position of guanine and a 3’ O methylation at a ribose (m7(3’OMeG)). It will be understood that the notation used in the above paragraph, e.g., “(m2 7,3’-O)G” or “m7(3’OMeG)”, applies to other structures described herein. [0551] In some embodiments, providing an RNA with a 5'-cap disclosed herein may be achieved by in vitro transcription, in which a 5'-cap is co-transcriptionally expressed into an RNA strand, or may be attached to an RNA post-transcriptionally using capping enzymes. In some embodiments, co-transcriptional capping with a cap disclosed improves the capping efficiency of an RNA compared to co-transcriptional capping with an appropriate reference comparator. In some embodiments, improving capping efficiency can increase a translation efficiency and/or translation rate of an RNA, and/or increase expression of an encoded polypeptide. In some embodiments, alterations to polynucleotides generates a non-hydrolyzable cap structure which can, for example, prevent decapping and increase RNA half-life. [0552] In some embodiments, a utilized 5’ caps is a cap0, a cap1, or cap2 structure. See, e.g., Fig. 1 of Ramanathan A et al., and Fig. 1 of Decroly E et al., each of which is incorporated herein by reference in its entirety. See, e.g., Fig. 1 of Ramanathan A et al., and Fig. 1 of Decroly E et al., each of which is incorporated herein by reference in its entirety. In some embodiments, an RNA described herein comprises a cap1 structure. In some embodiments, an RNA described herein comprises a cap2. [0553] In some embodiments, an RNA described herein comprises a cap0 structure. In some embodiments, a cap0 structure comprises a guanosine nucleoside methylated at the 7-position of guanine ((m7)G). In some embodiments, such a cap0 structure is connected to an RNA via a 5'- to 5'-triphosphate linkage and is also referred to herein as (m7)Gppp. In some embodiments, a cap0 structure comprises a guanosine nucleoside methylated at the 2’-position of the ribose of guanosine. In some embodiments, a cap0 structure comprises a guanosine nucleoside methylated at the 3’-position of the ribose of guanosine. In some embodiments, a guanosine nucleoside included in a 5’ cap comprises methylation at the 7-position of guanine and at the 2’-position of the ribose ((m2 7,2’-O)G). In some embodiments, a guanosine nucleoside included in a 5’ cap comprises methylation at the 7- position of guanine and at the 2’-position of the ribose ((m2 7,3’-O)G). [0554] In some embodiments, a cap1 structure comprises a guanosine nucleoside methylated at the 7- position of guanine ((m7)G) and optionally methylated at the 2’ or 3’ position pf the ribose, and a 2’O methylated first nucleotide in an RNA ((m2’-O)N1). In some embodiments, a cap1 structure comprises a guanosine nucleoside methylated at the 7-position of guanine ((m7)G) and the 3’ position of the ribose, and a 2’O methylated first nucleotide in an RNA
Figure imgf000332_0001
1). In some embodiments, a cap1 structure is connected to an RNA via a 5'- to 5'- triphosphate linkage and is also referred to herein as, e.g., ((m7)Gppp(2'-O)N1) or (m2 7,3’-O)Gppp(2'-O)N1), wherein N1 is as defined and described herein. In some embodiments, a cap1 structure comprises a second nucleotide, N2, which is at position 2 and is chosen from A, G, C, or U, e.g., (m7)Gppp(2'-O)N1pN2 or (m2 7,3’-O)Gppp(2'-O)N1pN2 , wherein each of N1 and N2 is as defined and described herein. [0555] In some embodiments, a cap2 structure comprises a guanosine nucleoside methylated at the 7- position of guanine ((m7)G) and optionally methylated at the 2’ or 3’ position of the ribose, and a 2’O methylated first and second nucleotides in an RNA ((m2’-O)N1p(m2’-O)N2). In some embodiments, a cap2 structure comprises a guanosine nucleoside methylated at the 7-position of guanine ((m7)G) and the 3’ position of the ribose, and a 2’O methylated first and second nucleotide in an RNA. In some embodiments, a cap2 structure is connected to an RNA via a 5'- to 5'-triphosphate linkage and is also referred to herein as, e.g., ((m7)Gppp(2'-O)N1p(2'-O)N2) or (m2 7,3’- O)Gppp(2'-O)N1p(2'-O)N2), wherein each of N1 and N2 is as defined and described herein. [0556] In some embodiments, the 5’ cap is a dinucleotide cap structure. In some embodiments, the 5’ cap is a dinucleotide cap structure comprising N1, wherein N1 is as defined and described herein. In some embodiments, the 5’ cap is a dinucleotide cap G*N1, wherein N1 is as defined above and herein, and G* comprises a structure of formula (I): or a salt thereof,
Figure imgf000332_0002
wherein each R2 and R3 is -OH or -OCH3; and X is O or S. [0557] In some embodiments, R2 is -OH. In some embodiments, R2 is -OCH3. In some embodiments, R3 is -OH. In some embodiments, R3 is -OCH3. In some embodiments, R2 is -OH and R3 is -OH. In some embodiments, R2 is -OH and R3 is -CH3. In some embodiments, R2 is -CH3 and R3 is -OH. In some embodiments, R2 is -CH3 and R3 is - CH3. [0558] In some embodiments, X is O. In some embodiments, X is S. [0559] In some embodiments, the 5’ cap is a dinucleotide cap0 structure (e.g., (m7)GpppN1, (m2 7,2’- O)GpppN1, (m2 7,3’-O)GpppN1, (m7)GppSpN1, (m2 7,2’-O)GppSpN1, or (m2 7,3’-O)GppSpN1), wherein N1 is as defined and described herein. In some embodiments, the 5’ cap is a dinucleotide cap0 structure (e.g., (m7)GpppN1, (m2 7,2’- O)GpppN1, (m2 7,3’-O)GpppN1, (m7)GppSpN1, (m2 7,2’-O)GppSpN1, or (m2 7,3’-O)GppSpN1), wherein N1 is G. In some embodiments, the 5’ cap is a dinucleotide cap0 structure (e.g., (m7)GpppN1, (m2 7,2’-O)GpppN1, (m2 7,3’-O)GpppN1, (m7)GppSpN1, (m2 7,2’-O)GppSpN1, or (m2 7,3’-O)GppSpN1), wherein N1 is A, U, or C. In some embodiments, the 5’ cap is a dinucleotide cap1 structure (e.g., (m7)Gppp(m2’-O)N1, (m2 7,2’-O)Gppp(m2’-O)N1, (m2 7,3’-O)Gppp(m2’-O)N1, (m7)GppSp(m2’-O)N1, (m2 7,2’-O)GppSp(m2’-O)N1, or (m2 7,3’-O)GppSp(m2’-O)N1), wherein N1 is as defined and described herein. In some embodiments, the 5’ cap is selected from the group consisting of (m7)GpppG (“Ecap0”), (m7)Gppp(m2’-O)G (“Ecap1”), (m2 7,3’-O)GpppG (“ARCA” or “D1”), and (m2 7,2’-O)GppSpG (“beta-S-ARCA”). In some embodiments, the 5’ cap is (m7)GpppG (“Ecap0”), having a structure: or a salt thereof.
Figure imgf000333_0001
[0560] In some embodiments, the 5’ cap is (m7)Gppp(m2’-O)G (“Ecap1”), having a structure: or a salt thereof.
Figure imgf000333_0002
[0561] In some embodiments, the 5’ cap is (m2 7,3’-O)GpppG (“ARCA” or “D1”), having a structure: or a salt thereof.
Figure imgf000334_0001
[0562] In some embodiments, the 5’ cap is (m27,2’-O)GppSpG (“beta-S-ARCA”), having a structure: or a salt thereof.
Figure imgf000334_0002
[0563] In some embodiments, the 5’ cap is a trinucleotide cap structure. In some embodiments, the 5’ cap is a trinucleotide cap structure comprising N1pN2, wherein N1 and N2 are as defined and described herein. In some embodiments, the 5’ cap is a dinucleotide cap G*N1pN2, wherein N1 and N2 are as defined above and herein, and G* comprises a structure of formula (I): or a salt thereof, wherein R2,
Figure imgf000334_0003
, . [0564] In some embodiments, the 5’ cap is a trinucleotide cap0 structure (e.g. (m7)GpppN1pN2, (m2 7,2’- O)GpppN1pN2, or (m2 7,3’-O)GpppN1pN2), wherein N1 and N2 are as defined and described herein). In some embodiments, the 5’ cap is a trinucleotide cap1 structure (e.g., (m7)Gppp(m2’-O)N1pN2, (m2 7,2’-O)Gppp(m2’-O)N1pN2, (m2 7,3’-O)Gppp(m2’-O)N1pN2), wherein N1 and N2 are as defined and described herein. In some embodiments, the 5’ cap is a trinucleotide cap2 structure (e.g., (m7)Gppp(m2’-O)N1p(m2’-O)N2, (m2 7,2’-O)Gppp(m2’-O)N1p(m2’-O)N2, (m2 7,3’- O)Gppp(m2’-O)N1p(m2’-O)N2), wherein N1 and N2 are as defined and described herein. In some embodiments, the 5’ cap is selected from the group consisting of (m2 7,3’-O)Gppp(m2’-O)ApG (“CleanCap AG”, “CC413”), (m2 7,3’-O)Gppp(m2’- O)GpG (“CleanCap GG”), (m7)Gppp(m2’-O)ApG, (m7)Gppp(m2’-O)GpG, (m2 7,3’-O)Gppp(m2 6,2’-O)ApG, and (m7)Gppp(m2’- O)ApU. [0565] In some embodiments, the 5’ cap is (m2 7,3’-O)Gppp(m2’-O)ApG (“CleanCap AG”, “CC413”), having a structure: or a salt there
Figure imgf000335_0001
[0566] In some embodiments, the 5’ cap is (m2 7,3’-O)Gppp(m2’-O)GpG (“CleanCap GG”), having a structure: or a salt thereo
Figure imgf000335_0002
f. [0567] In some embodiments, the 5’ cap is (m7)Gppp(m2’-O)ApG, having a structure:
or a salt there
Figure imgf000336_0001
[0568] In some embodiments, the 5’ cap is (m7)Gppp(m2’-O)GpG, having a structure: or a salt there
Figure imgf000336_0002
. [0569] In some embodiments, the 5’ cap is (m2 7,3’-O)Gppp(m2 6,2’-O)ApG, having a structure:
or a salt there
Figure imgf000337_0001
[0570] In some embodiments, the 5’ cap is (m7)Gppp(m2’-O)ApU, having a structure: or a salt thereof.
Figure imgf000337_0002
[0571] In some embodiments, the 5’ cap is a tetranucleotide cap structure. In some embodiments, the 5’ cap is a tetranucleotide cap structure comprising N1pN2pN3, wherein N1, N2, and N3 are as defined and described herein. In some embodiments, the 5’ cap is a tetranucleotide cap G*N1pN2pN3, wherein N1, N2, and N3 are as defined above and herein, and G* comprises a structure of formula (I): or a salt thereof, wherein R2,
Figure imgf000338_0001
[0572] In some embodiments, the 5’ cap is a tetranucleotide cap0 structure (e.g. (m7)GpppN1pN2pN3, (m2 7,2’-O)GpppN1pN2pN3, or (m2 7,3’-O)GpppN1N2pN3), wherein N1, N2, and N3 are as defined and described herein). In some embodiments, the 5’ cap is a tetranucleotide Cap1 structure (e.g., (m7)Gppp(m2’-O)N1pN2pN3, (m2 7,2’- O)Gppp(m2’-O)N1pN2pN3, (m2 7,3’-O)Gppp(m2’-O)N1pN2N3), wherein N1, N2, and N3 are as defined and described herein. In some embodiments, the 5’ cap is a tetranucleotide Cap2 structure (e.g., (m7)Gppp(m2’-O)N1p(m2’-O)N2pN3, (m2 7,2’- O)Gppp(m2’-O)N1p(m2’-O)N2pN3, (m2 7,3’-O)Gppp(m2’-O)N1p(m2’-O)N2pN3), wherein N1, N2, and N3 are as defined and described herein. In some embodiments, the 5’ cap is selected from the group consisting of (m2 7,3’-O)Gppp(m2’- O)Ap(m2’-O)GpG, (m2 7,3’-O)Gppp(m2’-O)Gp(m2’-O)GpC, (m7)Gppp(m2’-O)Ap(m2’-O)UpA, and (m7)Gppp(m2’-O)Ap(m2’-O)GpG. [0573] In some embodiments, the 5’ cap is (m2 7,3’-O)Gppp(m2’-O)Ap(m2’-O)GpG, having a structure: or a salt thereof.
Figure imgf000338_0002
[0574] In some embodiments, the 5’ cap is (m2 7,3’-O)Gppp(m2’-O)Gp(m2’-O)GpC, having a structure:
or a salt there
Figure imgf000339_0001
[0575] In some embodiments, the 5’ cap is (m7)Gppp(m2’-O)Ap(m2’-O)UpA, having a structure: or a salt thereo
Figure imgf000339_0002
f. [0576] In some embodiments, the 5’ cap is (m7)Gppp(m2’-O)Ap(m2’-O)GpG, having a structure:
Figure imgf000340_0001
comprises a cap proximal sequence, e.g., as disclosed herein. In some embodiments, a cap proximal sequence comprises a sequence adjacent to a 5’ cap. In some embodiments, a cap proximal sequence comprises nucleotides in positions +1, +2, +3, +4, and/or +5 of an RNA polynucleotide. [0578] In some embodiments, a cap structure comprises one or more polynucleotides of a cap proximal sequence. In some embodiments, a cap structure comprises an m7 Guanosine cap and nucleotide +1 (N1) of an RNA polynucleotide. In some embodiments, a cap structure comprises an m7 Guanosine cap and nucleotide +2 (N2) of an RNA polynucleotide. In some embodiments, a cap structure comprises an m7 Guanosine cap and nucleotides +1 and +2 (N1 and N2) of an RNA polynucleotide. In some embodiments, a cap structure comprises an m7 Guanosine cap and nucleotides +1, +2, and +3 (N1, N2, and N3) of an RNA polynucleotide. [0579] Those skilled in the art, reading the present disclosure, will appreciate that, in some embodiments, one or more residues of a cap proximal sequence (e.g., one or more of residues +1, +2, +3, +4, and/or +5) may be included in an RNA by virtue of having been included in a cap entity (e.g., a cap1 or cap2 structure, etc.); alternatively, in some embodiments, at least some of the residues in a cap proximal sequence may be enzymatically added (e.g., by a polymerase such as a T7 polymerase). For example, in certain exemplified embodiments where a m2 7,3’-OGppp(m1 2’-O)ApG cap is utilized, +1 (i.e., N1) and +2 (i.e. N2) are the (m1 2’-O)A and G residues of the cap, and +3, +4, and +5 are added by polymerase (e.g., T7 polymerase). [0580] In some embodiments, the 5’ cap is a dinucleotide cap structure, wherein the cap proximal sequence comprises N1 of the 5’ cap, where N1 is any nucleotide, e.g., A, C, G or U. In some embodiments, the 5’ cap is a trinucleotide cap structure (e.g., the trinucleotide cap structures described above and herein), wherein the cap proximal sequence comprises N1 and N2 of the 5’ cap, wherein N1 and N2 are independently any nucleotide, e.g., A, C, G or U. In some embodiments, the 5’ cap is a tetranucleotide cap structure (e.g., the trinucleotide cap structures described above and herein), wherein the cap proximal sequence comprises N1, N2, and N3 of the 5’ cap, wherein N1, N2, and N3 are any nucleotide, e.g., A, C, G or U. [0581] In some embodiments, e.g., where the 5’ cap is a dinucleotide cap structure, a cap proximal sequence comprises N1 of a the 5’ cap, and N2, N3, N4 and N5, wherein N1 to N5 correspond to positions +1, +2, +3, +4, and/or +5 of an RNA polynucleotide. In some embodiments, e.g., where the 5’ cap is a trinucleotide cap structure, a cap proximal sequence comprises N1 and N2 of a the 5’ cap, and N3, N4 and N5, wherein N1 to N5 correspond to positions +1, +2, +3, +4, and/or +5 of an RNA polynucleotide. In some embodiments, e.g., where the 5’ cap is a tetranucleotide cap structure, a cap proximal sequence comprises N1, N2, and N3 of a the 5’ cap, and N4 and N5, wherein N1 to N5 correspond to positions +1, +2, +3, +4, and/or +5 of an RNA polynucleotide. [0582] In some embodiments, N1 is A. In some embodiments, N1 is C. In some embodiments, N1 is G. In some embodiments, N1 is U. In some embodiments, N2 is A. In some embodiments, N2 is C. In some embodiments, N2 is G. In some embodiments, N2 is U. In some embodiments, N3 is A. In some embodiments, N3 is C. In some embodiments, N3 is G. In some embodiments, N3 is U. In some embodiments, N4 is A. In some embodiments, N4 is C. In some embodiments, N4 is G. In some embodiments, N4 is U. In some embodiments, N5 is A. In some embodiments, N5 is C. In some embodiments, N5 is G. In some embodiments, N5 is U. It will be understood that each of the embodiments described above and herein (e.g., for N1 through N5) may be taken singly or in combination and/or may be combined with other embodiments of variables described above and herein (e.g., 5’ caps). [0583] In some embodiments, a cap proximal sequence comprises A1 and G2 of the Cap1 structure, and a sequence comprising: A3A4U5 (SEQ ID NO: 150) at positions +3, +4 and +5 respectively of the polyribonucleotide. 3. 5’ UTR [0584] In some embodiments, a nucleic acid (e.g., DNA, RNA) utilized in accordance with the present disclosure comprises a 5'-UTR. In some embodiments, 5’-UTR may comprise a plurality of distinct sequence elements; in some embodiments, such plurality may be or comprise multiple copies of one or more particular sequence elements (e.g., as may be from a particular source or otherwise known as a functional or characteristic sequence element). In some embodiments, a 5’ UTR comprises multiple different sequence elements. [0585] The term “untranslated region” or “UTR” is commonly used in the art to a region in a DNA molecule which is transcribed but is not translated into an amino acid sequence, or to the corresponding region in an RNA polynucleotide, such as an mRNA molecule. An untranslated region (UTR) can be present 5' (upstream) of an open reading frame (5'-UTR) and/or 3' (downstream) of an open reading frame (3'-UTR). As used herein, the terms “five prime untranslated region” or “5' UTR” refer to a sequence of a polyribonucleotide between the 5' end of the polyribonucleotide (e.g., a transcription start site) and a start codon of a coding region of the polyribonucleotide. In some embodiments, “5' UTR” refers to a sequence of a polyribonucleotide that begins at the 5' end of the polyribonucleotide (e.g., a transcription start site) and ends one nucleotide (nt) before a start codon (usually AUG) of a coding region of the polyribonucleotide, e.g., in its natural context. In some embodiments, a 5' UTR comprises a Kozak sequence. A 5'-UTR is downstream of the 5'-cap (if present), e.g., directly adjacent to the 5'-cap. In some embodiments, a 5’ UTR disclosed herein comprises a cap proximal sequence, e.g., as defined and described herein. In some embodiments, a cap proximal sequence comprises a sequence adjacent to a 5’ cap. [0586] Example 5’ UTRs include a human alpha globin (hAg) 5’UTR or a portion thereof, a TEV 5’ UTR or a portion thereof, a HSP705’ UTR or a portion thereof, or a c-Jun 5’ UTR or a portion thereof. [0587] In some embodiments, an RNA disclosed herein comprises a hAg 5’ UTR or a portion thereof. [0588] In some embodiments, an RNA disclosed herein comprises a 5’ UTR having at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to a 5’ UTR with the sequence AGAATAAACTAGTATTCTTCTGGTCCCCACAGACTCAGAGAGAACCCGCCACC (SEQ ID NO: 151). In some embodiments, an RNA disclosed herein comprises a 5’ UTR having the sequence AGAATAAACTAGTATTCTTCTGGTCCCCACAGACTCAGAGAGAACCCGCCACC (SEQ ID NO: 151). [0589] In some embodiments, an RNA disclosed herein comprises a 5’ UTR having at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to a 5’ UTR with the sequence AACUAGUAUUCUUCUGGUCCCCACAGACUCAGAGAGAACCCGCCACC (SEQ ID NO: 152)(hAg- Kozak/5'UTR). In some embodiments, an RNA disclosed herein comprises a 5’ UTR having the sequence AACUAGUAUUCUUCUGGUCCCCACAGACUCAGAGAGAACCCGCCACC (SEQ ID NO: 152)(hAg-Kozak/5'UTR). 4. PolyA Tail [0590] In some embodiments, a polynucleotide (e.g., DNA, RNA) disclosed herein comprises a polyadenylate (polyA) sequence, e.g., as described herein. In some embodiments, a polyA sequence is situated downstream of a 3'-UTR, e.g., adjacent to a 3'-UTR. [0591] As used herein, the term “poly(A) sequence” or “poly-A tail” refers to an uninterrupted or interrupted sequence of adenylate residues which is typically located at the 3'-end of an RNA polynucleotide. Poly(A) sequences are known to those of skill in the art and may follow the 3’-UTR in the RNAs described herein. An uninterrupted poly(A) sequence is characterized by consecutive adenylate residues. In nature, an uninterrupted poly(A) sequence is typical. In some embodiments, polynucleotides disclosed herein comprise an uninterrupted Poly(A) sequence. In some embodiments, polynucleotides disclosed herein comprise interrupted Poly(A) sequence. In some embodiments, RNAs disclosed herein can have a poly(A) sequence attached to the free 3'-end of the RNA by a template-independent RNA polymerase after transcription or a poly(A) sequence encoded by DNA and transcribed by a template-dependent RNA polymerase. [0592] It has been demonstrated that a poly(A) sequence (SEQ ID NO: 829) of about 120 A nucleotides has a beneficial influence on the levels of RNA in transfected eukaryotic cells, as well as on the levels of protein that is translated from an open reading frame that is present upstream (5’) of the poly(A) sequence (Holtkamp et al., 2006, Blood, vol. 108, pp. 4009-4017, which is herein incorporated by reference). [0593] In some embodiments, a poly(A) sequence in accordance with the present disclosure is not limited to a particular length; in some embodiments, a poly(A) sequence is any length. In some embodiments, a poly(A) sequence (SEQ ID NO: 829) comprises, essentially consists of, or consists of at least 20, at least 30, at least 40, at least 80, or at least 100 and up to 500, up to 400, up to 300, up to 200, or up to 150 A nucleotides, and, in particular, about 120 A nucleotides. In this context, "essentially consists of" means that most nucleotides in the poly(A) sequence, typically at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% by number of nucleotides in the poly(A) sequence are A nucleotides, but permits that remaining nucleotides are nucleotides other than A nucleotides, such as U nucleotides (uridylate), G nucleotides (guanylate), or C nucleotides (cytidylate). In this context, "consists of" means that all nucleotides in the poly(A) sequence, i.e., 100% by number of nucleotides in the poly(A) sequence, are A nucleotides. The term “A nucleotide” or “A” refers to adenylate. [0594] In some embodiments, a poly(A) sequence is attached during RNA transcription, e.g., during preparation of in vitro transcribed RNA, based on a DNA template comprising repeated dT nucleotides (deoxythymidylate) in the strand complementary to the coding strand. The DNA sequence encoding a poly(A) sequence (coding strand) is referred to as poly(A) cassette. [0595] In some embodiments, the poly(A) cassette present in the coding strand of DNA essentially consists of dA nucleotides, but is interrupted by a random sequence of the four nucleotides (dA, dC, dG, and dT). Such random sequence may be 5 to 50, 10 to 30, or 10 to 20 nucleotides in length. Such a cassette is disclosed in WO 2016/005324 A1, hereby incorporated by reference. Any poly(A) cassette disclosed in WO 2016/005324 A1, which is incorporated herein by reference in its entirety, may be used in accordance with the present disclosure. A poly(A) cassette that essentially consists of dA nucleotides, but is interrupted by a random sequence having an equal distribution of the four nucleotides (dA, dC, dG, dT) and having a length of e.g., 5 to 50 nucleotides shows, on DNA level, constant propagation of plasmid DNA in E. coli and is still associated, on RNA level, with the beneficial properties with respect to supporting RNA stability and translational efficiency is encompassed. In some embodiments, the poly(A) sequence contained in an RNA polynucleotide described herein essentially consists of A nucleotides, but is interrupted by a random sequence of the four nucleotides (A, C, G, U). Such random sequence may be 5 to 50, 10 to 30, or 10 to 20 nucleotides in length. [0596] In some embodiments, no nucleotides other than A nucleotides flank a poly(A) sequence at its 3'- end, i.e., the poly(A) sequence is not masked or followed at its 3'-end by a nucleotide other than A. [0597] In some embodiments, the poly(A) sequence (SEQ ID NO: 829) may comprise at least 20, at least 30, at least 40, at least 80, or at least 100 and up to 500, up to 400, up to 300, up to 200, or up to 150 nucleotides. In some embodiments, the poly(A) sequence (SEQ ID NO: 829) may essentially consist of at least 20, at least 30, at least 40, at least 80, or at least 100 and up to 500, up to 400, up to 300, up to 200, or up to 150 nucleotides. In some embodiments, the poly(A) sequence (SEQ ID NO: 829) may consist of at least 20, at least 30, at least 40, at least 80, or at least 100 and up to 500, up to 400, up to 300, up to 200, or up to 150 nucleotides. In some embodiments, the poly(A) sequence (SEQ ID NO: 829) comprises at least 100 nucleotides. In some embodiments, the poly(A) sequence (SEQ ID NO: 829) comprises about 150 nucleotides. In some embodiments, the poly(A) sequence comprises about 120 nucleotides. [0598] In some embodiments, a poly A tail comprises a specific number of Adenosines, such as about 50 or more, about 60 or more, about 70 or more, about 80 or more, about 90 or more, about 100 or more, about 120, or about 150 or about 200. In some embodiments a poly A tail of a string construct may comprise 200 A residues or less. In some embodiments, a poly A tail of a string construct may comprise about 200 A residues. In some embodiments, a poly A tail of a string construct may comprise 180 A residues or less. In some embodiments, a poly A tail of a string construct may comprise about 180 A residues. In some embodiments, a poly A tail may comprise 150 residues or less. [0599] In some embodiments, RNA comprises a poly(A) sequence comprising the nucleotide sequence of AAAAAAAAAAAAAAAAAAAAAAAAAAAAAAGCATATGACTAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAA AAAAAAAAAAAAAAAAAAAAAAAA (SEQ ID NO: 153), or a nucleotide sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the nucleotide sequence of AAAAAAAAAAAAAAAAAAAAAAAAAAAAAAGCATATGACTAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAA AAAAAAAAAAAAAAAAAAAAAAAA (SEQ ID NO: 153). In some embodiments, a poly(A) tail comprises a plurality of A residues interrupted by a linker. In some embodiments, a linker comprises the nucleotide sequence GCATATGAC (SEQ ID NO: 154). [0600] In some embodiments, RNA comprises a poly(A) sequence comprising the nucleotide sequence of AAAAAAAAAAAAAAAAAAAAAAAAAAAAAAGCAUAUGACUAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAA AAAAAAAAAAAAAAAAAAAAAAAA (SEQ ID NO: 155), or a nucleotide sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the nucleotide sequence of AAAAAAAAAAAAAAAAAAAAAAAAAAAAAAGCAUAUGACUAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAA AAAAAAAAAAAAAAAAAAAAAAAA (SEQ ID NO: 155). In some embodiments, a poly(A) tail comprises a plurality of A residues interrupted by a linker. In some embodiments, a linker comprises the nucleotide sequence GCAUAUGAC (SEQ ID NO: 156). 5. 3' UTR [0601] In some embodiments, an RNA utilized in accordance with the present disclosure comprises a 3'- UTR. As used herein, the terms “three prime untranslated region,” “3' untranslated region,” or “3' UTR” refer to a sequence of an RNA molecule that begins following a stop codon of a coding region of an open reading frame sequence. In some embodiments, the 3' UTR begins immediately after a stop codon of a coding region of an open reading frame sequence, e.g., in its natural context. In other embodiments, the 3' UTR does not begin immediately after stop codon of the coding region of an open reading frame sequence, e.g., in its natural context. The term “3'- UTR” does preferably not include the poly(A) sequence. Thus, the 3'-UTR is upstream of the poly(A) sequence (if present), e.g., directly adjacent to the poly(A) sequence. [0602] In some embodiments, an RNA disclosed herein comprises a 3’ UTR comprising an F element d/ I l I b di 3’ UTR i l h i i i i e. In
Figure imgf000344_0001
, . , element sequence is a 3’-UTR of amino-terminal enhancer of split (AES). [0604] In some embodiments, an RNA disclosed herein comprises a 3’ UTR having at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to a 3’ UTR with the sequence of CTGGTACTGCATGCACGCAATGCTAGCTGCCCCTTTCCCGTCCTGGGTACCCCGAGTCTCCCCCGACCTCGGGTCCCAGGTATG CTCCCACCTCCACCTGCCCCACTCACCACCTCTGCTAGTTCCAGACACCTCCCAAGCACGCAGCAATGCAGCTCAAAACGCTTAG CCTAGCCACACCCCCACGGGAAACAGCAGTGATTAACCTTTAGCAATAAACGAAAGTTTAACTAAGCTATACTAACCCCAGGGTT GGTCAATTTCGTGCCAGCCACACC (SEQ ID NO: 157). In some embodiments, an RNA disclosed herein comprises a 3’ UTR with the sequence of CTGGTACTGCATGCACGCAATGCTAGCTGCCCCTTTCCCGTCCTGGGTACCCCGAGTCTCCCCCGACCTCGGGTCCCAGGTATG CTCCCACCTCCACCTGCCCCACTCACCACCTCTGCTAGTTCCAGACACCTCCCAAGCACGCAGCAATGCAGCTCAAAACGCTTAG CCTAGCCACACCCCCACGGGAAACAGCAGTGATTAACCTTTAGCAATAAACGAAAGTTTAACTAAGCTATACTAACCCCAGGGTT GGTCAATTTCGTGCCAGCCACACC (SEQ ID NO: 157). [0605] In some embodiments, an RNA disclosed herein comprises a 3’ UTR having at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to a 3’ UTR with the sequence of CUGGUACUGCAUGCACGCAAUGCUAGCUGCCCCUUUCCCGUCCUGGGUACCCCGAGUCUCCCCCGACCUCGGGUCCCAGGUA UGCUCCCACCUCCACCUGCCCCACUCACCACCUCUGCUAGUUCCAGACACCUCCCAAGCACGCAGCAAUGCAGCUCAAAACGCU UAGCCUAGCCACACCCCCACGGGAAACAGCAGUGAUUAACCUUUAGCAAUAAACGAAAGUUUAACUAAGCUAUACUAACCCCA GGGUUGGUCAAUUUCGUGCCAGCCACACC (SEQ ID NO: 158). In some embodiments, an RNA disclosed herein comprises a 3’ UTR with the sequence of CUGGUACUGCAUGCACGCAAUGCUAGCUGCCCCUUUCCCGUCCUGGGUACCCCGAGUCUCCCCCGACCUCGGGUCCCAGGUA UGCUCCCACCUCCACCUGCCCCACUCACCACCUCUGCUAGUUCCAGACACCUCCCAAGCACGCAGCAAUGCAGCUCAAAACGCU UAGCCUAGCCACACCCCCACGGGAAACAGCAGUGAUUAACCUUUAGCAAUAAACGAAAGUUUAACUAAGCUAUACUAACCCCA GGGUUGGUCAAUUUCGUGCCAGCCACACC (SEQ ID NO: 158). [0606] In some embodiments, a 3’UTR is an FI element as described in WO2017/060314, which is herein incorporated by reference in its entirety. B. RNA Formats [0607] At least three distinct formats useful for RNA compositions (e.g., pharmaceutical compositions) have been developed, namely non-modified uridine containing RNA (uRNA), nucleoside-modified mRNA (modRNA), and self-amplifying RNA (saRNA). Each of these platforms displays unique features. In general, in all three formats, RNA is capped, contains open reading frames (ORFs) flanked by untranslated regions (UTR), and have a polyA-tail at the 3' end. An ORF of an uRNA and modRNA vectors encode an antibody agent or portion thereof. An saRNA has multiple ORFs. [0608] In some embodiments, the RNA described herein may have modified nucleosides. In some embodiments, the RNA comprises a modified nucleoside in place of at least one (e.g., every) uridine. [0609] The term “uracil,” as used herein, describes one of the nucleobases that can occur in the nucleic acid of RNA. The structure of uracil is: . [0610] The term “uridine,” as used her one of the nucleosides that can occur in RNA. The
Figure imgf000346_0001
structure of uridine is: . [0611] UTP (uridine 5’-triphosphate)
Figure imgf000346_0002
as t e oowng structure: . [0612] Pseudo-UTP (pseudo
Figure imgf000346_0003
owing structure: . [0613] “Pseudouridine”
Figure imgf000346_0004
s o e exa pe o a o e uceos e at is an isomer of uridine, where the uracil is attached to the pentose ring via a carbon-carbon bond instead of a nitrogen-carbon glycosidic bond. [0614] Another exemplary modified nucleoside is N1-methyl-pseudouridine (m1Ψ), which has the structure: .
Figure imgf000347_0001
[0615] N1-methyl-pseudo-UTP has the following structure: . [0616] Another exempl
Figure imgf000347_0002
m5U), which has the structure: . [0617] In some embodiments, on
Figure imgf000347_0003
A described herein is replaced by a modified nucleoside. In some embodiments, the modified nucleoside is a modified uridine. [0618] In some embodiments, RNA comprises a modified nucleoside in place of at least one uridine. In some embodiments, RNA comprises a modified nucleoside in place of each uridine. [0619] In some embodiments, the modified nucleoside is independently selected from pseudouridine (ψ), N1-methyl-pseudouridine (m1ψ), and 5-methyl-uridine (m5U). In some embodiments, the modified nucleoside comprises pseudouridine (ψ). In some embodiments, the modified nucleoside comprises N1-methyl-pseudouridine (m1ψ). In some embodiments, the modified nucleoside comprises 5-methyl-uridine (m5U). In some embodiments, RNA may comprise more than one type of modified nucleoside, and the modified nucleosides are independently selected from pseudouridine (ψ), N1-methyl-pseudouridine (m1ψ), and 5-methyl-uridine (m5U). In some embodiments, the modified nucleosides comprise pseudouridine (ψ) and N1-methyl-pseudouridine (m1ψ). In some embodiments, the modified nucleosides comprise pseudouridine (ψ) and 5-methyl-uridine (m5U). In some embodiments, the modified nucleosides comprise N1-methyl-pseudouridine (m1ψ) and 5-methyl-uridine (m5U). In some embodiments, the modified nucleosides comprise pseudouridine (ψ), N1-methyl-pseudouridine (m1ψ), and 5- methyl-uridine (m5U). [0620] In some embodiments, the modified nucleoside replacing one or more, e.g., all, uridine in the RNA may be any one or more of 3-methyl-uridine (m3U), 5-methoxy-uridine (mo5U), 5-aza-uridine, 6-aza-uridine, 2-thio- 5-aza-uridine, 2-thio-uridine (s2U), 4-thio-uridine (s4U), 4-thio-pseudouridine, 2-thio-pseudouridine, 5-hydroxy- uridine (ho5U), 5-aminoallyl-uridine, 5-halo-uridine (e.g., 5-iodo-uridine or 5-bromo-uridine), uridine 5-oxyacetic acid (cmo5U), uridine 5-oxyacetic acid methyl ester (mcmo5U), 5-carboxymethyl-uridine (cm5U), 1-carboxymethyl- pseudouridine, 5-carboxyhydroxymethyl-uridine (chm5U), 5-carboxyhydroxymethyl-uridine methyl ester (mchm5U), 5-methoxycarbonylmethyl-uridine (mcm5U), 5-methoxycarbonylmethyl-2-thio-uridine (mcm5s2U), 5-aminomethyl-2- thio-uridine (nm5s2U), 5-methylaminomethyl-uridine (mnm5U), 1-ethyl-pseudouridine, 5-methylaminomethyl-2-thio- uridine (mnm5s2U), 5-methylaminomethyl-2-seleno-uridine (mnm5se2U), 5-carbamoylmethyl-uridine (ncm5U), 5- carboxymethylaminomethyl-uridine (cmnm5U), 5-carboxymethylaminomethyl-2-thio-uridine (cmnm5s2U), 5- propynyl-uridine, 1-propynyl-pseudouridine, 5-taurinomethyl-uridine (τm5U), 1-taurinomethyl-pseudouridine, 5- taurinomethyl-2-thio-uridine(τm5s2U), 1-taurinomethyl-4-thio-pseudouridine), 5-methyl-2-thio-uridine (m5s2U), 1- methyl-4-thio-pseudouridine (m1s4ψ), 4-thio-1-methyl-pseudouridine, 3-methyl-pseudouridine (m3ψ), 2-thio-1- methyl-pseudouridine, 1-methyl-1-deaza-pseudouridine, 2-thio-1-methyl-1-deaza-pseudouridine, dihydrouridine (D), dihydropseudouridine, 5,6-dihydrouridine, 5-methyl-dihydrouridine (m5D), 2-thio-dihydrouridine, 2-thio- dihydropseudouridine, 2-methoxy-uridine, 2-methoxy-4-thio-uridine, 4-methoxy-pseudouridine, 4-methoxy-2-thio- pseudouridine, N1-methyl-pseudouridine, 3-(3-amino-3-carboxypropyl)uridine (acp3U), 1-methyl-3-(3-amino-3- carboxypropyl)pseudouridine (acp3 ψ), 5-(isopentenylaminomethyl)uridine (inm5U), 5-(isopentenylaminomethyl)-2- thio-uridine (inm5s2U), α-thio-uridine, 2′-O-methyl-uridine (Um), 5,2′-O-dimethyl-uridine (m5Um), 2′-O-methyl- pseudouridine (ψm), 2-thio-2′-O-methyl-uridine (s2Um), 5-methoxycarbonylmethyl-2′-O-methyl-uridine (mcm5Um), 5-carbamoylmethyl-2′-O-methyl-uridine (ncm5Um), 5-carboxymethylaminomethyl-2′-O-methyl-uridine (cmnm5Um), 3,2′-O-dimethyl-uridine (m3Um), 5-(isopentenylaminomethyl)-2′-O-methyl-uridine (inm5Um), 1-thio-uridine, deoxythymidine, 2′-F-ara-uridine, 2′-F-uridine, 2′-OH-ara-uridine, 5-(2-carbomethoxyvinyl) uridine, 5-[3-(1-E- propenylamino)uridine, or any other modified uridine known in the art. [0621] In some embodiments, the RNA comprises other modified nucleosides or comprises further modified nucleosides, e.g., modified cytidine. For example, in some embodiments, in the RNA 5-methylcytidine is substituted partially or completely, preferably completely, for cytidine. In some embodiments, the RNA comprises 5- methylcytidine and one or more selected from pseudouridine (ψ), N1-methyl-pseudouridine (m1ψ), and 5-methyl- uridine (m5U). In some embodiments, the RNA comprises 5-methylcytidine and N1-methyl-pseudouridine (m1ψ). In some embodiments, the RNA comprises 5-methylcytidine in place of each cytidine and N1-methyl-pseudouridine (m1ψ) in place of each uridine. [0622] In some embodiments of the present disclosure, the RNA is “replicon RNA” or simply a “replicon,” in particular “self-replicating RNA” or “self-amplifying RNA.” In one particularly preferred embodiment, the replicon or self-replicating RNA is derived from or comprises elements derived from a single-stranded (ss) RNA virus, in particular a positive-stranded ssRNA virus, such as an alphavirus. Alphaviruses are typical representatives of positive- stranded RNA viruses. Alphaviruses replicate in the cytoplasm of infected cells (for review of the alphaviral life cycle see José et al., Future Microbiol., 2009, vol. 4, pp. 837–856, which is incorporated herein by reference in its entirety). The total genome length of many alphaviruses typically ranges between 11,000 and 12,000 nucleotides, and the genomic RNA typically has a 5’-cap, and a 3’ poly(A) tail. The genome of alphaviruses encodes non-structural proteins (involved in transcription, modification and replication of viral RNA and in protein modification) and structural proteins (forming the virus particle). There are typically two open reading frames (ORFs) in the genome. The four non-structural proteins (nsP1–nsP4) are typically encoded together by a first ORF beginning near the 5′ terminus of the genome, while alphavirus structural proteins are encoded together by a second ORF which is found downstream of the first ORF and extends near the 3’ terminus of the genome. Typically, the first ORF is larger than the second ORF, the ratio being roughly 2:1. In cells infected by an alphavirus, only the nucleic acid sequence encoding non-structural proteins is translated from the genomic RNA, while the genetic information encoding structural proteins is translatable from a subgenomic transcript, which is an RNA molecule that resembles eukaryotic messenger RNA (mRNA; Gould et al., 2010, Antiviral Res., vol. 87 pp. 111–124, which is incorporated herein by reference in its entirety). Following infection, i.e. at early stages of the viral life cycle, the (+) stranded genomic RNA directly acts like a messenger RNA for the translation of the open reading frame encoding the non-structural poly- protein (nsP1234). [0623] Alphavirus-derived vectors have been proposed for delivery of foreign genetic information into target cells or target organisms. In simple approaches, a first ORF encodes an alphavirus-derived RNA-dependent RNA polymerase (replicase), which upon translation mediates self-amplification of the RNA. A second ORF encoding alphaviral structural proteins is replaced by an open reading frame encoding an HSV-2 construct described herein. Alphavirus-based trans-replication systems rely on alphavirus nucleotide sequence elements on two separate nucleic acid molecules: one nucleic acid molecule encodes a viral replicase, and the other nucleic acid molecule is capable of being replicated by said replicase in trans (hence the designation trans-replication system). Trans-replication requires the presence of both these nucleic acid molecules in a given host cell. The nucleic acid molecule capable of being replicated by the replicase in trans must comprise certain alphaviral sequence elements to allow recognition and RNA synthesis by the alphaviral replicase. [0624] Features of a non-modified uridine platform may include, for example, one or more of intrinsic adjuvant effect, as well as good tolerability and safety. Features of modified uridine (e.g., pseudouridine) platform may include reduced adjuvant effect, blunted immune innate immune sensor activating capacity and thus good tolerability and safety. Features of self-amplifying platform may include, for example, long duration of protein expression, good tolerability and safety, higher likelihood for efficacy with very low composition (e.g., immunogenic composition, e.g., vaccine) dose. [0625] The present disclosure provides particular RNA constructs optimized, for example, for improved manufacturability, encapsulation, expression level (and/or timing), etc. Certain components are discussed below, and certain preferred embodiments are exemplified herein. C. Codon Optimization and GC Enrichment [0626] As used herein, the term “codon-optimized” refers to alteration of codons in a coding region of a nucleic acid molecule (e.g., a polyribonucleotide) to reflect the typical codon usage of a host organism (e.g., a subject receiving a nucleic acid molecule (e.g., a polyribonucleotide)) without preferably altering the amino acid sequence encoded by the nucleic acid molecule. Within the context of the present disclosure, in some embodiments, coding regions are codon-optimized for optimal expression in a subject to be treated using the RNA molecules described herein. In some embodiments, codon-optimization may be performed such that codons for which frequently occurring tRNAs are available are inserted in place of “rare codons.” In some embodiments, codon- optimization may include increasing guanosine/cytosine (G/C) content of a coding region of RNA described herein as compared to the G/C content of the corresponding coding sequence of a wild type RNA, wherein the amino acid sequence encoded by the RNA is preferably not modified compared to the amino acid sequence. [0627] In some embodiments, a coding sequence (also referred to as a “coding region”) is codon optimized for expression in the subject to whom a composition (e.g., a pharmaceutical composition) is to be administered (e.g., a human). Thus, in some embodiments, sequences in such a polynucleotide (e.g., a polyribonucleotide) may differ from wild type sequences encoding the relevant antigen or portion or epitope thereof, even when the amino acid sequence of the antigen or portion or epitope thereof is wild type. [0628] In some embodiments, strategies for codon optimization for expression in a relevant subject (e.g., a human), and even, in some cases, for expression in a particular cell or tissue. [0629] Various species exhibit particular bias for certain codons of a particular amino acid. Without wishing to be bound by any one theory, codon bias (differences in codon usage between organisms) often correlates with the efficiency of translation of messenger RNA (mRNA), which is in turn believed to be dependent on, among other things, the properties of the codons being translated and the availability of particular transfer RNA (tRNA) molecules. The predominance of selected tRNAs in a cell may generally be a reflection of the codons used most frequently in peptide synthesis. Accordingly, genes may be tailored for optimal gene expression in a given organism based on codon optimization. Codon usage tables are available, for example, at the "Codon Usage Database" available at www.kazusa.orjp/codon/ and these tables may be adapted in a number of ways. Computer algorithms for codon optimizing a particular sequence for expression in a particular subject or its cells are also available, such as Gene Forge (Aptagen; Jacobus, PA), are also available. [0630] In some embodiments, a polynucleotide (e.g., a polyribonucleotide) of the present disclosure is codon optimized, wherein the codons in the polynucleotide (e.g., the polyribonucleotide) are adapted to human codon usage (herein referred to as “human codon optimized polynucleotide”). In some embodiments, a portion of a polyribonucleotide is codon optimized (e.g., a portion of or the portion encoding a glycoprotein or a portion of or the portion encoding a secretory signal). In some embodiments, the entire polyribonucleotide is codon optimized. Codons encoding the same amino acid occur at different frequencies in a subject, e.g., a human. Accordingly, in some embodiments, the coding sequence of a polynucleotide of the present disclosure is modified such that the frequency of the codons encoding the same amino acid corresponds to the naturally occurring frequency of that codon according to the human codon usage, e.g., as shown in Table 20. For example, in the case of the amino acid Ala, the wild type coding sequence is preferably adapted in a way that the codon “GCC” is used with a frequency of 0.40, the codon “GCT” is used with a frequency of 0.28, the codon “GCA” is used with a frequency of 0.22 and the codon “GCG” is used with 30 a frequency of 0.10 etc. (see Table 20). Accordingly, in some embodiments, such a procedure Ĩas exemplified for Ala) is applied for each amino acid encoded by the coding sequence of a polynucleotide to obtain sequences adapted to human codon usage. Table 20: Human codon usage with frequencies indicated for each amino acid. Amino acid codon frequency Amino acid codon frequency Ala GCG 0.10 Pro CCG 0.11
Figure imgf000351_0001
Lou CTC 0.20 Tyr TAC* 0.58 Met ATG* 1 Stop TGA* 061 [063 e
Figure imgf000352_0001
described in WO2002/098443, which is incorporated by reference herein in its entirety. In some embodiments, a coding sequence may be optimized using a multiparametric optimization strategy. In some embodiments, optimization parameters may include parameters that influence protein expression, which can be, for example, impacted on a transcription level, an RNA level, and/or a translational level. In some embodiments, exemplary optimization parameters include, but are not limited to transcription-level parameters (including, e.g., GC content, consensus splice sites, cryptic splice sites, SD sequences, TATA boxes, termination signals, artificial recombination sites, and combinations thereof); RNA-level parameters (including, e.g., RNA instability motifs, ribosomal entry sites, repetitive sequences, and combinations thereof); translation-level parameters (including, e.g., codon usage, premature poly(A) sites, ribosomal entry sites, secondary structures, and combinations thereof); or combinations thereof. In some embodiments, a coding sequence may be optimized by a GeneOptimizer algorithm as described in Fath et al. “Multiparameter RNA and Codon Optimization: A Standardized Tool to Assess and Enhance Autologous Mammalian Gene Expression” PLoS ONE 6(3): e17596; Rabb et al., which is incorporated herein by reference in its entirety, “The GeneOptimizer Algorithm: using a sliding window approach to cope with the vast sequence space in multiparameter DNA sequence optimization” Systems and Synthetic Biology (2010) 4:215-225; and Graft et al. “Codon-optimized genes that enable increased heterologous expression in mammalian cells and elicit efficient immune responses in mice after vaccination of naked DNA” Methods Mol Med (2004) 94:197-210, the entire content of each of which is incorporated herein for the purposes described herein. In some embodiments, a coding sequence may be optimized by Eurofins’ adaption and optimization algorithm “GENEius” as described in Eurofins’ Application Notes: Eurofins’ adaption and optimization software “GENEius” in comparison to other optimization algorithms, the entire content of which is incorporated by reference for the purposes described herein. [0632] In some embodiments, a coding sequence utilized in accordance with the present disclosure has G/C content that is increased compared to a coding sequence for an HSV gC, gD, and/or gE (or portion thereof) construct described herein. In some embodiments, guanosine/cytidine (G/C) content of a coding region is modified relative to a comparable coding sequence for an HSV gC, gD, and/or gE (or portion thereof) construct described herein, but the amino acid sequence encoded by the polyribonucleotide not modified. [0633] Without wishing to be bound by any particular theory, it is proposed that GC enrichment may improve translation of a payload sequence. Typically, sequences having an increased G (guanosine)/C (cytidine) content are more stable than sequences having an increased A (adenosine)/U (uridine) content. In respect to the fact that several codons code for one and the same amino acid (so-called degeneration of the genetic code), the most favorable codons for the stability can be determined (so-called alternative codon usage). Depending on the amino acid to be encoded by a polyribonucleotide, there are various possibilities for modification of the ribonucleic acid sequence, compared to its wild type sequence. In particular, codons which contain A and/or U nucleosides can be modified by substituting these codons by other codons, which code for the same amino acids but contain no A and/or U or contain a lower content of A and/or U nucleosides. [0634] In some embodiments, G/C content of a coding region of a polyribonucleotide described herein is increased by at least 1%, at least 2%, at least 3%, at least 4%, at least 5%, at least 6%, or even more compared to the G/C content of the coding region prior to codon optimization, e.g., of the wild type RNA. In some embodiments, G/C content of a coding region of a polyribonucleotide described herein is decreased by at least 1%, at least 2%, at least 3%, at least 4%, at least 5%, at least 6%, or even more compared to the G/C content of the coding region prior to codon optimization, e.g., of the wild type RNA. [0635] In some embodiments, G/C content of a coding region of a polyribonucleotide provided herein is decreased by at least 1%, at least 2%, at least 3%, at least 4%, at least 5%, at least 6%, or even more compared to the G/C content of the coding region prior to codon optimization, e.g., of the wild type RNA. In some embodiments, G/C content of a coding region of a polyribonucleotide provided herein comprises at the most 65%, such as at the most 64%, such as at the most 63%, such as at the most 62%, such as at the most 61%, such as at the most 60% G/C content compared to the G/C content of the coding region prior to codon optimization, e.g., of the wild type RNA. [0636] In some embodiments, C content of a coding region of a polyribonucleotide provided herein is decreased by at least 1%, at least 2%, at least 3%, at least 4%, at least 5%, at least 6%, or even more compared to the C content of the coding region prior to codon optimization, e.g., of the wild type RNA. In some embodiments, C content of a coding region of a polyribonucleotide provided herein comprises at the most 40%, such as at the most 39%, such as at the most 38%, such as at the most 37%, such as at the most 36%, such as at the most 35% C content compared to the C content of the coding region prior to codon optimization, e.g., of the wild type RNA. [0637] In some embodiments, stability and translation efficiency of a polyribonucleotide may incorporate one or more elements established to contribute to stability and/or translation efficiency of the polyribonucleotide; exemplary such elements are described, for example, in PCT/EP2006/009448 incorporated herein by reference. In some embodiments, to increase expression of a polyribonucleotide used according to the present disclosure, a polyribonucleotide may be modified within the coding region, i.e., the sequence encoding the expressed peptide or protein, without altering the sequence of the expressed peptide or protein, for example so as to increase the GC- content to increase RNA stability and/or to perform a codon optimization and, thus, enhance translation in cells. D. Certain Example RNA Constructs [0638] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 65. In some embodiments, an RNA construct comprises a 5’ cap, a 5’UTR, a polyribonucleotide that encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 65, a 3’ UTR, and a polyA tail. [0639] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 70. In some embodiments, an RNA construct comprises a 5’ cap, a 5’UTR, a polyribonucleotide that encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 70, a 3’ UTR, and a polyA tail. [0640] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 73. In some embodiments, an RNA construct comprises a 5’ cap, a 5’UTR, a polyribonucleotide that encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 73, a 3’ UTR, and a polyA tail. [0641] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 67. In some embodiments, an RNA construct comprises a 5’ cap, a 5’UTR, a polyribonucleotide that encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 67, a 3’ UTR, and a polyA tail. [0642] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 68. In some embodiments, an RNA construct comprises a 5’ cap, a 5’UTR, a polyribonucleotide that encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 68, a 3’ UTR, and a polyA tail. [0643] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 75. In some embodiments, an RNA construct comprises a 5’ cap, a 5’UTR, a polyribonucleotide that encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 75, a 3’ UTR, and a polyA tail. [0644] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 131. In some embodiments, an RNA construct comprises a 5’ cap, a 5’UTR, a polyribonucleotide that encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 131, a 3’ UTR, and a polyA tail. [0645] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 132. In some embodiments, an RNA construct comprises a 5’ cap, a 5’UTR, a polyribonucleotide that encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 132, a 3’ UTR, and a polyA tail. [0646] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 159. In some embodiments, an RNA construct comprises a 5’ cap, a 5’UTR, a polyribonucleotide that encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 159, a 3’ UTR, and a polyA tail. [0647] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 160. In some embodiments, an RNA construct comprises a 5’ cap, a 5’UTR, a polyribonucleotide that encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 160, a 3’ UTR, and a polyA tail. [0648] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 161. In some embodiments, an RNA construct comprises a 5’ cap, a 5’UTR, a polyribonucleotide that encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 161, a 3’ UTR, and a polyA tail. [0649] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 162. In some embodiments, an RNA construct comprises a 5’ cap, a 5’UTR, a polyribonucleotide that encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 162, a 3’ UTR, and a polyA tail. [0650] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 163. In some embodiments, an RNA construct comprises a 5’ cap, a 5’UTR, a polyribonucleotide that encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 163, a 3’ UTR, and a polyA tail. [0651] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 164. In some embodiments, an RNA construct comprises a 5’ cap, a 5’UTR, a polyribonucleotide that encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 164, a 3’ UTR, and a polyA tail. [0652] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 327. In some embodiments, an RNA construct comprises a 5’ cap, a 5’UTR, a polyribonucleotide that encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 327, a 3’ UTR, and a polyA tail. [0653] In some embodiments, a polyribonucleotide provided herein encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 328. In some embodiments, an RNA construct comprises a 5’ cap, a 5’UTR, a polyribonucleotide that encodes a polypeptide, wherein the polypeptide comprises an amino acid sequence according to SEQ ID NO: 328, a 3’ UTR, and a polyA tail. IV. RNA Delivery Technologies [0654] Provided polyribonucleotides may be delivered for therapeutic applications described herein using any appropriate methods known in the art, including, e.g., delivery as naked RNAs, or delivery mediated by viral and/or non-viral vectors, polymer-based vectors, lipid compositions, nanoparticles (e.g., lipid nanoparticles, polymeric nanoparticles, lipid-polymer hybrid nanoparticles, etc.), and/or peptide-based vectors. See, e.g., Wadhwa et al. “Opportunities and Challenges in the Delivery of mRNA-Based Vaccines” Pharmaceutics (2020) 102 (27 pages), the content of which is incorporated herein by reference, for information on various approaches that may be useful for delivery polyribonucleotides described herein. [0655] In some embodiments, one or more polyribonucleotides can be formulated with lipid nanoparticles for delivery (e.g., administration). [0656] In some embodiments, lipid nanoparticles can be designed to protect polyribonucleotides from extracellular RNases and/or engineered for systemic delivery of the RNA to target cells. In some embodiments, such lipid nanoparticles may be particularly useful to deliver polyribonucleotides when polyribonucleotides are intravenously or intramuscularly administered to a subject. A. Lipid Compositions 1. Lipids and Lipid-Like Materials [0657] The terms "lipid" and "lipid-like material" are broadly defined herein as molecules which comprise one or more hydrophobic moieties or groups and optionally also one or more hydrophilic moieties or groups. Molecules comprising hydrophobic moieties and hydrophilic moieties are also frequently denoted as amphiphiles. Lipids are usually poorly soluble in water. In an aqueous environment, the amphiphilic nature allows the molecules to self-assemble into organized structures and different phases. One of those phases consists of lipid bilayers, as they are present in vesicles, multilamellar/unilamellar liposomes, or membranes in an aqueous environment. Hydrophobicity can be conferred by the inclusion of a polar groups that include, but are not limited to, long-chain saturated and unsaturated aliphatic hydrocarbon groups and such groups substituted by one or more aromatic, cycloaliphatic, or heterocyclic group(s). The hydrophilic groups may comprise polar and/or charged groups and include carbohydrates, phosphate, carboxylic, sulfate, amino, sulfhydryl, nitro, hydroxyl, and other like groups. [0658] Often, an amphiphilic compound has a polar head attached to a long hydrophobic tail. In some embodiments, the polar portion is soluble in water, while the non-polar portion is insoluble in water. In addition, the polar portion may have either a formal positive charge, or a formal negative charge. Alternatively, the polar portion may have both a formal positive and a negative charge, and be a zwitterion or inner salt. For purposes of the disclosure, the amphiphilic compound can be, but is not limited to, one or a plurality of natural or non-natural lipids and lipid-like compounds. [0659] A "lipid-like material" is a substance that is structurally and/or functionally related to a lipid but may not be considered a lipid in a strict sense. For example, the term includes compounds that are able to form amphiphilic layers as they are present in vesicles, multilamellar/unilamellar liposomes, or membranes in an aqueous environment and includes surfactants, or synthesized compounds with both hydrophilic and hydrophobic moieties. Generally speaking, the term refers to molecules, which comprise hydrophilic and hydrophobic moieties with different structural organization, which may or may not be similar to that of lipids. [0660] Specific examples of amphiphilic compounds that may be included in an amphiphilic layer include, but are not limited to, phospholipids, aminolipids and sphingolipids. [0661] Generally, lipids may be divided into eight categories: fatty acids, glycerolipids, glycerophospholipids, sphingolipids, saccharolipids, polyketides (derived from condensation of ketoacyl subunits), sterols and prenol lipids (derived from condensation of isoprene subunits). Although the term "lipid" is sometimes used as a synonym for fats, fats are a subgroup of lipids called triglycerides. Lipids also encompass molecules such as fatty acids and their derivatives (including tri-, di-, monoglycerides, and phospholipids), as well as sterol- containing metabolites such as cholesterol. [0662] Fatty acids are a diverse group of molecules made of a hydrocarbon chain that terminates with a carboxylic acid group; this arrangement confers the molecule with a polar, hydrophilic end, and a nonpolar, hydrophobic end that is insoluble in water. The carbon chain, typically between four and 24 carbons long, may be saturated or unsaturated, and may be attached to functional groups containing oxygen, halogens, nitrogen, and sulfur. If a fatty acid contains a double bond, there is the possibility of either a cis or trans geometric isomerism, which significantly affects the molecule's configuration. Cis-double bonds cause the fatty acid chain to bend, an effect that is compounded with more double bonds in the chain. Other major lipid classes in the fatty acid category are the fatty esters and fatty amides. [0663] Glycerolipids are composed of mono-, di-, and tri-substituted glycerols, the best-known being the fatty acid triesters of glycerol, called triglycerides. The word "triacylglycerol" is sometimes used synonymously with "triglyceride". In these compounds, the three hydroxyl groups of glycerol are each esterified, typically by different fatty acids. Additional subclasses of glycerolipids are represented by glycosylglycerols, which are characterized by the presence of one or more sugar residues attached to glycerol via a glycosidic linkage. [0664] Glycerophospholipids are amphipathic molecules (containing both hydrophobic and hydrophilic regions) that contain a glycerol core linked to two fatty acid-derived "tails" by ester linkages and to one "head" group by a phosphate ester linkage. Examples of glycerophospholipids, usually referred to as phospholipids (though sphingomyelins are also classified as phospholipids) are phosphatidylcholine (also known as PC, GPCho or lecithin), phosphatidylethanolamine (PE or GPEtn) and phosphatidylserine (PS or GPSer). [0665] Sphingolipids are members of a complex family of compounds that share a common structural feature, a sphingoid base backbone. The major sphingoid base in mammals is commonly referred to as sphingosine. Ceramides (N-acyl-sphingoid bases) are a major subclass of sphingoid base derivatives with an amide-linked fatty acid. The fatty acids are typically saturated or mono-unsaturated with chain lengths from 16 to 26 carbon atoms. The major phosphosphingolipids of mammals are sphingomyelins (ceramide phosphocholines), whereas insects contain mainly ceramide phosphoethanolamines and fungi have phytoceramide phosphoinositols and mannose-containing headgroups. The glycosphingolipids are a diverse family of molecules composed of one or more sugar residues linked via a glycosidic bond to the sphingoid base. Examples of these are the simple and complex glycosphingolipids such as cerebrosides and gangliosides. [0666] Sterols, such as cholesterol and its derivatives, or tocopherol and its derivatives, are important components of membrane lipids, along with the glycerophospholipids and sphingomyelins. [0667] Saccharolipids are compounds in which fatty acids are linked directly to a sugar backbone, forming structures that are compatible with membrane bilayers. In the saccharolipids, a monosaccharide substitutes for the glycerol backbone present in glycerolipids and glycerophospholipids. The most familiar saccharolipids are the acylated glucosamine precursors of the Lipid A component of the lipopolysaccharides in Gram-negative bacteria. Typical lipid A molecules are disaccharides of glucosamine, which are derivatized with as many as seven fatty-acyl chains. The minimal lipopolysaccharide required for growth in E. coli is Kdo2-Lipid A, a hexa-acylated disaccharide of glucosamine that is glycosylated with two 3-deoxy-D-manno-octulosonic acid (Kdo) residues. [0668] Polyketides are synthesized by polymerization of acetyl and propionyl subunits by classic enzymes as well as iterative and multimodular enzymes that share mechanistic features with the fatty acid synthases. They comprise a large number of secondary metabolites and natural products from animal, plant, bacterial, fungal and marine sources, and have great structural diversity. Many polyketides are cyclic molecules whose backbones are often further modified by glycosylation, methylation, hydroxylation, oxidation, or other processes. [0669] Lipids and lipid-like materials may be cationic, anionic or neutral. Neutral lipids or lipid-like materials exist in an uncharged or neutral zwitterionic form at a selected pH. [0670] In some embodiments, suitable lipids or lipid-like materials for use in the present disclosure include those described in WO2020/128031 and US20200163878, the entire contents of each of which are incorporated herein by reference for the purposes described herein. 2. Cationic or cationically ionizable lipids or lipid-like materials [0671] In some embodiments cationic or cationically ionizable lipids or lipid-like materials contemplated for use herein include any cationic or cationically ionizable lipids or lipid-like materials which are able to electrostatically bind nucleic acid. In one embodiment, cationic or cationically ionizable lipids or lipid-like materials contemplated for use herein can be associated with nucleic acid, e.g., by forming complexes with the nucleic acid or forming vesicles in which the nucleic acid is enclosed or encapsulated. [0672] Cationic lipids or lipid-like materials are characterized in that they have a net positive charge (e.g., at a relevant pH). Cationic lipids or lipid-like materials bind negatively charged nucleic acid by electrostatic interaction. Generally, cationic lipids possess a lipophilic moiety, such as a sterol, an acyl chain, a diacyl or more acyl chains, and the head group of the lipid typically carries the positive charge. [0673] In certain embodiments, a cationic lipid or lipid-like material has a net positive charge only at certain pH, in particular acidic pH, while it has preferably no net positive charge, preferably has no charge, i.e., it is neutral, at a different, preferably higher pH such as physiological pH. This ionizable behavior is thought to enhance efficacy through helping with endosomal escape and reducing toxicity as compared with particles that remain cationic at physiological pH. [0674] In some embodiments, a cationic or cationically ionizable lipid or lipid-like material comprises a head group which includes at least one nitrogen atom (N) which is positive charged or capable of being protonated. [0675] Examples of cationic lipids include, but are not limited to 1,2-dioleoyl-3-trimethylammonium propane (DOTAP); N,N-dimethyl-2,3-dioleyloxypropylamine (DODMA), 1,2-di-O-octadecenyl-3-trimethylammonium propane (DOTMA), 3-(N—(N′,N′-dimethylaminoethane)-carbamoyl)cholesterol (DC-Chol), dimethyldioctadecylammonium (DDAB); 1,2-dioleoyl-3-dimethylammonium-propane (DODAP); 1,2-diacyloxy-3- dimethylammonium propanes; 1,2-dialkyloxy-3-dimethylammonium propanes; dioctadecyldimethyl ammonium chloride (DODAC), 1,2-distearyloxy-N,N-dimethyl-3-aminopropane (DSDMA), 2,3-di(tetradecoxy)propyl-(2- hydroxyethyl)-dimethylazanium (DMRIE), 1,2-dimyristoyl-sn-glycero-3-ethylphosphocholine (DMEPC), l,2-dimyristoyl- 3-trimethylammonium propane (DMTAP), 1,2-dioleyloxypropyl-3-dimethyl-hydroxyethyl ammonium bromide (DORIE), and 2,3-dioleoyloxy- N-[2(spermine carboxamide)ethyl]-N,N-dimethyl-l-propanamium trifluoroacetate (DOSPA), 1,2- dilinoleyloxy-N,N-dimethylaminopropane (DLinDMA), 1,2-dilinolenyloxy-N,N-dimethylaminopropane (DLenDMA), dioctadecylamidoglycyl spermine (DOGS), 3-dimethylamino-2-(cholest-5-en-3-beta-oxybutan-4-oxy)-1-(cis,cis-9,12- oc-tadecadienoxy)propane (CLinDMA), 2-[5′-(cholest-5-en-3-beta-oxy)-3′-oxapentoxy)-3-dimethyl-1-(cis,cis-9′,12′- octadecadienoxy)propane (CpLinDMA), N,N-dimethyl-3,4-dioleyloxybenzylamine (DMOBA), 1,2-N,N′-dioleylcarbamyl- 3-dimethylaminopropane (DOcarbDAP), 2,3-Dilinoleoyloxy-N,N-dimethylpropylamine (DLinDAP), 1,2-N,N′- Dilinoleylcarbamyl-3-dimethylaminopropane (DLincarbDAP), 1,2-Dilinoleoylcarbamyl-3-dimethylaminopropane (DLinCDAP), 2,2-dilinoleyl-4-dimethylaminomethyl-[1,3]-dioxolane (DLin-K-DMA), 2,2-dilinoleyl-4-dimethylaminoethyl- [1,3]-dioxolane (DLin-K-XTC2-DMA), 2,2-dilinoleyl-4-(2-dimethylaminoethyl)-[1,3]-dioxolane (DLin-KC2-DMA), heptatriaconta-6,9,28,31-tetraen-19-yl-4-(dimethylamino)butanoate (DLin-MC3-DMA), N-(2-Hydroxyethyl)-N,N- dimethyl-2,3-bis(tetradecyloxy)-1-propanaminium bromide (DMRIE), (±)-N-(3-aminopropyl)-N,N-dimethyl-2,3-bis(cis- 9-tetradecenyloxy)-1-propanaminium bromide (GAP-DMORIE), (±)-N-(3-aminopropyl)-N,N-dimethyl-2,3- bis(dodecyloxy)-1-propanaminium bromide (GAP-DLRIE), (±)-N-(3-aminopropyl)-N,N-dimethyl-2,3- bis(tetradecyloxy)-1-propanaminium bromide (GAP-DMRIE), N-(2-Aminoethyl)-N,N-dimethyl-2,3-bis(tetradecyloxy)-1- propanaminium bromide (βAE-DMRIE), N-(4-carboxybenzyl)-N,N-dimethyl-2,3-bis(oleoyloxy)propan-1-aminium (DOBAQ), 2-({8-[(3β)-cholest-5-en-3-yloxy]octyl}oxy)-N,N-dimethyl-3-[(9Z,12Z)-octadeca-9,12-dien-1-yloxy]propan- 1-amine (Octyl-CLinDMA), 1,2-dimyristoyl-3-dimethylammonium-propane (DMDAP), 1,2-dipalmitoyl-3- dimethylammonium-propane (DPDAP), N1-[2-((1S)-1-[(3-aminopropyl)amino]-4-[di(3-amino- propyl)amino]butylcarboxamido)ethyl]-3,4-di[oleyloxy]-benzamide (MVL5), 1,2-dioleoyl-sn-glycero-3- ethylphosphocholine (DOEPC), 2,3-bis(dodecyloxy)-N-(2-hydroxyethyl)-N,N-dimethylpropan-1-amonium bromide (DLRIE), N-(2-aminoethyl)-N,N-dimethyl-2,3-bis(tetradecyloxy)propan-1-aminium bromide (DMORIE), di((Z)-non-2- en-1-yl) 8,8'-((((2(dimethylamino)ethyl)thio)carbonyl)azanediyl)dioctanoate (ATX), N,N-dimethyl-2,3- bis(dodecyloxy)propan-1-amine (DLDMA), N,N-dimethyl-2,3-bis(tetradecyloxy)propan-1-amine (DMDMA), Di((Z)-non- 2-en-1-yl)-9-((4-(dimethylaminobutanoyl)oxy)heptadecanedioate (L319), N-Dodecyl-3-((2-dodecylcarbamoyl-ethyl)- {2-[(2-dodecylcarbamoyl-ethyl)-2-{(2-dodecylcarbamoyl-ethyl)-[2-(2-dodecylcarbamoyl-ethylamino)-ethyl]-amino}- ethylamino)propionamide (lipidoid 98N12-5), 1-[2-[bis(2-hydroxydodecyl)amino]ethyl-[2-[4-[2-[bis(2 hydroxydodecyl)amino]ethyl]piperazin-1-yl]ethyl]amino]dodecan-2-ol (lipidoid C12-200), LIPOFECTIN® (commercially available cationic liposomes comprising DOTMA and 1 ,2-dioleoyl-sn-3phosphoethanolamine (DOPE), from GIBCO/BRL, Grand Island, N.Y.); LIPOFECTAMINE® (commercially available cationic liposomes comprising N-(1 -(2,3dioleyloxy)propyl)-N-(2-(sperminecarboxamido)ethyl)-N,N-dimethylammonium trifluoroacetate (DOSPA) and (DOPE), from GIBCO/BRL); and TRANSFECTAM® (commercially available cationic lipids comprising dioctadecylamidoglycyl carboxyspermine (DOGS) in ethanol from Promega Corp., Madison, Wis.) or any combination of any of the foregoing. Further suitable cationic lipids for use in the present disclosure include those described in WO2020/128031 and US20200163878, the entire contents of each of which are incorporated herein by reference for the purposes described herein. Further suitable cationic lipids for use in the present disclosure include those described in WO2010/053572 (including Cl 2-200 described at paragraph [00225]) and WO2012/170930, both of which are incorporated herein by reference for the purposes described herein. Additional suitable cationic lipids for use in the present disclosure include HGT4003, HGT5000, HGTS001, HGT5001, HGT5002 (see US20150140070A1, which is incorporated herein by reference in its entirety). [0676] In some embodiments, formulations that are useful for pharmaceutical compositions (e.g., immunogenic compositions, e.g., vaccines) compositions as described herein can comprise at least one cationic lipid. Representative cationic lipids include, but are not limited to, 1 ,2-dilinoleyoxy-3-(dimethylamino)acetoxypropane (DLin-DAC), 1 ,2-dilinoleyoxy-3morpholinopropane (DLin-MA), 1,2-dilinoleoyl-3-dimethylaminopropane (DLinDAP), 1 ,2-dilinoleylthio-3-dimethylaminopropane (DLin-S-DMA), 1 -linoleoyl-2-linoleyloxy-3dimethylaminopropane (DLin-2- DMAP), 1 ,2-dilinoleyloxy-3-trimethylaminopropane chloride salt (DLin-TMA.CI), 1 ,2-dilinoleoyl-3- trimethylaminopropane chloride salt (DLin-TAP.CI), 1 ,2-dilinoleyloxy-3-(N-methylpiperazino)propane (DLin-MPZ), 3- (N,Ndilinoleylamino)-1 ,2-propanediol (DLinAP), 3-(N,N-dioleylamino)-1 ,2-propanediol (DOAP), 1 ,2-dilinoleyloxo-3- (2-N,N-dimethylamino)ethoxypropane (DLin-EG-DMA), and 2,2-dilinoleyl-4-dimethylaminomethyl-[1 ,3]-dioxolane (DLin-K-DMA), 2,2-dilinoleyl-4-(2-dimethylaminoethyl)-[1 ,3]-dioxolane (DLin-KC2-DMA); dilinoleyl-methyl-4- dimethylaminobutyrate (DLin-MC3-DMA); MC3 (US20100324120, which is incorporated herein by reference in its entirety). [0677] In some embodiments, amino or cationic lipids useful in accordance with the present disclosure have at least one protonatable or deprotonatable group, such that the lipid is positively charged at a pH at or below physiological pH (e.g. pH 7.4), and neutral at a second pH, preferably at or above physiological pH. It will, of course, be understood that the addition or removal of protons as a function of pH is an equilibrium process, and that the reference to a charged or a neutral lipid refers to the nature of the predominant species and does not require that all of lipids have to be present in the charged or neutral form. Lipids having more than one protonatable or deprotonatable group, or which are zwitterionic, are not excluded and may likewise be suitable in the context of the present invention. [0678] In some embodiments, a protonatable lipid has a pKa of the protonatable group in the range of about 4 to about 11, e.g., a pKa of about 5 to about 7. [0679] In some embodiments, a cationic lipid may comprise from about 10 mol % to about 100 mol %, about 20 mol % to about 100 mol %, about 30 mol % to about 100 mol %, about 40 mol % to about 100 mol %, or about 50 mol % to about 100 mol % of total lipid present in a lipid composition utilized in accordance with the present disclosure. 3. Additional lipids or lipid-like materials [0680] In some embodiments, formulations utilized in accordance with the present disclosure may comprise lipids or lipid-like materials other than cationic or cationically ionizable lipids or lipid-like materials, i.e., non- cationic lipids or lipid-like materials (including non-cationically ionizable lipids or lipid-like materials). Collectively, anionic and neutral lipids or lipid-like materials are referred to herein as non-cationic lipids or lipid-like materials. In some embodiments, optimizing a formulation of nucleic acid particles by addition of other hydrophobic moieties, such as cholesterol and lipids, in addition to an ionizable/cationic lipid or lipid-like material may, for example, enhance particle stability and efficacy of nucleic acid delivery. [0681] In some embodiments, a lipid or lipid-like material may be incorporated which may or may not affect the overall charge of particles. In certain embodiments, such lipid or lipid-like material is a non-cationic lipid or lipid-like material. [0682] In some embodiments, a non-cationic lipid may comprise, e.g., one or more anionic lipids and/or neutral lipids. An "anionic lipid" is negatively charged (e.g., at a selected pH). [0683] A "neutral lipid" exists either in an uncharged or neutral zwitterionic form (e.g., at a selected pH). In some embodiments, a formulation comprises one of the following neutral lipid components: (1) a phospholipid, (2) cholesterol or a derivative thereof; or (3) a mixture of a phospholipid and cholesterol or a derivative thereof. Examples of cholesterol derivatives include, but are not limited to, cholestanol, cholestanone, cholestenone, coprostanol, cholesteryl-2'-hydroxyethyl ether, cholesteryl-4'- hydroxybutyl ether, tocopherol and derivatives thereof, and mixtures thereof. [0684] Specific example phospholipids that can be used include, but are not limited to, phosphatidylcholines, phosphatidylethanolamines, phosphatidylglycerols, phosphatidic acids, phosphatidylserines or sphingomyelin. Such phospholipids include in particular diacylphosphatidylcholines, such as distearoylphosphatidylcholine (DSPC), dioleoylphosphatidylcholine (DOPC), dimyristoylphosphatidylcholine (DMPC), dipentadecanoylphosphatidylcholine, dilauroylphosphatidylcholine, dipalmitoylphosphatidylcholine (DPPC), diarachidoylphosphatidylcholine (DAPC), dibehenoylphosphatidylcholine (DBPC), ditricosanoylphosphatidylcholine (DTPC), dilignoceroylphatidylcholine (DLPC), palmitoyloleoyl-phosphatidylcholine (POPC), 1,2-di-O-octadecenyl-sn- glycero-3-phosphocholine (18:0 Diether PC), 1-oleoyl-2-cholesterylhemisuccinoyl-sn-glycero-3-phosphocholine (OChemsPC), 1-hexadecyl-sn-glycero-3-phosphocholine (C16 Lyso PC) and phosphatidylethanolamines, in particular diacylphosphatidylethanolamines, such as dioleoylphosphatidylethanolamine (DOPE), distearoyl- phosphatidylethanolamine (DSPE), dipalmitoyl-phosphatidylethanolamine (DPPE), dimyristoyl- phosphatidylethanolamine (DMPE), dilauroyl-phosphatidylethanolamine (DLPE), diphytanoyl- phosphatidylethanolamine (DPyPE), and further phosphatidylethanolamine lipids with different hydrophobic chains. [0685] In certain embodiments, a formulation utilized in accordance with the present disclosure includes DSPC or DSPC and cholesterol. [0686] In certain embodiments, formulations utilized in accordance with the present disclosure include both a cationic lipid and an additional (non-cationic) lipid. [0687] In some embodiments, formulations herein include a polymer conjugated lipid such as a pegylated lipid. "Pegylated lipids" comprise both a lipid portion and a polyethylene glycol portion. Pegylated lipids are known in the art. [0688] Without wishing to be bound by theory, the amount of (total) cationic lipid compared to the amount of other lipid(s) in formulation may affect important characteristics, such as charge, particle size, stability, tissue selectivity, and bioactivity of the nucleic acid. In some embodiments, the molar ratio of the at least one cationic lipid to the at least one additional lipid is from about 10:0 to about 1:9, about 4:1 to about 1:2, or about 3:1 to about 1:1. [0689] In some embodiments, a non-cationic lipid, in particular a neutral lipid, (e.g., one or more phospholipids and/or cholesterol) may comprise from about 0 mol % to about 90 mol %, from about 0 mol % to about 80 mol %, from about 0 mol % to about 70 mol %, from about 0 mol % to about 60 mol %, or from about 0 mol % to about 50 mol %, of the total lipid present in a formulation. 4. Lipoplex Particles [0690] In certain embodiments of the present disclosure, the RNA described herein may be present in RNA lipoplex particles. [0691] An "RNA lipoplex particle" contains lipid, in particular cationic lipid, and RNA. Electrostatic interactions between positively charged liposomes and negatively charged RNA results in complexation and spontaneous formation of RNA lipoplex particles. Positively charged liposomes may be generally synthesized using a cationic lipid, such as DOTMA, and additional lipids, such as DOPE. In one embodiment, a RNA lipoplex particle is a nanoparticle. [0692] In certain embodiments, RNA lipoplex particles include both a cationic lipid and an additional lipid. In some embodiments, a cationic lipid is DOTMA and the additional lipid is DOPE. [0693] In some embodiments, the molar ratio of the at least one cationic lipid to the at least one additional lipid is from about 10:0 to about 1:9, about 4:1 to about 1:2, or about 3:1 to about 1:1. In specific embodiments, the molar ratio may be about 3:1, about 2.75:1, about 2.5:1, about 2.25:1, about 2:1, about 1.75:1, about 1.5:1, about 1.25:1, or about 1:1. In some embodiments, the molar ratio of the at least one cationic lipid to the at least one additional lipid is about 2:1. [0694] In some embodiments, RNA lipoplex particles have an average diameter that in one embodiment ranges from about 200 nm to about 1000 nm, from about 200 nm to about 800 nm, from about 250 to about 700 nm, from about 400 to about 600 nm, from about 300 nm to about 500 nm, or from about 350 nm to about 400 nm. In specific embodiments, the RNA lipoplex particles have an average diameter of about 200 nm, about 225 nm, about 250 nm, about 275 nm, about 300 nm, about 325 nm, about 350 nm, about 375 nm, about 400 nm, about 425 nm, about 450 nm, about 475 nm, about 500 nm, about 525 nm, about 550 nm, about 575 nm, about 600 nm, about 625 nm, about 650 nm, about 700 nm, about 725 nm, about 750 nm, about 775 nm, about 800 nm, about 825 nm, about 850 nm, about 875 nm, about 900 nm, about 925 nm, about 950 nm, about 975 nm, or about 1000 nm. In an embodiment, the RNA lipoplex particles have an average diameter that ranges from about 250 nm to about 700 nm. In another embodiment, the RNA lipoplex particles have an average diameter that ranges from about 300 nm to about 500 nm. In some embodiments, RNA lipoplex particles have an average diameter of about 400 nm. [0695] RNA lipoplex particles and compositions comprising RNA lipoplex particles described herein are useful for delivery of RNA to a target tissue after parenteral administration, in particular after intravenous administration. The RNA lipoplex particles may be prepared using liposomes that may be obtained by injecting a solution of the lipids in ethanol into water or a suitable aqueous phase. In one embodiment, the aqueous phase has an acidic pH. In one embodiment, the aqueous phase comprises acetic acid, e.g., in an amount of about 5 mM. Liposomes may be used for preparing RNA lipoplex particles by mixing the liposomes with RNA. In one embodiment, the liposomes and RNA lipoplex particles comprise at least one cationic lipid and at least one additional lipid. In one embodiment, the at least one cationic lipid comprises 1,2-di-O-octadecenyl-3-trimethylammonium propane (DOTMA) and/or 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP). In one embodiment, the at least one additional lipid comprises 1,2-di-(9Z-octadecenoyl)-sn-glycero-3-phosphoethanolamine (DOPE), cholesterol (Chol) and/or 1,2- dioleoyl-sn-glycero-3-phosphocholine (DOPC). In one embodiment, the at least one cationic lipid comprises 1,2-di-O- octadecenyl-3-trimethylammonium propane (DOTMA) and the at least one additional lipid comprises 1,2-di-(9Z- octadecenoyl)-sn-glycero-3-phosphoethanolamine (DOPE). In one embodiment, the liposomes and RNA lipoplex particles comprise 1,2-di-O-octadecenyl-3-trimethylammonium propane (DOTMA) and 1,2-di-(9Z-octadecenoyl)-sn- glycero-3-phosphoethanolamine (DOPE). [0696] Spleen targeting RNA lipoplex particles are described in WO 2013/143683, herein incorporated by reference. It has been found that RNA lipoplex particles having a net negative charge may be used to preferentially target spleen tissue or spleen cells such as antigen-presenting cells, in particular dendritic cells. Accordingly, following administration of the RNA lipoplex particles, RNA accumulation and/or RNA expression in the spleen occurs. Thus, RNA lipoplex particles of the disclosure may be used for expressing RNA in the spleen. In an embodiment, after administration of the RNA lipoplex particles, no or essentially no RNA accumulation and/or RNA expression in the lung and/or liver occurs. In one embodiment, after administration of the RNA lipoplex particles, RNA accumulation and/or RNA expression in antigen presenting cells, such as professional antigen presenting cells in the spleen occurs. Thus, RNA lipoplex particles of the disclosure may be used for expressing RNA in such antigen presenting cells. In one embodiment, the antigen presenting cells are dendritic cells and/or macrophages. 5. Lipid Nanoparticles (LNPs) [0697] In some embodiments, nucleic acid such as RNA described herein is administered in the form of lipid nanoparticles (LNPs). In some embodiments, LNPs may comprise any lipid capable of forming a particle to which the one or more nucleic acid molecules are attached, or in which the one or more nucleic acid molecules are encapsulated. [0698] In some embodiments, an LNP comprises one or more cationic lipids, and one or more stabilizing lipids. Stabilizing lipids include neutral lipids and pegylated lipids. [0699] In some embodiments, an LNP comprises a cationic lipid, a neutral lipid, a sterol, a polymer conjugated lipid; and an RNA, encapsulated within or associated with the lipid nanoparticle. [0700] In some embodiments, a neutral lipid is selected from the group consisting of DSPC, DPPC, DMPC, DOPC, POPC, DOPE, DOPG, DPPG, POPE, DPPE, DMPE, DSPE, and SM. In some embodiments, the neutral lipid is selected from the group consisting of DSPC, DPPC, DMPC, DOPC, POPC, DOPE and SM. In some embodiments, the neutral lipid is DSPC. [0701] In some embodiments, a sterol is cholesterol. [0702] In some embodiments, a polymer conjugated lipid is a pegylated lipid. In some embodiments, a pegylated lipid has the following structure: or a pharmaceutically acceptable
Figure imgf000363_0001
salt, tautomer or stereoisomer thereof, wherein: R12 and R13 are each independently a straight or branched, saturated or unsaturated alkyl chain containing from 10 to 30 carbon atoms, wherein the alkyl chain is optionally interrupted by one or more ester bonds; and w has a mean value ranging from 30 to 60. In some embodiments, R12 and R13 are each independently straight, saturated alkyl chains containing from 12 to 16 carbon atoms. In some embodiments, w has a mean value ranging from 40 to 55. In some embodiments, the average w is about 45. In some embodiments, R12 and R13 are each independently a straight, saturated alkyl chain containing about 14 carbon atoms, and w has a mean value of about 45. [0703] In some embodiments, a pegylated lipid is DMG-PEG 2000, e.g., having the following structure: [0704] rmula (III):
Figure imgf000364_0001
(III) or a pharmaceutically acceptable salt, tautomer, prodrug or stereoisomer thereof, wherein: one of L1 or L2 is –O(C=O)-, -(C=O)O-, -C(=O)-, -O-, -S(O)x-, -S-S-, -C(=O)S-, SC(=O)-, -NRaC(=O)-, -C(=O)NRa-, NRaC(=O)NRa-, -OC(=O)NRa- or -NRaC(=O)O-, and the other of L1 or L2 is –O(C=O)-, -(C=O)O-, -C(=O)-, -O-, - S(O)x-, -S-S-, -C(=O)S-, SC(=O)-, -NRaC(=O)-, -C(=O)NRa-, NRaC(=O)NRa-, -OC(=O)NRa- or -NRaC(=O)O- or a direct bond; G1 and G2 are each independently unsubstituted C1-C12 alkylene or C1-C12 alkenylene; G3 is C1-C24 alkylene, C1-C24 alkenylene, C3-C8 cycloalkylene, C3-C8 cycloalkenylene; Ra is H or C1-C12 alkyl; R1 and R2 are each independently C6-C24 alkyl or C6-C24 alkenyl; R3 is H, OR5, CN, -C(=O)OR4, -OC(=O)R4 or –NR5C(=O)R4; R4 is C1-C12 alkyl; R5 is H or C1-C6 alkyl; and x is 0, 1 or 2. [0705] In some of the foregoing embodiments of Formula (III), the lipid has one of the following structures (IIIA) or (IIIB):
Figure imgf000364_0002
( ) ( ) wherein: A is a 3 to 8-membered cycloalkyl or cycloalkylene ring; R6 is, at each occurrence, independently H, OH or C1-C24 alkyl; and n is an integer ranging from 1 to 15. [0706] In some of the foregoing embodiments of Formula (III), the lipid has structure (IIIA), and in other embodiments, the lipid has structure (IIIB). [0707] In other embodiments of Formula (III), the lipid has one of the following structures (IIIC) or (IIID):
Figure imgf000365_0001
wherein y and z are each independently integers ranging from 1 to 12. [0708] In any of the foregoing embodiments of Formula (III), one of L1 or L2 is -O(C=O)-. For example, in some embodiments each of L1 and L2 are -O(C=O)-. In some different embodiments of any of the foregoing, L1 and L2 are each independently -(C=O)O- or -O(C=O)-. For example, in some embodiments each of L1 and L2 is -(C=O)O-. [0709] In some different embodiments of Formula (III), the lipid has one of the following structures (IIIE) or (IIIF): .
Figure imgf000365_0002
[0710] In some of the foregoing embodiments of Formula (III), the lipid has one of the following structures (IIIG), (IIIH), (IIII), or (IIIJ): ;
Figure imgf000365_0003
.
Figure imgf000366_0001
[0711] In some of the foregoing embodiments of Formula (III), n is an integer ranging from 2 to 12, for example from 2 to 8 or from 2 to 4. For example, in some embodiments, n is 3, 4, 5 or 6. In some embodiments, n is 3. In some embodiments, n is 4. In some embodiments, n is 5. In some embodiments, n is 6. [0712] In some other of the foregoing embodiments of Formula (III), y and z are each independently an integer ranging from 2 to 10. For example, in some embodiments, y and z are each independently an integer ranging from 4 to 9 or from 4 to 6. [0713] In some of the foregoing embodiments of Formula (III), R6 is H. In other of the foregoing embodiments, R6 is C1-C24 alkyl. In other embodiments, R6 is OH. [0714] In some embodiments of Formula (III), G3 is unsubstituted. In other embodiments, G3 is substituted. In various different embodiments, G3 is linear C1-C24 alkylene or linear C1-C24 alkenylene. [0715] In some other foregoing embodiments of Formula (III), R1 or R2, or both, is C6-C24 alkenyl. For example, in some embodiments, R1 and R2 each, independently have the following structure: , wherein:
Figure imgf000366_0002
R7a and R7b are, at each occurrence, independently H or C1-C12 alkyl; and a is an integer from 2 to 12, and wherein R7a, R7b and a are each selected such that R1 and R2 each independently comprise from 6 to 20 carbon atoms. For example, in some embodiments a is an integer ranging from 5 to 9 or from 8 to 12. [0716] In some of the foregoing embodiments of Formula (III), at least one occurrence of R7a is H. For example, in some embodiments, R7a is H at each occurrence. In other different embodiments of the foregoing, at least one occurrence of R7b is C1-C8 alkyl. For example, in some embodiments, C1-C8 alkyl is methyl, ethyl, n-propyl, iso-propyl, n-butyl, iso-butyl, tert-butyl, n-hexyl or n-octyl. [0717] In different embodiments of Formula (III), R1 or R2, or both, has one of the following structures: ; =O)R4 or –
Figure imgf000367_0001
NHC(=O)R4. In some embodiments, R4 is methyl or ethyl. [0719] In various different embodiments, the cationic lipid of Formula (III) has one of the structures set forth in in Table 21 below. Table 21: Example Compounds of Formula (III). No. Structure
Figure imgf000367_0002
No. Structure
Figure imgf000368_0001
No. Structure
Figure imgf000369_0001
No. Structure
Figure imgf000370_0001
No. Structure
Figure imgf000371_0001
No. Structure [0720] In va
Figure imgf000372_0002
rious different embodiments, a cationic lipid has one of the structures set forth in Table 22 below. Table 22: Example Cationic Lipid Structures No. Structure
Figure imgf000372_0001
No. Structure H O O N [0721] In
Figure imgf000373_0002
so e e o e s, a co p ses a cao c p a s a o za e lipid-like material (lipidoid). In some embodiments, a cationic lipid has the following structure: [0722]
Figure imgf000373_0001
, p p v v g .g., ameter) of about 30 nm to about 150 nm, about 40 nm to about 150 nm, about 50 nm to about 150 nm, about 60 nm to about 130 nm, about 70 nm to about 110 nm, about 70 nm to about 100 nm, about 70 to about 90 nm, or about 70 nm to about 80 nm. In some embodiments, lipid nanoparticles in accordance with the present disclosure can have an average size (e.g., mean diameter) of about 50 nm to about 100 nm. In some embodiments, lipid nanoparticles may have an average size (e.g., mean diameter) of about 50 nm to about 150 nm. In some embodiments, lipid nanoparticles may have an average size (e.g., mean diameter) of about 60 nm to about 120 nm. In some embodiments, lipid nanoparticles in accordance with the present disclosure can have an average size (e.g., mean diameter) of about 30 nm, 35 nm, 40 nm, 45 nm, 50 nm, 55 nm, 60 nm, 65 nm, 70 nm, 75 nm, 80 nm, 85 nm, 90 nm, 95 nm, 100 nm, 105 nm, 110 nm, 115 nm, 120 nm, 125 nm, 130 nm, 135 nm, 140 nm, 145 nm, or 150 nm. The term “average diameter” or “mean diameter” refers to the mean hydrodynamic diameter of particles as measured by dynamic laser light scattering (DLS) with data analysis using the so-called cumulant algorithm, which provides as results the so-called Z-average with the dimension of a length, and the polydispersity index (PI), which is dimensionless (Koppel, D., J. Chem. Phys.57, 1972, pp 4814-4820, ISO 13321, which is herein incorporated by reference). Here “average diameter,” “mean diameter,” “diameter,” or “size” for particles is used synonymously with this value of the Z-average. [0723] In some embodiments, lipid nanoparticles described herein may exhibit a polydispersity index less than about 0.5, less than about 0.4, less than about 0.3, or about 0.2 or less. By way of example, lipid nanoparticles can exhibit a polydispersity index in a range of about 0.1 to about 0.3 or about 0.2 to about 0.3. The “polydispersity index” is preferably calculated based on dynamic light scattering measurements by the so-called cumulant analysis as mentioned in the definition of the “average diameter.” Under certain prerequisites, it can be taken as a measure of the size distribution of an ensemble of ribonucleic acid nanoparticles (e.g., ribonucleic acid nanoparticles). [0724] Lipid nanoparticles described herein can be characterized by an “N/P ratio,” which is the molar ratio of cationic (nitrogen) groups (the “N” in N/P) in the cationic polymer to the anionic (phosphate) groups (the “P” in N/P) in RNA. It is understood that a cationic group is one that is either in cationic form (e.g., N+), or one that is ionizable to become cationic. Use of a single number in an N/P ratio (e.g., an N/P ratio of about 5) is intended to refer to that number over 1, e.g., an N/P ratio of about 5 is intended to mean 5:1. In some embodiments, a lipid nanoparticle described herein has an N/P ratio greater than or equal to 5. In some embodiments, a lipid nanoparticle described herein has an N/P ratio that is about 5, 6, 7, 8, 9, or 10. In some embodiments, an N/P ratio for a lipid nanoparticle described herein is from about 10 to about 50. In some embodiments, an N/P ratio for a lipid nanoparticle described herein is from about 10 to about 70. In some embodiments, an N/P ratio for a lipid nanoparticle described herein is from about 10 to about 120. B. Example Methods of Making Lipid Nanoparticles [0725] Lipids and lipid nanoparticles comprising nucleic acids and their method of preparation are known in the art, including, e.g., as described in U.S. Patent Nos. 8,569,256, 5,965,542 and U.S. Patent Publication Nos. 2016/0199485, 2016/0009637, 2015/0273068, 2015/0265708, 2015/0203446, 2015/0005363, 2014/0308304, 2014/0200257, 2013/086373, 2013/0338210, 2013/0323269, 2013/0245107, 2013/0195920, 2013/0123338, 2013/0022649, 2013/0017223, 2012/0295832, 2012/0183581, 2012/0172411, 2012/0027803, 2012/0058188, 2011/0311583, 2011/0311582, 2011/0262527, 2011/0216622, 2011/0117125, 2011/0091525, 2011/0076335, 2011/0060032, 2010/0130588, 2007/0042031, 2006/0240093, 2006/0083780, 2006/0008910, 2005/0175682, 2005/017054, 2005/0118253, 2005/0064595, 2004/0142025, 2007/0042031, 1999/009076 and PCT Pub. Nos. WO 99/39741, WO 2018/081480, WO 2017/004143, WO 2017/075531, WO 2015/199952, WO 2014/008334, WO 2013/086373, WO 2013/086322, WO 2013/016058, WO 2013/086373, W02011/141705, and WO 2001/07548, the full disclosures each of which are herein incorporated by reference in their entirety for the purposes described herein. [0726] For example, in some embodiments, cationic lipids, neutral lipids (e.g., DSPC, and/or cholesterol) and polymer-conjugated lipids can be solubilized in ethanol at a pre-determined molar ratio (e.g., ones described herein). In some embodiments, lipid nanoparticles (lipid nanoparticle) are prepared at a total lipid to polyribonucleotides weight ratio of approximately 10: 1 to 30: 1. In some embodiments, such polyribonucleotides can be diluted to 0.2 mg/mL in acetate buffer. [0727] In some embodiments, using an ethanol injection technique, a colloidal lipid dispersion comprising polyribonucleotides can be formed as follows: an ethanol solution comprising lipids, such as cationic lipids, neutral lipids, and polymer-conjugated lipids, is injected into an aqueous solution comprising polyribonucleotides (e.g., ones described herein). [0728] In some embodiments, lipid and polyribonucleotide solutions can be mixed at room temperature by pumping each solution at controlled flow rates into a mixing unit, for example, using piston pumps. In some embodiments, the flow rates of a lipid solution and a RNA solution into a mixing unit are maintained at a ratio of 1:3. Upon mixing, nucleic acid-lipid particles are formed as the ethanolic lipid solution is diluted with aqueous polyribonucleotides. The lipid solubility is decreased, while cationic lipids bearing a positive charge interact with the negatively charged RNA. [0729] In some embodiments, a solution comprising RNA-encapsulated lipid nanoparticles can be processed by one or more of concentration adjustment, buffer exchange, formulation, and/or filtration. [0730] In some embodiments, RNA-encapsulated lipid nanoparticles can be processed through filtration. [0731] In some embodiments, particle size and/or internal structure of lipid nanoparticles (with or without RNAs) may be monitored by appropriate techniques such as, e.g., small-angle X-ray scattering (SAXS) and/or transmission electron cryomicroscopy (CryoTEM). V. Pharmaceutical Compositions [0732] The present disclosure provides compositions, e.g., pharmaceutical compositions comprising one or more polyribonucleotides described herein. Pharmaceutical formulations may additionally comprise a pharmaceutically acceptable excipient, which, as used herein, includes any and all solvents, dispersion media, diluents, or other liquid vehicles, dispersion or suspension aids, surface active agents, isotonic agents, thickening or emulsifying agents, preservatives, solid binders, lubricants and the like, as suited to the particular dosage form desired. Remington's The Science and Practice of Pharmacy, 21st Edition, A. R. Gennaro (Lippincott, Williams & Wilkins, Baltimore, MD, 2006; incorporated herein by reference) discloses various excipients used in formulating pharmaceutical compositions and known techniques for the preparation thereof. Except insofar as any conventional excipient medium is incompatible with a substance or its derivatives, such as by producing any undesirable biological effect or otherwise interacting in a deleterious manner with any other component(s) of the pharmaceutical composition, its use is contemplated to be within the scope of this disclosure. [0733] In some embodiments, an excipient is approved for use in humans and for veterinary use. In some embodiments, an excipient is approved by the United States Food and Drug Administration. In some embodiments, an excipient is pharmaceutical grade. In some embodiments, an excipient meets the standards of the United States Pharmacopoeia (USP), the European Pharmacopoeia (EP), the British Pharmacopoeia, and/or the International Pharmacopoeia. [0734] Pharmaceutically acceptable excipients used in the manufacture of pharmaceutical compositions include, but are not limited to, inert diluents, dispersing and/or granulating agents, surface active agents and/or emulsifiers, disintegrating agents, binding agents, preservatives, buffering agents, lubricating agents, and/or oils. Such excipients may optionally be included in pharmaceutical formulations. Excipients such as cocoa butter and suppository waxes, coloring agents, coating agents, sweetening, flavoring, and/or perfuming agents can be present in the composition, according to the judgment of the formulator. [0735] General considerations in the formulation and/or manufacture of pharmaceutical agents may be found, for example, in Remington: The Science and Practice of Pharmacy 21st ed., Lippincott Williams & Wilkins, 2005 (incorporated herein by reference). [0736] In some embodiments, pharmaceutical compositions provided herein may be formulated with one or more pharmaceutically acceptable carriers or diluents as well as any other known adjuvants and excipients in accordance with conventional techniques such as those disclosed in Remington: The Science and Practice of Pharmacy 21st ed., Lippincott Williams & Wilkins, 2005 (incorporated herein by reference). [0737] Pharmaceutical compositions described herein can be administered by appropriate methods known in the art. As will be appreciated by a skilled artisan, the route and/or mode of administration may depend on a number of factors, including, e.g., but not limited to stability and/or pharmacokinetics and/or pharmacodynamics of pharmaceutical compositions described herein. [0738] In some embodiments, pharmaceutical compositions described herein are formulated for parenteral administration, which includes modes of administration other than enteral and topical administration, usually by injection, and includes, without limitation, intravenous, intramuscular, intraarterial, intradermal, subcutaneous, subcuticular, or intraarticular injection and infusion. In preferred embodiments, pharmaceutical compositions described herein are formulated for intravenous, intramuscular, or subcutaneous administration. In particularly preferred embodiments, pharmaceutical compositions described herein are formulated for intramuscular administration. [0739] In some embodiments, pharmaceutical compositions described herein are formulated for intravenous administration. In some embodiments, pharmaceutically acceptable excipients that may be useful for intravenous administration include sterile aqueous solutions or dispersions and sterile powders for preparation of sterile injectable solutions or dispersions. [0740] Therapeutic compositions typically must be sterile and stable under the conditions of manufacture and storage. The composition can be formulated as a solution, microemulsion, lipid nanoparticles, or other ordered structure suitable to high drug concentration. The carrier can be a solvent or dispersion medium containing, for example, water, ethanol, polyol (for example, glycerol, propylene glycol, and liquid polyethylene glycol, and the like), and suitable mixtures thereof. Proper fluidity can be maintained, for example, by the use of surfactants. In many cases, it will be preferable to include isotonic agents, for example, sugars, polyalcohols such as mannitol, sorbitol, or sodium chloride in the composition. In some embodiments, prolonged absorption of the injectable compositions can be brought about by including in the composition an agent that delays absorption, for example, monostearate salts and gelatin. [0741] Sterile injectable solutions can be prepared by incorporating the active compound in the required amount in an appropriate solvent with one or a combination of ingredients enumerated above, as required, followed by sterilization and/or microfiltration. In some embodiments, pharmaceutical compositions can be prepared as described herein and/or methods known in the art. In some embodiments, a pharmaceutical composition includes ALC-0315; ALC-0159; DSPC; Cholesterol; Sucrose; NaCl; KCl; Na2HPO4; KH2PO4; Water for injection. In some embodiments, normal saline (isotonic 0.9% NaCl) is used as diluent. [0742] These compositions may also contain adjuvants such as preservatives, wetting agents, emulsifying agents and dispersing agents. Prevention of the presence of microorganisms may be ensured both by sterilization procedures, and by the inclusion of various antibacterial and antifungal agents, for example, paraben, chlorobutanol, phenol sorbic acid, and the like. It may also be desirable to include isotonic agents, such as sugars, sodium chloride, and the like into pharmaceutical compositions described herein. In addition, prolonged absorption of the injectable pharmaceutical form may be brought about by the inclusion of agents which delay absorption such as aluminum monostearate and gelatin. [0743] Formulations of pharmaceutical compositions described herein may be prepared by any method known or hereafter developed in the art of pharmacology. In general, such preparatory methods include the step of bringing active ingredient(s) into association with a diluent or another excipient and/or one or more other accessory ingredients, and then, if necessary and/or desirable, shaping and/or packaging the product into a desired single- or multi-dose unit. [0744] A pharmaceutical composition in accordance with the present disclosure may be prepared, packaged, and/or sold in bulk, as a single unit dose, and/or as a plurality of single unit doses. As used herein, a “unit dose” is discrete amount of the pharmaceutical composition comprising a predetermined amount of at least one RNA product produced using a system and/or method described herein. [0745] Relative amounts of polyribonucleotides encapsulated in lipid nanoparticles, a pharmaceutically acceptable excipient, and/or any additional ingredients in a pharmaceutical composition can vary, depending upon the subject to be treated, target cells, diseases or disorders, and may also further depend upon the route by which the composition is to be administered. [0746] In some embodiments, pharmaceutical compositions described herein are formulated into pharmaceutically acceptable dosage forms by conventional methods known to those of skill in the art. Actual dosage levels of the active ingredients (e.g., polyribonucleotides encapsulated in lipid nanoparticles) in the pharmaceutical compositions described herein may be varied so as to obtain an amount of the active ingredient which is effective to achieve the desired therapeutic response for a particular patient, composition, and mode of administration, without being toxic to the patient. The selected dosage level will depend upon a variety of pharmacokinetic factors including the activity of the particular compositions of the present disclosure employed, the route of administration, the time of administration, the rate of excretion of the particular compound being employed, the duration of the treatment, other drugs, compounds and/or materials used in combination with the particular compositions employed, the age, sex, weight, condition, general health and prior medical history of the patient being treated, and like factors well known in the medical arts. [0747] A physician having ordinary skill in the art can readily determine and prescribe the effective amount of the pharmaceutical composition required. For example, a physician could start doses of active ingredients (e.g., polyribonucleotides encapsulated in lipid nanoparticles) employed in the pharmaceutical composition at levels lower than that required in order to achieve the desired therapeutic effect and gradually increase the dosage until the desired effect is achieved. [0748] In some embodiments, a pharmaceutical composition is formulated (e.g., but not limited to, for intravenous, intramuscular, or subcutaneous administration) to deliver a dose of about 5 mg RNA/kg. [0749] In some embodiments, a pharmaceutical composition described herein may further comprise one or more additives, for example, in some embodiments that may enhance stability of such a composition under certain conditions. Examples of additives may include but are not limited to salts, buffer substances, preservatives, and carriers. For example, in some embodiments, a pharmaceutical composition may further comprise a cryoprotectant (e.g., sucrose) and/or an aqueous buffered solution, which may in some embodiments include one or more salts, including, e.g., alkali metal salts or alkaline earth metal salts such as, e.g., sodium salts, potassium salts, and/or calcium salts. [0750] In some embodiments, a pharmaceutical composition provided herein is a preservative-free, sterile RNA-lipid nanoparticle dispersion in an aqueous buffer for intravenous or intramuscular administration. [0751] Although the descriptions of pharmaceutical compositions provided herein are principally directed to pharmaceutical compositions that are suitable for administration to humans, it will be understood by the skilled artisan that such compositions are generally suitable for administration to animals of all sorts. Modification of pharmaceutical compositions suitable for administration to humans in order to render the compositions suitable for administration to various animals is well understood, and the ordinarily skilled veterinary pharmacologist can design and/or perform such modification with merely ordinary, if any, experimentation. A. Certain Example Pharmaceutical Compositions [0752] Provided herein are combinations comprising a plurality of polyribonucleotides, wherein the plurality of polyribonucleotides comprises a first set of polyribonucleotides that encode one or more glycoprotein (GP) polypeptides as described herein. In some embodiments, a combination comprises one polyribonucleotide that encodes a (GP) polypeptide. In some embodiments, a combination comprises two or more polyribonucleotides that encode one or more (GP) polypeptides. In some embodiments, a combination comprises three or more polyribonucleotides that encode three or more (GP) polypeptides. In some embodiments, a combination comprises four or more polyribonucleotides that encode four or more (GP) polypeptides. In some embodiments, a combination comprises three or more polyribonucleotides that encode three or more (GP) polypeptides and a polyribonucleotide that encodes a T-cell string polypeptide. [0753] Also provided herein are combinations that comprise two or more pharmaceutical compositions, wherein each pharmaceutical composition comprises one or more polyribonucleotides as described herein. Also provided herein are combinations that comprise three or more pharmaceutical compositions, wherein each pharmaceutical composition comprises one or more polyribonucleotides as described herein. [0754] Provided herein are combinations that comprise two or more RNA constructs as described herein. In some embodiments, a combination comprises three or more RNA constructs as described herein. In some embodiments, a combination comprises four or more RNA constructs as described herein. In some embodiments, a combination comprises five or more RNA constructs as described herein. [0755] Also provided herein are combinations that comprise two or more pharmaceutical compositions, wherein each pharmaceutical composition comprises one or more RNA constructs as described herein. Also provided herein are combinations that comprise three or more pharmaceutical compositions, wherein each pharmaceutical composition comprises one or more RNA constructs as described herein. Also provided herein are combinations that comprise four or more pharmaceutical compositions, wherein each pharmaceutical composition comprises one or more RNA constructs as described herein. [0756] In some embodiments, a combination comprises a plurality of polyribonucleotides comprising a first set of polyribonucleotides that comprises a polyribonucleotide encoding a GP polypeptide comprising an HSV-2 gC or antigenic portion thereof, and a polyribonucleotide encodes a GP polypeptide comprising an HSV-2 gD or antigenic portion thereof. [0757] In some embodiments, a combination comprises a plurality of polyribonucleotides comprising a first set of polyribonucleotides that comprises a polyribonucleotide encoding a GP polypeptide comprising an antigenic portion of HSV-2 gC, and a polyribonucleotide encoding a GP polypeptide comprising an antigenic portion of HSV-2 gD. In some embodiments, a GP polypeptide comprising an antigenic portion of HSV-2 gC comprises an amino acid sequence according to SEQ ID NO: 65. In some embodiments, a GP polypeptide comprising an antigenic portion of HSV-2 gD comprises an amino acid sequence according to SEQ ID NO: 70. [0758] In some embodiments, a combination comprises (i) a first pharmaceutical composition comprising a polyribonucleotide that encodes a GP polypeptide comprising an HSV-2 gC or antigenic portion thereof, and (ii) a second pharmaceutical composition comprising a polyribonucleotide that encodes a GP polypeptide comprising an HSV-2 gD antigen or antigenic portion thereof. [0759] In some embodiments, a combination comprises (i) a first pharmaceutical composition comprising a first polyribonucleotide that encodes a GP polypeptide comprising an antigenic portion of HSV-2 gC, and (ii) a second pharmaceutical composition comprising a polyribonucleotide that encodes a GP comprising an antigenic portion of HSV-2 gD. In some embodiments, a GP polypeptide comprising an antigenic portion of HSV-2 gC comprises an amino acid sequence according to SEQ ID NO: 65. In some embodiments, a GP polypeptide comprising an antigenic portion of HSV-2 gD comprises an amino acid sequence according to SEQ ID NO: 70. In some embodiments, a GP polypeptide comprising an antigenic portion of HSV-2 gC comprises an amino acid sequence according to SEQ ID NO: 159. In some embodiments, a GP polypeptide comprising an antigenic portion of HSV-2 gD comprises an amino acid sequence according to SEQ ID NO: 70. [0760] In some embodiments, a combination comprises a plurality of polyribonucleotides comprising a first set of polyribonucleotides that comprises a polyribonucleotide encoding a GP polypeptide comprising an HSV-2 gC or antigenic portion thereof, and a polyribonucleotide encodes a GP polypeptide comprising an HSV-2 gE or antigenic portion thereof. [0761] In some embodiments, a combination comprises a plurality of polyribonucleotides comprising a first set of polyribonucleotides that comprises a polyribonucleotide encoding a GP polypeptide comprising an antigenic portion of HSV-2 gC, and a polyribonucleotide encoding a GP polypeptide comprising an antigenic portion of HSV-2 gE. In some embodiments, a GP polypeptide comprising an antigenic portion of HSV-2 gC comprises an amino acid sequence according to SEQ ID NO: 65. In some embodiments, a GP polypeptide comprising an antigenic portion of HSV-2 gE comprises an amino acid sequence according to SEQ ID NO: 73. [0762] In some embodiments, a combination comprises (i) a first pharmaceutical composition comprising a polyribonucleotide that encodes a GP polypeptide comprising an HSV-2 gC or antigenic portion thereof, and (ii) a second pharmaceutical composition comprising a polyribonucleotide that encodes a GP polypeptide comprising an HSV-2 gE antigen or antigenic portion thereof. [0763] In some embodiments, a combination comprises (i) a first pharmaceutical composition comprising a first polyribonucleotide that encodes a GP polypeptide comprising an antigenic portion of HSV-2 gC, and (ii) a second pharmaceutical composition comprising a polyribonucleotide that encodes a GP comprising an antigenic portion of HSV-2 gE. In some embodiments, a GP polypeptide comprising an antigenic portion of HSV-2 gC comprises an amino acid sequence according to SEQ ID NO: 65. In some embodiments, a GP polypeptide comprising an antigenic portion of HSV-2 gE comprises an amino acid sequence according to SEQ ID NO: 73.In some embodiments, a combination comprises a plurality of polyribonucleotides comprising a first set of polyribonucleotides that comprises a polyribonucleotide encoding a GP polypeptide comprising an HSV-2 gD or antigenic portion thereof, and a polyribonucleotide encodes a GP polypeptide comprising an HSV-2 gE or antigenic portion thereof. [0764] In some embodiments, a combination comprises a plurality of polyribonucleotides comprising a first set of polyribonucleotides that comprises a polyribonucleotide encoding a GP polypeptide comprising an antigenic portion of HSV-2 gD, and a polyribonucleotide encoding a GP polypeptide comprising an antigenic portion of HSV-2 gE. In some embodiments, a GP polypeptide comprising an antigenic portion of HSV-2 gD comprises an amino acid sequence according to SEQ ID NO: 70. In some embodiments, a GP polypeptide comprising an antigenic portion of HSV-2 gE comprises an amino acid sequence according to SEQ ID NO: 73. [0765] In some embodiments, a combination comprises (i) a first pharmaceutical composition comprising a polyribonucleotide that encodes a GP polypeptide comprising an HSV-2 gD or antigenic portion thereof, and (ii) a second pharmaceutical composition comprising a polyribonucleotide that encodes a GP polypeptide comprising an HSV-2 gE antigen or antigenic portion thereof. [0766] In some embodiments, a combination comprises (i) a first pharmaceutical composition comprising a first polyribonucleotide that encodes a GP polypeptide comprising an antigenic portion of HSV-2 gD, and (ii) a second pharmaceutical composition comprising a polyribonucleotide that encodes a GP comprising an antigenic portion of HSV-2 gE. In some embodiments, a GP polypeptide comprising an antigenic portion of HSV-2 gD comprises an amino acid sequence according to SEQ ID NO: 70. In some embodiments, a GP polypeptide comprising an antigenic portion of HSV-2 gE comprises an amino acid sequence according to SEQ ID NO: 73. [0767] In some embodiments, a GP polypeptide that comprises an amino acid sequence according to SEQ ID NO: 65 is encoded by a ribonucleic acid sequence according to 106. In some embodiments, a GP polypeptide that comprises an amino acid sequence according to SEQ ID NO: 70 is encoded by a ribonucleic acid sequence according to 118. In some embodiments, a GP polypeptide that comprises an amino acid sequence according to SEQ ID NO: 73 is encoded by a ribonucleic acid sequence according to 123. VI. Patient Populations [0768] In some embodiments, technologies of the present disclosure are used for therapeutic and/or prophylactic purposes. In some embodiments, technologies of the present disclosure are used in the treatment and/or prophylactic of an HSV infection. Prophylactic purposes of the present disclosure comprise pre-exposure prophylaxis and/or post-exposure prophylaxis. [0769] In some embodiments, technologies of the present disclosure are used in the treatment and/or prophylaxis of a disorder related to such an HSV (e.g., HSV-1 and/or HSV-2) infection. A disordered related to such an HSV (e.g., HSV-1 and/or HSV-2) infection comprises, for example, a typical symptom and/or a complication of an HSV (e.g., HSV-1 and/or HSV-2) infection. [0770] In some embodiments, provided compositions (e.g., that are or comprise one or more GP polypeptides and/or one or more T-cell string polypeptides) may be useful to detect and/or characterize one or more features of an anti-HSV (e.g., anti-HSV-1 and/or anti-HSV-2) immune response (e.g., by detecting binding to a provided antigen by serum from an infected subject). [0771] In some embodiments, provided compositions (e.g., that are or comprise one or more GP polypeptides and/or one or more T-cell string polypeptides) are useful to raise antibodies to one or more epitopes included therein; such antibodies may themselves be useful, for example for detection or treatment of an HSV infection. [0772] The present disclosure provides use of encoding nucleic acids (e.g., DNA or RNA) to produce encoded antigens and/or use of DNA constructs to produce RNA. [0773] In some embodiments, technologies of the present disclosure are utilized in a non-limited subject population; in some embodiments, technologies of the present disclosure are utilized in particular subject populations. [0774] In some embodiments, a subject population comprises an adult population. In some embodiments, an adult population comprises subjects between the ages of about 18 years and about 55 years of age (e.g., about 19, 20, 25, 30, 35, 40, 45, 50, 51, 52, 53, 54, or 55,). [0775] In some embodiments, a subject population comprises an elderly population. In some embodiments, an elderly population comprises subjects of, about 56 years of age, about 60 years of age, about 70 years of age, or older (e.g., about 60, 65, 70, 75, 80, 85, 90, 95, or 100 years of age). [0776] In some embodiments, a subject has a weight of at least about 50 kg. In some embodiments, a subject has a weight of at least about 51 kg (e.g., about 52, 53, 54, 55, 56,57, 58, 59, 60 kg). [0777] In some embodiments, a subject has a body mass index (BMI) in a range of about 17.5 kg/m2 to about 37 kg/m2, such as about 18 kg/m2 to about 36 kg/m2, such as about 18.5 kg/m2 to about 35 kg/m2. In some embodiments, a subject has a BMI of at least 17 kg/m2, such as at least 17.5 kg/m2, such as at least 18 kg/m2, such as at least 18.5 kg/m2. In some embodiments, a subject has a BMI of at the most 40 kg/m2, such as at the most 39 kg/m2, such as at the most 38 kg/m2, such as at the most 37 kg/m2, such as at the most 36 kg/m2, such as at the most 35 kg/m2. [0778] In some embodiments, a subject population comprises a pediatric population. In some embodiments, a pediatric population comprises subjects approximately 18 years old or younger. In some such embodiments, a pediatric population comprises subjects between the ages of about 1 year and about 18 years (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, or 18 years of age). [0779] In some embodiments, a subject population comprises a newborn population. In some embodiments, a newborn population comprises subjects about 12 months or younger (e.g., 12, 11, 10, 9, 8, 7, 6, 5, 4, 3, 2, 1 months or younger). In some embodiments, subject populations to be treated with technologies described herein include infants (e.g., about 12 months or younger) whose mothers did not receive such technologies described herein during pregnancy. In some embodiments, subject populations to be treated with technologies described herein may include pregnant women; in some embodiments, infants whose mothers were treated with disclosed technologies during pregnancy (e.g., who received at least one dose, or alternatively only who received both doses), are not vaccinated during the first weeks, months, or even years (e.g., 1, 2, 3, 4, 5, 6, 7, 8 weeks or more, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24 months or more, or 1, 2, 3, 4, 5 years or more) post-birth. Alternatively or additionally, in some embodiments, infants whose mothers were treated with disclosed technologies during pregnancy (e.g., who received at least one dose, or alternatively only who received both doses), receive reduced treated with disclosed technologies (e.g., lower doses and/or smaller numbers of administrations – e.g., boosters – and/or lower total exposure over a given period of time) after birth, for example during the first weeks, months, or even years (e.g., 1, 2, 3, 4, 5, 6, 7, 8 weeks or more, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24 months or more, or 1, 2, 3, 4, 5 years or more) post-birth or may need reduced vaccination (e.g., lower doses and/or smaller numbers of administrations – e.g., boosters – over a given period of time), In some embodiments, compositions as provided herein are administered to subject populations that do not include pregnant women. [0780] In some embodiments, a subject population is or comprises children aged 6 weeks to up to 17 months of age. [0781] In some embodiments, a provided pharmaceutical composition (e.g., immunogenic composition, e.g., vaccine) may be administered in combination with (i.e., so that subject(s) are simultaneously exposed to both) another pharmaceutical composition (e.g., immunogenic composition, e.g., vaccine) or therapeutic intervention, e.g., to treat or prevent an HSV infection, or another disease, disorder, or condition. [0782] In some embodiments, a provided pharmaceutical composition (e.g., immunogenic composition, e.g., vaccine) may be administered with a protein vaccine, a DNA vaccine, an RNA vaccine, a cellular vaccine, a conjugate vaccine, etc. In some embodiments, one or more doses of a provided pharmaceutical composition (e.g., immunogenic composition, e.g., vaccine) may be administered together with (e.g., in a single visit) another composition (e.g., vaccine) or other therapy. [0783] In some embodiments, a provided pharmaceutical composition (e.g., immunogenic composition, e.g., vaccine) may be administered to subjects who have been exposed, or expect they have been exposed, to HSV (e.g., HSV-1 and/or HSV-2). In some embodiments, a provided pharmaceutical composition (e.g., immunogenic composition, e.g., vaccine) may be administered to subjects who do not have symptoms of an HSV (e.g., HSV-1 and/or HSV-2). [0784] In some embodiments, a subject has no prior history of known or suspected herpes simplex vaccination prior to administration of one or more doses of a composition as disclosed herein. [0785] In some embodiments, a subject does not have febrile illness prior to administration of one or more doses of a composition as disclosed herein. In some embodiments, a subject does not have febrile illness about 72 hours, about 48 hours, about 36 hours, about 24 hours, or about 12 hours prior to administration of one or more doses of a composition as disclosed herein. [0786] In some embodiments, a subject does not have an acute illness prior to administration of one or more doses of a composition as disclosed herein. In some embodiments, a subject does not have an acute illness about 72 hours, about 48 hours, about 36 hours, about 24 hours, or about 12 hours prior to administration of one or more doses of a composition as disclosed herein. [0787] In some embodiments, a subject has not received a vaccine 0 to 300 days, 0 to 290 days, 0 to 280 days, 0 to 270 days, 0 to 260 days, 0 to 250 days, 0 to 240 days, 0 to 230 days, 0 to 220 days, 0 to 210, 0 to 200 days, 0 to 190 days, 0 to 180 days, 0 to 170 days, 0 to 160 days, 0 to 150 days, 0 to 140 days, 0 to 130 days, 0 to 120 days, 0 to 110 days, 0 to 100 days, 0 to 90 days, 0 to 80 days, 0 to 70 days, 0 to 60 days, 0 to 50 days, 0 to 40 days, 0 to 35 days, 0 to 30 days, 0 to 29 days, 0 to 28 days before being administered one or more doses of a therapeutically effective amount of a composition as disclosed herein. In some embodiments, a subject has not received a vaccine about 7 days, about 14 days, about 21 days, about 28 days, about 35 days, about 42 days, about 49 days, about 56 days, about 60 days, about 70 days, about 80 days, about 90 days, about 100 days, about 125 days, about 150 days, about 175 days, about 190 days, about 200 days, about 210 days, or about 210 days before being administered one or more doses of a therapeutically effective amount of a composition as disclosed herein. In some embodiments, a vaccine is not a seasonal influenza vaccine or a medically indicated vaccine. [0788] In some embodiments, a subject does not receive a vaccine at least 2 weeks to 35 weeks, at least 3 weeks to 34 weeks, or at least 4 weeks to 33 weeks after administration of one or more doses of a therapeutically effective amount of a composition as disclosed herein. In some embodiments, a subject does not receive a vaccine at least about 2 weeks, at least about 4 weeks, at least about 6 weeks, at least about 8 weeks, at least about 10 weeks, at least about 12 weeks, at least about 14 weeks, at least about 16 weeks, at least about 18 weeks, at least about 20 weeks, at least about 22 weeks, at least about 24 weeks, at least about 26 weeks, at least about 28 weeks, or at least about 30 weeks after administration of one or more doses of a therapeutically effective amount of a composition as disclosed herein. In some embodiments, the vaccine is not a seasonal influenza vaccine or a medically indicated vaccine. [0789] In some embodiments, a subject has not received blood, plasma products, or immunoglobulins about 0 to 600 days, about 0 to 590 days, about 0 to 580 days, about 0 to 570 days, about 0 to 560 days, about 0 to 550 days, or about 0 to 545 days before administration of one or more doses of a therapeutically effective amount of a composition disclosed herein. [0790] In some embodiments, a subject has not received an allergy treatment 8 to 45 days, 12 to 40 days, 16 to 38 days, 21 to 35 days, 23 to 32 days, 25 to 30 days, or 26 to 29 days before administration of one or more doses of a therapeutically effective amount of a composition disclosed herein. In some embodiments, a subject has not received an allergy treatment about 14 days, about 16 days, about 18 days, about 20 days, about 22 days, about 24 days, about 26 days, about 28 days before administration of one or more doses of a therapeutically effective amount of a composition disclosed herein. In some embodiments, an allergy treatment comprises antigen injections. [0791] In some embodiments, a subject has not received an immunosuppressive medication 7 to 56 days, 14 to 56 days, 21 to 56 days, 28 to 56 days, 35 to 56 days, 42 to 56 days, 49 to 56 days, 7 to 49 days, 14 to 49 days, 21 to 49 days, 28 to 49 days, 35 to 49 days, 42 to 49 days, 7 to 42 days, 14 to 42 days, 21 to 42 days, 28 to 42 days, 35 to 42 days, 7 to 35 days, 14 to 35 days, 21 to 35 days, 28 to 35 days, 7 to 28 days, 14 to 28 days, 21 to 28 days, 7 to 21 days, 14 to 21 days, or 7 to 14 days before administration of one or more doses of a therapeutically effective amount of a composition disclosed herein. In some embodiments, a subject has not received an immunosuppressive medication about 7 days, about 14 days, about 21 days, about 28 days, about 35 days, about 42 days, about 49 days, or about 56 days before administration of one or more doses of a therapeutically effective amount of a composition disclosed herein. In some embodiments, a subject has not received an immunosuppressive medication about 28 days before administration of one or more doses of a therapeutically effective amount of a composition disclosed herein. [0792] In some embodiments, an immunosuppressive medication comprises a systemic corticosteroid or radiotherapy. In some embodiments, a systemic coriticosteroid is selected from, but not limited to, methylprednisolone, dexamethasone, hydrocortisone, prednisone, prednisolone, fluticasone, flumethasone, fluocinolone, budesonide, beclomethasone, ciclesonide, cortisone, triamcinolone, betamethasone, deflazacort, difluprednate, loteprednol, paramethasone, tixocortol, aldosterone, cloprednol, cortivazol, deoxycortone, desonide, desoximetasone, difluorocortolone, fluclorolone, fludrocortisone, flunisolide, fluocinonide, fluocortin butyl, fluorocortisone, fluorocortolone, fluorometholone, flurandrenolone, halcinonide, icomethasone, meprednisone, mometasone, rofleponide, RPR 106541, and their respective pharmaceutically acceptable derivatives, such as beclomethasone dipropionate (anhydrous or monohydrate), beclomethasone monopropionate, dexamethasone 21- isonicotinate, fluticasone propionate, icomethasone enbutate, tixocortol 21-pivalate, triamcinolone acetonide, and pharmaceutically acceptable salts and/or derivatives thereof. In some embodiments, a coriticosteroid is prednisone. [0793] In some embodiments, a subject has not received a prophylactic antipyretic and/or an analgesic medication 0 to 600 days, 0 to 550 days, 0 to 500 days, 0 to 500 days, 0 to 450 days, 0 to 400 days, 0 to 350 days, 0 to 300 days, 0 to 250 days, 0 to 200 days, 0 to 150 days, 0 to 150 days, 0 to 150 days, or 0 to 50 days before administration of one or more doses of a therapeutically effective amount of a composition disclosed herein. [0794] In some embodiments, a prophylactic antipyretic medication is selected from, but not limited to, acetaminophen, a non-steroidal anti-inflammatory drug (NSAID), salicylamide, salicyl salicylate, methyl salicylate, magnesium salicylate, faislamine, ethenzamide, diflunisal, choline magnesium salicylate, benorylate/benorilatem and amoxiprin, acetylsalicylate, ceclofenac, acemetacin, alclofenac, bromfenac, diclofenac, etodolac, indomethacin, nabumetone, oxametacin, proglumetacin, sulindac, tolmetin, iminoprofen, benoxaprofen, carprofen, dexibuprofen, dexketoprofen, fenbufen, fenoprofen, flunoxaprofen, flurbiprofen, ibuprofen, ibuproxam, indoprofen, ketoprofen, ketorolac, loxoprofen, naproxen, oxaprozin, pirprofen, suprofen, tiaprofenic acid, mefenamic acid, flufenamic acid, meclofenamic acid, tolfenamic acid, droxicam, lornoxicam, meloxicam, piroxicam, tenoxicam, dipyrone, azapropazone, clofezone, kebuzone, metamizole, mofebutazone, oxyphenbutazone, phenazone, phenylbutazone, sulfinpyrazone, decoxib, rofecoxib, parecoxib, and etoricoxib. [0795] In some embodiments, a prophylactic analgesic medication is selected from, but not limited to, acetaminophen, salicylamide, salicyl salicylate, methyl salicylate, magnesium salicylate, faislamine, ethenzamide, diflunisal, choline magnesium salicylate, benorylate/benorilatem and amoxiprin, acetylsalicylate, ceclofenac, acemetacin, alclofenac, bromfenac, diclofenac, etodolac, indomethacin, nabumetone, oxametacin, proglumetacin, sulindac, tolmetin, iminoprofen, benoxaprofen, carprofen, dexibuprofen, dexketoprofen, fenbufen, fenoprofen, flunoxaprofen, flurbiprofen, ibuprofen, ibuproxam, indoprofen, ketoprofen, ketorolac, loxoprofen, naproxen, oxaprozin, pirprofen, suprofen, tiaprofenic acid, mefenamic acid, flufenamic acid, meclofenamic acid, tolfenamic acid, droxicam, lornoxicam, meloxicam, piroxicam, tenoxicam, dipyrone, azapropazone, clofezone, kebuzone, metamizole, mofebutazone, oxyphenbutazone, phenazone, phenylbutazone, sulfinpyrazone, decoxib, rofecoxib, parecoxib, etoricoxib, codeine, dihydrocodeine, morphine or a morphine derivative or pharmaceutically acceptable salt thereof, diacetylmorphine, hydrocodone, hydromorphone, levorphanol, oxymorphone, alfentanil, buprenorphine, butorphanol, fentanyl, sufentanil, meperidine, methadone, nalbuphine, propoxyphene, pentazocine, and pharmaceutically acceptable salts thereof. VII. Treatment or Prevention Methods [0796] In some embodiments, technologies of the present disclosure may be administered to subjects according to a particular dosing regimen. In some embodiments, a dosing regimen may involve a single administration; in some embodiments, a dosing regimen may comprise one or more “booster” administrations after the initial administration. In some embodiments, initial and boost doses are the same amount; in some embodiments they differ. In some embodiments, two or more booster doses are administered. In some embodiments, a plurality of doses are administered at regular intervals. In some embodiments, periods of time between doses become longer. In some embodiments, one or more subsequent doses is administered if a particular clinical (e.g., reduction in neutralizing antibody levels) or situational (e.g., local development of a new strain) even arises or is detected. [0797] In some embodiments, administered pharmaceutical compositions (e.g., immunogenic compositions, e.g., vaccines) comprising RNA constructs that encode HSV-2 gC, gD and/or gE constructs are administered in RNA doses of from about 0.1 μg to about 300 μg, about 0.5 μg to about 200 μg, or about 1 μg to about 100 μg, such as about 1 μg, about 3 μg, about 10 μg, about 30 μg, about 50 μg, or about 100 μg. In some embodiments, an saRNA construct is administered at a lower dose (e.g., 2, 4, 5, 10 fold or more lower) than a modRNA or uRNA construct. [0798] In some embodiments, a first booster dose is administered about six months of the initial dose, and preferably about 5, 4, 3, 2, or 1 months. In some embodiments, a first booster dose is administered in a time period that begins about 1, 2, 3, or 4 weeks after the first dose, and ends about 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12 weeks of the first dose (e.g., between about 1 and about 12 weeks after the first dose, or between about 2 or 3 weeks and about 5 and 6 weeks after the first dose, or about 3 weeks or about 4 weeks after the first dose). [0799] In some embodiments, a plurality of booster doses (e.g., 2, 3, or 4) doses are administered within 6 months of the first dose, or within 12 months of the first dose. [0800] In some embodiments, 3 doses or fewer are required to achieve effective vaccination (e.g., greater than 60%, and in some embodiments greater than about 70%, about 75%, about 80%, about 85%, about 90% or more) reduction in risk of infection, or of serious disease. In some embodiments, not more than two doses are required. In some embodiments, a single dose is sufficient. In some embodiments, an RNA dose is about 60 μg or lower, 50 μg or lower, 40 μg or lower, 30 μg or lower, 20 μg or lower, 10 μg or lower, 5 μg or lower, 2.5 μg or lower, or 1 μg or lower. In some embodiments, an RNA dose is about 0.25 μg, at least 0.5 μg, at least 1 μg, at least 2 μg, at least 3 μg, at least 4 μg, at least 5 μg, at least 10 μg, at least 20 μg, at least 30 μg, or at least 40 μg. In some embodiments, an RNA dose is about 0.25 μg to 60 μg, 0.5 μg to 55 μg, 1 μg to 50 μg, 5 μg to 40 μg, or 10 μg to 30 μg may be administered per dose. In some embodiments, an RNA dose is about 30 μg. In some embodiments, at least two such doses are administered. For example, a second dose may be administered about 21 days following administration of the first dose. In some embodiments, a first booster dose is administered about one month after an initial dose. In some such embodiments, at least one further booster is administered at one-month interval(s). In some embodiments, after 2 or 3 boosters, a longer interval is introduced and no further booster is administered for at least 6, 9, 12, 18, 24, or more months. In some embodiments, a single further booster is administered after about 18 months. In some embodiments, no further booster is required unless, for example, a material change in clinical or environmental situation is observed. VIII. Methods of Manufacture [0801] Individual polyribonucleotides can be produced by methods known in the art. For example, in some embodiments, polyribonucleotides can be produced by in vitro transcription, for example, using a DNA template. A plasmid DNA used as a template for in vitro transcription to generate a polyribonucleotide described herein is also within the scope of the present disclosure. [0802] A DNA template is used for in vitro RNA synthesis in the presence of an appropriate RNA polymerase (e.g., a recombinant RNA-polymerase such as a T7 RNA-polymerase) with ribonucleotide triphosphates (e.g., ATP, CTP, GTP, UTP). In some embodiments, polyribonucleotides (e.g., ones described herein) can be synthesized in the presence of modified ribonucleotide triphosphates. By way of example only, in some embodiments, pseudouridine (ψ), N1-methyl-pseudouridine (m1ψ), or 5-methyl-uridine (m5U) can be used to replace uridine triphosphate (UTP). In some embodiments, pseudouridine (ψ) can be used to replace uridine triphosphate (UTP). In some embodiments, N1-methyl-pseudouridine (m1ψ) can be used to replace uridine triphosphate (UTP). In some embodiments, 5-methyl-uridine (m5U) can be used to replace uridine triphosphate (UTP). [0803] As will be clear to those skilled in the art, during in vitro transcription, an RNA polymerase (e.g., as described and/or utilized herein) typically traverses at least a portion of a single-stranded DNA template in the 3'→ 5' direction to produce a single-stranded complementary RNA in the 5'→ 3' direction. [0804] In some embodiments where a polyribonucleotide comprises a polyA tail, one of those skill in the art will appreciate that such a polyA tail may be encoded in a DNA template, e.g., by using an appropriately tailed PCR primer, or it can be added to a polyribonucleotide after in vitro transcription, e.g., by enzymatic treatment (e.g., using a poly(A) polymerase such as an E. coli Poly(A) polymerase). Suitable poly(A) tails are described herein above. For example, in some embodiments, a poly(A) tail comprises a nucleotide sequence of AAAAAAAAAAAAAAAAAAAAAAAAAAAAAAGCATATGACTAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAA AAAAAAAAAAAAAAAAAAAAAAAA (SEQ ID NO: 153). In some embodiments, a poly(A) tail comprises a plurality of A residues interrupted by a linker. In some embodiments, a linker comprises the nucleotide sequence GCATATGAC (SEQ ID NO: 154). [0805] In some embodiments, those skilled in the art will appreciate that addition of a 5' cap to an RNA (e.g., mRNA) can facilitate recognition and attachment of the RNA to a ribosome to initiate translation and enhances translation efficiency. Those skilled in the art will also appreciate that a 5' cap can also protect an RNA product from 5' exonuclease mediated degradation and thus increases half-life. Methods for capping are known in the art; one of ordinary skill in the art will appreciate that in some embodiments, capping may be performed after in vitro transcription in the presence of a capping system (e.g., an enzyme-based capping system such as, e.g., capping enzymes of vaccinia virus). In some embodiments, a cap may be introduced during in vitro transcription, along with a plurality of ribonucleotide triphosphates such that a cap is incorporated into a polyribonucleotide during transcription (also known as co-transcriptional capping). In some embodiments, a GTP fed-batch procedure with multiple additions in the course of the reaction may be used to maintain a low concentration of GTP in order to effectively cap the RNA. Suitable 5' cap are described herein above. For example, in some embodiments, a 5' cap comprises m7(3'OMeG)(5')ppp(5')(2'OMeA)pG. [0806] Following RNA transcription, a DNA template is digested. In some embodiments, digestion can be achieved with the use of DNase I under appropriate conditions.
Figure imgf000387_0001
solution, for example, in a buffer such as HEPES, a phosphate buffer solution, a citrate buffer solution, an acetate buffer solution; in some embodiments, such solution may be buffered to a pH within a range of, for example, about 6.5 to about 7.5; in some embodiments approximately 7.0. In some embodiments, production of polyribonucleotides may further include one or more of the following steps: purification, mixing, filtration, and/or filling. [0808] In some embodiments, polyribonucleotides can be purified (e.g., in some embodiments after in vitro transcription reaction), for example, to remove components utilized or formed in the course of the production, like, e.g., proteins, DNA portions, and/or or nucleotides. Various nucleic acid purifications that are known in the art can be used in accordance with the present disclosure. Certain purification steps may be or include, for example, one or more of precipitation, column chromatography (including, e.g., but not limited to anionic, cationic, hydrophobic interaction chromatography (HIC)), solid substrate-based purification (e.g., magnetic bead-based purification). In some embodiments, polyribonucleotides may be purified using magnetic bead-based purification, which in some embodiments may be or comprise magnetic bead-based chromatography. In some embodiments, polyribonucleotides may be purified using hydrophobic interaction chromatography (HIC) and/or diafiltration. In some embodiments, polyribonucleotides may be purified using HIC followed by diafiltration. [0809] In some embodiments, dsRNA may be obtained as side product during in vitro transcription. In some such embodiments, a second purification step may be performed to remove dsRNA contamination. For example, in some embodiments, cellulose materials (e.g., microcrystalline cellulose) may be used to remove dsRNA contamination, for examples in some embodiments in a chromatographic format. In some embodiments, cellulose materials (e.g., microcrystalline cellulose) can be pretreated to inactivate potential RNase contamination, for example in some embodiments by autoclaving followed by incubation with aqueous basic solution, e.g., NaOH. In some embodiments, cellulose materials may be used to purify polyribonucleotides according to methods described in WO 2017/182524, the entire content of which is incorporated herein by reference. [0810] In some embodiments, a batch of polyribonucleotides may be further processed by one or more steps of filtration and/or concentration. For example, in some embodiments, polyribonucleotide(s), for example, after removal of dsRNA contamination, may be further subject to diafiltration (e.g., in some embodiments by tangential flow filtration), for example, to adjust the concentration of polyribonucleotides to a desirable RNA concentration and/or to exchange buffer to a drug substance buffer. [0811] In some embodiments, polyribonucleotides may be processed through 0.2 μm filtration before they are filled into appropriate containers. [0812] In some embodiments, polyribonucleotides and compositions thereof may be manufactured in accordance with a process as described herein, or as otherwise known in the art. [0813] In some embodiments, polyribonucleotides and compositions thereof may be manufactured at a large scale. For example, in some embodiments, a batch of polyribonucleotides can be manufactured at a scale of greater than 1 g, greater than 2 g, greater than 3 g, greater than 4 g, greater than 5 g, greater than 6 g, greater than 7 g, greater than 8 g, greater than 9 g, greater than 10 g, greater than 15 g, greater than 20 g, or higher. [0814] In some embodiments, RNA quality control may be performed and/or monitored at any time during production process of polyribonucleotides and/or compositions comprising the same. For example, in some embodiments, RNA quality control parameters, including one or more of RNA identity (e.g., sequence, length, and/or RNA natures), RNA integrity, RNA concentration, residual DNA template, and residual dsRNA, may be assessed and/or monitored after each or certain steps of a polyribonucleotide manufacturing process, e.g., after in vitro transcription, and/or each purification step. [0815] In some embodiments, the stability of polyribonucleotides (e.g., produced by in vitro transcription) and/or compositions comprising polyribonucleotides can be assessed under various test storage conditions, for example, at room temperatures vs. fridge or sub-zero temperatures over a period of time (e.g., at least 3 months, at least 6 months, at least 9 months, at least 12 months, or longer). In some embodiments, polyribonucleotides (e.g., ones described herein) and/or compositions thereof may be stored stable at a fridge temperature (e.g., about 4qC to about 10qC) for at least 1 month or longer including, at least 2 months, at least 3 months, at least 4 months, at least 5 months, at least 6 months, at least 7 months, at least 8 months, at least 9 months, at least 10 months, at least 11 months, or at least 12 months or longer. In some embodiments, polyribonucleotides (e.g., ones described herein) and/or compositions thereof may be stored stable at a sub-zero temperature (e.g., -20qC or below) for at least 1 month or longer including, at least 2 months, at least 3 months, at least 4 months, at least 5 months, at least 6 months, at least 7 months, at least 8 months, at least 9 months, at least 10 months, at least 11 months, or at least 12 months or longer. In some embodiments, polyribonucleotides (e.g., ones described herein) and/or compositions thereof may be stored stable at room temperature (e.g., at about 25°C) for at least 1 month or longer. [0816] In some embodiments, one or more assessments may be utilized during manufacture, or other preparation or use of polyribonucleotides (e.g., as a release test). [0817] In some embodiments, one or more quality control parameters may be assessed to determine whether polyribonucleotides described herein meet or exceed acceptance criteria (e.g., for subsequent formulation and/or release for distribution). In some embodiments, such quality control parameters may include, but are not limited to RNA integrity, RNA concentration, residual DNA template and/or residual dsRNA. Certain methods for assessing RNA quality are known in the art; for example, one of skill in the art will recognize that in some embodiments, one or more analytical tests can be used for RNA quality assessment. Examples of such certain analytical tests may include but are not limited to gel electrophoresis, UV absorption, and/or PCR assay. [0818] In some embodiments, a batch of polyribonucleotides may be assessed for one or more features as described herein to determine next action step(s). For example, a batch of polyribonucleotides can be designated for one or more further steps of manufacturing and/or formulation and/or distribution if RNA quality assessment indicates that such a batch of polyribonucleotides meet or exceed the relevant acceptance criteria. Otherwise, an alternative action can be taken (e.g., discarding the batch) if such a batch of polyribonucleotides does not meet or exceed the acceptance criteria. [0819] In some embodiments, a batch of polyribonucleotides that satisfy assessment results can be utilized for one or more further steps of manufacturing and/or formulation and/or distribution. IX. DNA Constructs [0820] Among other things, the present disclosure provides DNA constructs, for example that may encode one or more antibody agents as described herein, or components thereof. In some embodiments, DNA constructs provided by and/or utilized in accordance with the present disclosure are comprised in a vector. [0821] Non-limiting examples of a vector include plasmid vectors, cosmid vectors, phage vectors such as lambda phage, viral vectors such as retroviral, adenoviral or baculoviral vectors, or artificial chromosome vectors such as bacterial artificial chromosomes (BAC), yeast artificial chromosomes (YAC), or P1 artificial chromosomes (PAC). In some embodiments, a vector is an expression vector. In some embodiments, a vector is a cloning vector. In general, a vector is a nucleic acid construct that can receive or otherwise become linked to a nucleic acid element of interest (e.g., a construct that is or encodes a payload, or that imparts a particular functionality, etc.). [0822] Expression vectors, which may be plasmid or viral or other vectors, typically include an expressible sequence of interest (e.g., a coding sequence) that is functionally linked with one or more control elements (e.g., promoters, enhancers, transcription terminators, etc.). Typically, such control elements are selected for expression in a system of interest. In some embodiments, a system is ex vivo (e.g., an in vitro transcription system); in some embodiments, a system is in vivo (e.g., a bacterial, yeast, plant, insect, fish, vertebrate, mammalian cell or tissue, etc.). [0823] Cloning vectors are generally used to modify, engineer, and/or duplicate (e.g., by replication in vivo, for example in a simple system such as bacteria or yeast, or in vitro, such as by amplification such as polymerase chain reaction or other amplification process). In some embodiments, a cloning vector may lack expression signals. [0824] In many embodiments, a vector may include replication elements such as primer binding site(s) and/or origin(s) of replication. In many embodiments, a vector may include insertion or modification sites such as restriction endonuclease recognition sites and/or guide RNA binding sites, etc. [0825] In some embodiments, a vector is a viral vector (e.g., an AAV vector). In some embodiments, a vector is a non-viral vector. In some embodiments, a vector is a plasmid. [0826] Those skilled in the art are aware of a variety of technologies useful for the production of recombinant polynucleotides (e.g., DNA or RNA) as described herein. For example, restriction digestion, reverse transcription, amplification (e.g., by polymerase chain reaction), Gibson assembly, etc., are well established and useful tools and technologies. Alternatively or additionally, certain nucleic acids may be prepared or assembled by chemical and/or enzymatic synthesis. In some embodiments, a combination of known methods is utilized to prepare a recombinant polynucleotide. [0827] In some embodiments, polynucleotide(s) of the present disclosure are included in a DNA construct (e.g., a vector) amenable to transcription and/or translation. [0828] In some embodiments, an expression vector comprises a polynucleotide that encodes proteins and/or polypeptides of the present disclosure operatively linked to a sequence or sequences that control expression (e.g., promoters, start signals, stop signals, polyadenylation signals, activators, repressors, etc.). In some embodiments, a sequence or sequences that control expression are selected to achieve a desired level of expression. In some embodiments, more than one sequence that controls expression (e.g., promoters) are utilized. In some embodiments, more than one sequence that controls expression (e.g., promoters) are utilized to achieve a desired level of expression of a plurality of polynucleotides that encode a plurality of proteins and/or polypeptides. In some embodiments, a plurality of recombinant proteins and/or polypeptides are expressed from the same vector (e.g., a bi-cistronic vector, a tri-cistronic vector, multi-cistronic). In some embodiments, a plurality of polypeptides are expressed, each of which is expressed from a separate vector. [0829] In some embodiments, an expression vector comprising a polynucleotide of the present disclosure is used to produce an RNA and/or protein and/or polypeptide in a host cell. In some embodiments, a host cell may be in vitro (e.g., a cell line) – for example a cell or cell line (e.g., Human Embryonic Kidney (HEK cells), Chinese Hamster Ovary cells, etc.) suitable for producing polynucleotides of the present disclosure and proteins and/or polypeptides encoded by said polynucleotides. [0830] In some embodiments, an expression vector is an RNA expression vector. In some embodiments, an RNA expression vector comprises a polynucleotide template used to produce a RNA in cell-free enzymatic mix. In some embodiments, an RNA expression vector comprising a polynucleotide template is enzymatically linearized prior to in vitro transcription. In some embodiments, a polynucleotide template is generated through PCR as a linear polynucleotide template. In some embodiments, a linearized polynucleotide is mixed with enzymes suitable for RNA synthesis, RNA capping and/or purification. In some embodiments, the resulting RNA is suitable for producing proteins encoded by the RNA. [0831] A variety of methods are known in the art to introduce an expression vector into host cells. In some embodiments, a vector may be introduced into host cells using transfection. In some embodiments, transfection is completed, for example, using calcium phosphate transfection, lipofection, or polyethylenimine- mediated transfection. In some embodiments, a vector may be introduced into a host cell using transduction. [0832] In some embodiments, transformed host cells are cultured following introduction of a vector into a host cell to allow for expression of said recombinant polynucleotides. In some embodiments, a transformed host cells are cultured for at least 12 hours, 16 hours, 20 hours, 24 hours, 28 hours, 32 hours, 36 hours 40 hours, 44 hours, 48 hours, 52 hours, 56 hours, 60 hours, 64 hours, 68 hours, 72 hours or longer. Transformed host cells are cultured in growth conditions (e.g., temperature, carbon-dioxide levels, growth medium) in accordance with the requirements of a host cell selected. A skilled artisan would recognize culture conditions for host cells selected are well known in the art. X. Dosage Regimens [0833] In some embodiments, the present disclosure provides a method of treating or preventing herpes simplex virus (HSV) infection comprising administering to a subject in need thereof a therapeutically effective amount of a composition disclosed herein, in a treatment cycle comprising one or more doses (e.g., one dose, two doses, three doses, four doses, five doses, six doses, seven doses, eight doses, nine doses, or ten doses) of the composition. In some embodiments, a treatment cycle comprises two or more doses (e.g., two doses, three doses, four doses, five doses, six doses, seven doses, eight doses, nine doses, or ten doses) of the composition. In some embodiments, a treatment cycle comprises two doses. In some embodiments, a first dose is a priming dose of a composition disclosed herein. In some embodiments, a second dose is a booster dose of a composition disclosed herein. [0834] In some embodiments, a subject is administered one or more doses of a composition disclosed herein prior to infection with HSV (e.g., HSV-1, HSV-2, or a combination thereof). In some embodiments, a subject is administered two or more doses of a composition disclosed herein prior to infection with HSV (e.g., HSV-1, HSV-2, or a combination thereof). In some embodiments, a subject is administered three or more doses of a composition disclosed herein prior to infection with HSV (e.g., HSV-1, HSV-2, or a combination thereof). [0835] In some embodiments, a second dose of the therapeutically effective amount of a composition disclosed herein is administered to a subject 1 day to 24 weeks, 3.5 days to 24 weeks, 1 week to 24 weeks, 2 weeks to 24 weeks, 4 weeks to 24 weeks, 6 weeks to 24 weeks, 8 weeks to 24 weeks, 10 weeks to 24 weeks, 12 weeks to 24 weeks, 16 weeks to 24 weeks, 20 weeks to 24 weeks, 1 day to 20 weeks, 3.5 days to 20 weeks, 1 week to 20 weeks, 2 weeks to 20 weeks, 4 weeks to 20 weeks, 6 weeks to 20 weeks, 8 weeks to 20 weeks, 10 weeks to 20 weeks, 12 weeks to 20 weeks, 16 weeks to 20 weeks, 1 day to 16 weeks, 3.5 days to 16 weeks, 1 week to 16 weeks, 2 weeks to 16 weeks, 4 weeks to 16 weeks, 6 weeks to 16 weeks, 8 weeks to 16 weeks, 10 weeks to 16 weeks, 12 weeks to 16 weeks, 1 day to 12 weeks, 3.5 days to 12 weeks, 1 week to 12 weeks, 2 weeks to 12 weeks, 4 weeks to 12 weeks, 6 weeks to 12 weeks, 8 weeks to 12 weeks, 10 weeks to 12 weeks, 1 day to 10 weeks, 3.5 days to 10 weeks, 1 week to 10 weeks, 2 week to 10 weeks, 4 weeks to 10 weeks, 6 weeks to 10 weeks, 8 weeks to 10 weeks, 1 day to 8 weeks, 3.5 days to 8 weeks, 1 week to 8 weeks, 2 weeks to 8 weeks, 4 weeks to 8 weeks, 6 weeks to 8 weeks, 1 day to 6 weeks, 3.5 days to 6 weeks, 1 week to 6 weeks, 2 weeks to 6 weeks, 4 weeks to 6 weeks, 1 day to 4 weeks, 3.5 days to 4 weeks, 1 week to 4 weeks, 2 week to 4 weeks, 1 day to 2 weeks, 3.5 days to 2 weeks, 1 week to 2 weeks, 1 day to 1 week, 3.5 days to 1 week, or 1 day to 3.5 days, after administration of a first dose of the therapeutically effective amount of the composition to the subject. In some embodiments, a second dose of the therapeutically effective amount of a composition disclosed herein is administered 1 week to 14 weeks after administration of the first dose of the therapeutically effective amount of the composition to the subject. In some embodiments, a second dose of the therapeutically effective amount of a composition disclosed herein is administered 4 weeks to 12 weeks after administration of the first dose of the therapeutically effective amount of the composition to the subject. In some embodiments, a second dose of the therapeutically effective amount of a composition disclosed herein is administered 6 weeks to 10 weeks after administration of the first dose of the therapeutically effective amount of the composition to the subject. [0836] In some embodiments, a second dose of the therapeutically effective amount of a composition disclosed herein is administered about 1 week, about 2 weeks, about 4 weeks, about 6 weeks, about 8 weeks, about 10 weeks, about 12 weeks, about 16 weeks, about 20 weeks, or about 24 weeks after administration of the first dose of the therapeutically effective amount of the composition to the subject. In some embodiments, a second dose of the therapeutically effective amount of a composition disclosed herein is administered about 8 weeks after administration of the first dose of the therapeutically effective amount of the composition to the subject. [0837] In some embodiments, a second dose of the therapeutically effective amount of a composition disclosed herein is administered about 1 day, about 7 days, about 14 days, about 28 days, about 35 days, about 40 days, about 45 days, about 50 days, about 51 days, about 52 days, about 53 days, about 54 days, about 55 days, about 56 days, about 57 days, about 58 days, about 59 days, or about 60 days after administration of the first dose of the therapeutically effective amount of the composition to the subject. [0838] In some embodiments, a third dose of the therapeutically effective amount of a composition disclosed herein is administered to a subject 1 day to 24 weeks, 3.5 days to 24 weeks, 1 week to 24 weeks, 2 weeks to 24 weeks, 4 weeks to 24 weeks, 6 weeks to 24 weeks, 8 weeks to 24 weeks, 10 weeks to 24 weeks, 12 weeks to 24 weeks, 16 weeks to 24 weeks, 20 weeks to 24 weeks, 1 day to 20 weeks, 3.5 days to 20 weeks, 1 week to 20 weeks, 2 weeks to 20 weeks, 4 weeks to 20 weeks, 6 weeks to 20 weeks, 8 weeks to 20 weeks, 10 weeks to 20 weeks, 12 weeks to 20 weeks, 16 weeks to 20 weeks, 1 day to 16 weeks, 3.5 days to 16 weeks, 1 week to 16 weeks, 2 weeks to 16 weeks, 4 weeks to 16 weeks, 6 weeks to 16 weeks, 8 weeks to 16 weeks, 10 weeks to 16 weeks, 12 weeks to 16 weeks, 1 day to 12 weeks, 3.5 days to 12 weeks, 1 week to 12 weeks, 2 weeks to 12 weeks, 4 weeks to 12 weeks, 6 weeks to 12 weeks, 8 weeks to 12 weeks, 10 weeks to 12 weeks, 1 day to 10 weeks, 3.5 days to 10 weeks, 1 week to 10 weeks, 2 week to 10 weeks, 4 weeks to 10 weeks, 6 weeks to 10 weeks, 8 weeks to 10 weeks, 1 day to 8 weeks, 3.5 days to 8 weeks, 1 week to 8 weeks, 2 weeks to 8 weeks, 4 weeks to 8 weeks, 6 weeks to 8 weeks, 1 day to 6 weeks, 3.5 days to 6 weeks, 1 week to 6 weeks, 2 weeks to 6 weeks, 4 weeks to 6 weeks, 1 day to 4 weeks, 3.5 days to 4 weeks, 1 week to 4 weeks, 2 week to 4 weeks, 1 day to 2 weeks, 3.5 days to 2 weeks, 1 week to 2 weeks, 1 day to 1 week, 3.5 days to 1 week, or 1 day to 3.5 days, after administration of a first dose of the therapeutically effective amount of the composition to the subject. In some embodiments, a third dose of the therapeutically effective amount of a composition disclosed herein is administered 4 weeks to 24 weeks after administration of the first dose of the therapeutically effective amount of the composition to the subject. In some embodiments, a third dose of the therapeutically effective amount of a composition disclosed herein is administered 12 weeks to 20 weeks after administration of the first dose of the therapeutically effective amount of the composition to the subject. In some embodiments, a third dose of the therapeutically effective amount of a composition disclosed herein is administered 14 weeks to 18 weeks after administration of the first dose of the therapeutically effective amount of the composition to the subject. [0839] In some embodiments, a third dose of the therapeutically effective amount of a composition disclosed herein is administered about 6 weeks, about 8 weeks, about 10 weeks, about 12 weeks, about 13 weeks, about 14 weeks, about 15 weeks, about 16 weeks, about 17 weeks, about 18 weeks, about 19 weeks, about 20 weeks, or about 24 weeks after administration of the first dose of the therapeutically effective amount of the composition to the subject. In some embodiments, a third dose of the therapeutically effective amount of a composition disclosed herein is administered about 8 weeks after administration of the first dose of the therapeutically effective amount of the composition to the subject. [0840] In some embodiments, a third dose of the therapeutically effective amount of a composition disclosed herein is administered about 10 days, about 20 days, about 30 days, about 40 days, about 50 days, about 60 days, about 70 days, about 80 days, about 90 days, about 100 days, about 105 days, about 110 days, about 111 days, about 112 days, about 113 days, about 114 days, about 115 days, about 116 days, or about 117 days after administration of the first dose of the therapeutically effective amount of the composition to the subject. [0841] In some embodiments, each of the one or more doses of a therapeutically effective amount of a composition disclosed herein is administered to a subject intramuscularly, subcutaneously, orally, or intranasally. In some embodiments, each of the one or more doses of a therapeutically effective amount of a composition disclosed herein is administered to a subject intramuscularly. [0842] In some embodiments, each of the one or more doses comprises 0.1 to 500 μg, 0.2 to 500 μg, 0.5 to 500 μg, 0.75 to 500 μg, 1 to 500 μg. 2.5 to 500 μg, 5 to 500 μg, 7.5 to 500 μg, 10 to 500 μg, 12.5 to 500 15 to 500 μg, 17.5 to 500 μg, 20 to 500 μg, 25 to 500 μg, 30 to 500 μg, 35 to 500 μg, 40 to 500 μg, 45 to 500 μg, 50 to 500 μg, 55 to 500 μg, 60 to 500 μg, 65 to 500 μg, 70 to 500 μg, 75 to 500 μg, 80 to 500 μg, 85 to 500 μg, 90 to 500 μg, 95 to 500 μg, 100 to 500 μg, 125 to 500 μg, 150 to 500 μg, 175 to 500 μg, 200 to 500 μg, 225 to 500 μg, 250 to 500 μg, 275 to 500 μg, 300 to 500 μg, 325 to 500 μg, 350 to 500 μg, 375 to 500 μg, 400 to 500 μg, 425 to 500 μg, 450 to 500 μg, 475 to 500 μg, 0.1 to 450 μg, 0.2 to 450 μg, 0.5 to 450 μg, 0.75 to 450 μg, 1 to 450 μg. 2.5 to 450 μg, 5 to 450 μg, 7.5 to 450 μg, 10 to 450 μg, 12.5 to 450 15 to 450 μg, 17.5 to 450 μg, 20 to 450 μg, 25 to 450 μg, 30 to 450 μg, 35 to 450 μg, 40 to 450 μg, 45 to 450 μg, 50 to 450 μg, 55 to 450 μg, 60 to 450 μg, 65 to 450 μg, 70 to 450 μg, 75 to 450 μg, 80 to 450 μg, 85 to 450 μg, 90 to 450 μg, 95 to 450 μg, 100 to 450 μg, 125 to 450 μg, 150 to 450 μg, 175 to 450 μg, 200 to 450 μg, 225 to 450 μg, 250 to 450 μg, 275 to 450 μg, 300 to 450 μg, 325 to 450 μg, 350 to 450 μg, 375 to 450 μg, 400 to 450 μg, 425 to 450 μg, 0.1 to 400 μg, 0.2 to 400 μg, 0.5 to 400 μg, 0.75 to 400 μg, 1 to 400 μg. 2.5 to 400 μg, 5 to 400 μg, 7.5 to 400 μg, 10 to 400 μg, 12.5 to 400 15 to 400 μg, 17.5 to 400 μg, 20 to 400 μg, 25 to 400 μg, 30 to 400 μg, 35 to 400 μg, 40 to 400 μg, 45 to 400 μg, 50 to 400 μg, 55 to 400 μg, 60 to 400 μg, 65 to 400 μg, 70 to 400 μg, 75 to 400 μg, 80 to 400 μg, 85 to 400 μg, 90 to 400 μg, 95 to 400 μg, 100 to 400 μg, 125 to 400 μg, 150 to 400 μg, 175 to 400 μg, 200 to 400 μg, 225 to 400 μg, 250 to 400 μg, 275 to 400 μg, 300 to 400 μg, 325 to 400 μg, 350 to 400 μg, 375 to 400 μg, 0.1 to 350 μg, 0.2 to 350 μg, 0.5 to 350 μg, 0.75 to 350 μg, 1 to 350 μg. 2.5 to 350 μg, 5 to 350 μg, 7.5 to 350 μg, 10 to 350 μg, 12.5 to 350 15 to 350 μg, 17.5 to 350 μg, 20 to 350 μg, 25 to 350 μg, 30 to 350 μg, 35 to 350 μg, 40 to 350 μg, 45 to 350 μg, 50 to 350 μg, 55 to 350 μg, 60 to 350 μg, 65 to 350 μg, 70 to 350 μg, 75 to 350 μg, 80 to 350 μg, 85 to 350 μg, 90 to 350 μg, 95 to 350 μg, 100 to 350 μg, 125 to 350 μg, 150 to 350 μg, 175 to 350 μg, 200 to 350 μg, 225 to 350 μg, 250 to 350 μg, 275 to 350 μg, 300 to 350 μg, 325 to 350 μg, 0.1 to 300 μg, 0.2 to 300 μg, 0.5 to 300 μg, 0.75 to 300 μg, 1 to 300 μg. 2.5 to 300 μg, 5 to 300 μg, 7.5 to 300 μg, 10 to 300 μg, 12.5 to 300 15 to 300 μg, 17.5 to 300 μg, 20 to 300 μg, 25 to 300 μg, 30 to 300 μg, 35 to 300 μg, 40 to 300 μg, 45 to 300 μg, 50 to 300 μg, 55 to 300 μg, 60 to 300 μg, 65 to 300 μg, 70 to 300 μg, 75 to 300 μg, 80 to 300 μg, 85 to 300 μg, 90 to 300 μg, 95 to 300 μg, 100 to 300 μg, 125 to 300 μg, 150 to 300 μg, 175 to 300 μg, 200 to 300 μg, 225 to 300 μg, 250 to 300 μg, 275 to 300 μg, 0.1 to 250 μg, 0.2 to 250 μg, 0.5 to 250 μg, 0.75 to 250 μg, 1 to 250 μg. 2.5 to 250 μg, 5 to 250 μg, 7.5 to 250 μg, 10 to 250 μg, 12.5 to 250 15 to 250 μg, 17.5 to 250 μg, 20 to 250 μg, 25 to 250 μg, 30 to 250 μg, 35 to 250 μg, 40 to 250 μg, 45 to 250 μg, 50 to 250 μg, 55 to 250 μg, 60 to 250 μg, 65 to 250 μg, 70 to 250 μg, 75 to 250 μg, 80 to 250 μg, 85 to 250 μg, 90 to 250 μg, 95 to 250 μg, 100 to 250 μg, 125 to 250 μg, 150 to 250 μg, 175 to 250 μg, 200 to 250 μg, 225 to 250 μg, 0.1 to 200 μg, 0.2 to 200 μg, 0.5 to 200 μg, 0.75 to 200 μg, 1 to 200 μg. 2.5 to 200 μg, 5 to 200 μg, 7.5 to 200 μg, 10 to 200 μg, 12.5 to 200 15 to 200 μg, 17.5 to 200 μg, 20 to 200 μg, 25 to 200 μg, 30 to 200 μg, 35 to 200 μg, 40 to 200 μg, 45 to 200 μg, 50 to 200 μg, 55 to 200 μg, 60 to 200 μg, 65 to 200 μg, 70 to 200 μg, 75 to 200 μg, 80 to 200 μg, 85 to 200 μg, 90 to 200 μg, 95 to 200 μg, 100 to 200 μg, 125 to 200 μg, 150 to 200 μg, 175 to 200 μg, 0.1 to 150 μg, 0.2 to 150 μg, 0.5 to 150 μg, 0.75 to 150 μg, 1 to 150 μg. 2.5 to 150 μg, 5 to 150 μg, 7.5 to 150 μg, 10 to 150 μg, 12.5 to 150 15 to 150 μg, 17.5 to 150 μg, 20 to 150 μg, 25 to 150 μg, 30 to 150 μg, 35 to 150 μg, 40 to 150 μg, 45 to 150 μg, 50 to 150 μg, 55 to 150 μg, 60 to 150 μg, 65 to 150 μg, 70 to 150 μg, 75 to 150 μg, 80 to 150 μg, 85 to 150 μg, 90 to 150 μg, 95 to 150 μg, 100 to 150 μg, 125 to 150 μg, 0.1 to 100 μg, 0.2 to 100 μg, 0.5 to 100 μg, 0.75 to 100 μg, 1 to 100 μg. 2.5 to 100 μg, 5 to 100 μg, 7.5 to 100 μg, 10 to 100 μg, 12.5 to 100 15 to 100 μg, 17.5 to 100 μg, 20 to 100 μg, 25 to 100 μg, 30 to 100 μg, 35 to 100 μg, 40 to 100 μg, 45 to 100 μg, 50 to 100 μg, 55 to 100 μg, 60 to 100 μg, 65 to 100 μg, 70 to 100 μg, 75 to 100 μg, 80 to 100 μg, 85 to 100 μg, 90 to 100 μg, 95 to 100 μg, 0.1 to 90 μg, 0.2 to 90 μg, 0.5 to 90 μg, 0.75 to 90 μg, 1 to 90 μg. 2.5 to 90 μg, 5 to 90 μg, 7.5 to 90 μg, 10 to 90 μg, 12.5 to 90 15 to 90 μg, 17.5 to 90 μg, 20 to 90 μg, 25 to 90 μg, 30 to 90 μg, 35 to 90 μg, 40 to 90 μg, 45 to 90 μg, 50 to 90 μg, 55 to 90 μg, 60 to 90 μg, 65 to 90 μg, 70 to 90 μg, 75 to 90 μg, 80 to 90 μg, 85 to 90 μg, 0.1 to 80 μg, 0.2 to 80 μg, 0.5 to 80 μg, 0.75 to 80 μg, 1 to 80 μg. 2.5 to 80 μg, 5 to 80 μg, 7.5 to 80 μg, 10 to 80 μg, 12.5 to 80 μg, 15 to 80 μg, 17.5 to 80 μg, 20 to 80 μg, 25 to 80 μg, 30 to 80 μg, 35 to 80 μg, 40 to 80 μg, 45 to 80 μg, 50 to 80 μg, 55 to 80 μg, 60 to 80 μg, 65 to 80 μg, 70 to 80 μg, 75 to 80 μg, 0.1 to 70 μg, 0.2 to 70 μg, 0.5 to 70 μg, 0.75 to 70 μg, 1 to 70 μg. 2.5 to 70 μg, 5 to 70 μg, 7.5 to 70 μg, 10 to 70 μg, 12.5 to 70 μg, 15 to 70 μg, 17.5 to 70 μg, 20 to 70 μg, 25 to 70 μg, 30 to 70 μg, 35 to 70 μg, 40 to 70 μg, 45 to 70 μg, 50 to 70 μg, 55 to 70 μg, 60 to 70 μg, 65 to 70 μg, 0.1 to 60 μg, 0.2 to 60 μg, 0.5 to 60 μg, 0.75 to 60 μg, 1 to 60 μg. 2.5 to 60 μg, 5 to 60 μg, 7.5 to 60 μg, 10 to 60 μg, 12.5 to 60 μg, 15 to 60 μg, 17.5 to 60 μg, 20 to 60 μg, 25 to 60 μg, 30 to 60 μg, 35 to 60 μg, 40 to 60 μg, 45 to 60 μg, 50 to 60 μg, 55 to 60 μg, 0.1 to 50 μg, 0.2 to 50 μg, 0.5 to 50 μg, 0.75 to 50 μg, 1 to 50 μg. 2.5 to 50 μg, 5 to 50 μg, 7.5 to 50 μg, 10 to 50 μg, 12.5 to 50 μg, 15 to 50 μg, 17.5 to 50 μg, 20 to 50 μg, 25 to 50 μg, 30 to 50 μg, 35 to 50 μg, 40 to 50 μg, 45 to 50 μg, 0.1 to 40 μg, 0.2 to 40 μg, 0.5 to 40 μg, 0.75 to 40 μg, 1 to 40 μg. 2.5 to 40 μg, 5 to 40 μg, 7.5 to 40 μg, 10 to 40 μg, 12.5 to 40 μg, 15 to 40 μg, 17.5 to 40 μg, 20 to 40 μg, 25 to 40 μg, 30 to 40 μg, 35 to 40 μg, 0.1 to 30 μg, 0.2 to 30 μg, 0.5 to 30 μg, 0.75 to 30 μg, 1 to 30 μg. 2.5 to 30 μg, 5 to 30 μg, 7.5 to 30 μg, 10 to 30 μg, 12.5 to 30 15 to 30 μg, 17.5 to 30 μg, 20 to 30 μg, 25 to 30 μg, 0.1 to 20 μg, 0.2 to 20 μg, 0.5 to 20 μg, 0.75 to 20 μg, 1 to 20 μg. 2.5 to 20 μg, 5 to 20 μg, 7.5 to 20 μg, 10 to 20 μg, 12.5 to 20 15 to 20 μg, 17.5 to 20 μg, 0.1 to 10 μg, 0.2 to 10 μg, 0.5 to 10 μg, 0.75 to 10 μg, 1 to 10 μg. 2.5 to 10 μg, 5 to 10 μg, 7.5 to 10 μg, 0.1 to 1 μg, 0.2 to 1 μg, 0.5 to 1 μg, or 0.75 to 1 μg of one or more polyribonucleotides encoding one or more HSV glycoprotein antigens or antigenic fragments thereof. [0843] In some embodiments, each of the one or more doses comprises 1 μg to 250 μg of one or more polyribonucleotides encoding one or more HSV glycoproteins or antigenic fragments thereof. In some embodiments, each of the one or more doses comprises 2 μg to 200 μg of one or more polyribonucleotides encoding one or more HSV glycoproteins or antigenic fragments thereof. In some embodiments, each of the one or more doses comprises 3 μg to 100 μg of one or more polyribonucleotides encoding one or more HSV glycoproteins or antigenic fragments thereof. In some embodiments, each of the one or more doses comprises 3 μg to 60 μg of one or more polyribonucleotides encoding one or more HSV glycoproteins or antigenic fragments thereof. In some embodiments, each of the one or more doses comprises 3 μg of one or more polyribonucleotides encoding one or more HSV glycoproteins or antigenic fragments thereof. In some embodiments, each of the one or more doses comprises 10 μg of one or more polyribonucleotides encoding one or more HSV glycoproteins or antigenic fragments thereof. In some embodiments, each of the one or more doses comprises 30 μg of one or more polyribonucleotides encoding one or more HSV glycoproteins or antigenic fragments thereof. In some embodiments, each of the one or more doses comprises 60 μg of one or more polyribonucleotides encoding one or more HSV glycoproteins or antigenic fragments thereof. [0844] In some embodiments, each of the one or more doses comprises 0.5 μg, 0.75 μg, 1.0 μg, 1.25 μg, 1.33 μg, 1.5μg, 1.75 μg, 2.0 μg, 2.25 μg, 2.5 μg, 5 μg, 7.5 μg, 10 μg, 15 μg, 20 μg, or 25 μg of one or more polyribonucleotides encoding one or more HSV glycoproteins or antigenic fragments thereof. For example, in some embodiments, each of the one or more doses comprises 1 μg of each of three polyribonucleotides, wherein each polyribonucleotide encodes a different HSV glycoprotein or antigenic fragment thereof. In another example, in some embodiments, each of the one or more doses comprises 3.33 μg of each of three polyribonucleotides, wherein each polyribonucleotide encodes a different HSV glycoprotein or antigenic fragment thereof. In yet another example, in some embodiments, each of the one or more doses comprises 10 μg of each of three polyribonucleotides, wherein each polyribonucleotide encodes a different HSV glycoprotein or antigenic fragment thereof. In some embodiments, each of the one or more doses comprises 20 μg of each of three polyribonucleotides, wherein each polyribonucleotide encodes a different HSV glycoprotein or antigenic fragment thereof. XI. Concomitant Therapies A. Combinatorial Use with Antipyretic and/or Analgesic Medications [0845] In some embodiments, the present disclosure provides a method of treating or preventing HSV infection comprising administering to a subject in need thereof one or more doses of a therapeutically effective amount of a composition as disclosed herein as part of a combination therapy. [0846] In some embodiments, the present disclosure provides a method of treating or preventing HSV infection comprising administering to a subject in need thereof one or more doses of a therapeutically effective amount of a composition as disclosed herein concomitantly with administration of an antipyretic medication. In some embodiments, an antipyretic medication is administered within 60 minutes, within 30 minutes, or within 15 minutes after administration of one or more doses of a therapeutically effective amount of a composition as disclosed herein. In some embodiments, an antipyretic medication is administered concurrently with administration of one or more doses of a therapeutically effective amount of a composition as disclosed herein. In some embodiments, an antipyretic medication is administered within 60 minutes, within 30 minutes, or within 15 minutes before administration of one of more doses of a therapeutically effective amount of a composition as disclosed herein. [0847] In some embodiments, the antipyretic comprises acetaminophen, a non-steroidal anti-inflammatory drug (NSAID), salicylamide, salicyl salicylate, methyl salicylate, magnesium salicylate, faislamine, ethenzamide, diflunisal, choline magnesium salicylate, benorylate/benorilatem and amoxiprin, acetylsalicylate, ceclofenac, acemetacin, alclofenac, bromfenac, diclofenac, etodolac, indomethacin, nabumetone, oxametacin, proglumetacin, sulindac, tolmetin, iminoprofen, benoxaprofen, carprofen, dexibuprofen, dexketoprofen, fenbufen, fenoprofen, flunoxaprofen, flurbiprofen, ibuprofen, ibuproxam, indoprofen, ketoprofen, ketorolac, loxoprofen, naproxen, oxaprozin, pirprofen, suprofen, tiaprofenic acid, mefenamic acid, flufenamic acid, meclofenamic acid, tolfenamic acid, droxicam, lornoxicam, meloxicam, piroxicam, tenoxicam, dipyrone, azapropazone, clofezone, kebuzone, metamizole, mofebutazone, oxyphenbutazone, phenazone, phenylbutazone, sulfinpyrazone, decoxib, rofecoxib, parecoxib, or etoricoxib. In certain embodiments, the antipyretic is an NSAID. In certain embodiments, the antipyretic is acetaminophen. [0848] In some embodiments, the present disclosure provides a method of treating or preventing HSV (HSV-1, HSV-2, or a combination thereof) infection comprising administering to a subject in need thereof one or more doses of a therapeutically effective amount of a composition as disclosed herein concomitantly with administration of an analgesic medication. In some embodiments, an analgesic medication is administered within 60 minutes, within 30 minutes, or within 15 minutes after administration of one or more doses of a therapeutically effective amount of a composition as disclosed herein. In some embodiments, an analgesic medication is administered concurrently with administration of one or more doses of a therapeutically effective amount of a composition as disclosed herein. In some embodiments, an analgesic medication is administered within 60 minutes, within 30 minutes, or within 15 minutes before administration of one of more doses of a therapeutically effective amount of a composition as disclosed herein. [0849] In some embodiments, an analgesic comprises acetaminophen, salicylamide, salicyl salicylate, methyl salicylate, magnesium salicylate, faislamine, ethenzamide, diflunisal, choline magnesium salicylate, benorylate/benorilatem and amoxiprin, acetylsalicylate, ceclofenac, acemetacin, alclofenac, bromfenac, diclofenac, etodolac, indomethacin, nabumetone, oxametacin, proglumetacin, sulindac, tolmetin, iminoprofen, benoxaprofen, carprofen, dexibuprofen, dexketoprofen, fenbufen, fenoprofen, flunoxaprofen, flurbiprofen, ibuprofen, ibuproxam, indoprofen, ketoprofen, ketorolac, loxoprofen, naproxen, oxaprozin, pirprofen, suprofen, tiaprofenic acid, mefenamic acid, flufenamic acid, meclofenamic acid, tolfenamic acid, droxicam, lornoxicam, meloxicam, piroxicam, tenoxicam, dipyrone, azapropazone, clofezone, kebuzone, metamizole, mofebutazone, oxyphenbutazone, phenazone, phenylbutazone, sulfinpyrazone, decoxib, rofecoxib, parecoxib, etoricoxib, codeine, dihydrocodeine, morphine or a morphine derivative or pharmaceutically acceptable salt thereof, diacetylmorphine, hydrocodone, hydromorphone, levorphanol, oxymorphone, alfentanil, buprenorphine, butorphanol, fentanyl, sufentanil, meperidine, methadone, nalbuphine, propoxyphene, pentazocine, or pharmaceutically acceptable salts thereof. In some embodiments, the analgesic is acetaminophen. [0850] In some embodiments, the present disclosure provides a method of treating or preventing HSV (HSV-1, HSV-2, or a combination thereof) infection comprising administering to a subject in need thereof one or more doses of a therapeutically effective amount of a composition as disclosed herein concomitantly with administration of an antipyretic medication and an analgesic medication. In some embodiments, an antipyretic medication and analgesic medication is administered within 60 minutes, within 30 minutes, or within 15 minutes after administration of one or more doses of a therapeutically effective amount of a composition as disclosed herein. In some embodiments, an antipyretic medication and analgesic medication is administered concurrently with administration of one or more doses of a therapeutically effective amount of a composition as disclosed herein. In some embodiments, an antipyretic medication and analgesic medication is administered within 60 minutes, within 30 minutes, or within 15 minutes before administration of one of more doses of a therapeutically effective amount of a composition as disclosed herein. [0851] In some embodiments, an antipyretic medication and/or analgesic medication is acetaminophen. In some embodiments, acetaminophen is administered to the subject at a dose of 0.1 g/day to 20 g/day, 0.25 g/day to 20 g/day, 0.5 g/day to 20 g/day, 0.75 g/day to 20 g/day, 1.0 g/day to 20 g/day, 1.25 g/day to 20 g/day, 1.5 g/day to 20 g/day, 1.75 g/day to 20 g/day, 2.0 g/day to 20 g/day, 2.25 g/day to 201 g/day, 2.5 g/day to 20 g/day, 2.75 g/day to 20 g/day, 3.0 g/day to 20 g/day, 3.25 g/day to 20 g/day, 3.5 g/day to 20 g/day, 3.75 g/day to 20 g/day, 4.0 g/day to 20 g/day, 4.25 g/day to 20 g/day, 4.5 g/day to 20 g/day, 4.75 g/day to 20 g/day, 5.0 g/day to 20 g/day, 5.25 g/day to 20 g/day, 5.5 g/day to 20 g/day, 5.75 g/day to 20 g/day, 6.0 g/day to 20 g/day, 6.25 g/day to 20 g/day, 6.5 g/day to 20 g/day, 6.75 g/day to 20 g/day, 7.0 g/day to 20 g/day, 7.25 g/day to 20 g/day, 7.5 g/day to 20 g/day, 7.75 g/day to 20 g/day, 8.0 g/day to 20 g/day, 8.25 g/day to 20 g/day, 8.5 g/day to 20 g/day, 8.75 g/day to 20 g/day, 9.0 g/day to 20 g/day, 9.25 g/day to 20 g/day, 9.5 g/day to 20 g/day, 9.75 g/day to 20 g/day, 10 g/day to 20 g/day, 12.5 g/day to 20 g/day, 15 g/day to 20 g/day, 17.5 g/day to 20 g/day, 0.1 g/day to 15 g/day, 0.25 g/day to 15 g/day, 0.5 g/day to 15 g/day, 0.75 g/day to 15 g/day, 1.0 g/day to 15 g/day, 1.25 g/day to 15 g/day, 1.5 g/day to 15 g/day, 1.75 g/day to 15 g/day, 2.0 g/day to 15 g/day, 2.25 g/day to 151 g/day, 2.5 g/day to 15 g/day, 2.75 g/day to 15 g/day, 3.0 g/day to 15 g/day, 3.25 g/day to 15 g/day, 3.5 g/day to 15 g/day, 3.75 g/day to 15 g/day, 4.0 g/day to 15 g/day, 4.25 g/day to 15 g/day, 4.5 g/day to 15 g/day, 4.75 g/day to 15 g/day, 5.0 g/day to 15 g/day, 5.25 g/day to 15 g/day, 5.5 g/day to 15 g/day, 5.75 g/day to 15 g/day, 6.0 g/day to 15 g/day, 6.25 g/day to 15 g/day, 6.5 g/day to 15 g/day, 6.75 g/day to 15 g/day, 7.0 g/day to 15 g/day, 7.25 g/day to 15 g/day, 7.5 g/day to 15 g/day, 7.75 g/day to 15 g/day, 8.0 g/day to 15 g/day, 8.25 g/day to 15 g/day, 8.5 g/day to 15 g/day, 8.75 g/day to 15 g/day, 9.0 g/day to 15 g/day, 9.25 g/day to 15 g/day, 9.5 g/day to 15 g/day, 9.75 g/day to 15 g/day, 10 g/day to 15 g/day, 12.5 g/day to 15 g/day, 0.1 g/day to 10 g/day, 0.25 g/day to 10 g/day, 0.5 g/day to 10 g/day, 0.75 g/day to 10 g/day, 1.0 g/day to 10 g/day, 1.25 g/day to 10 g/day, 1.5 g/day to 10 g/day, 1.75 g/day to 10 g/day, 2.0 g/day to 10 g/day, 2.25 g/day to 101 g/day, 2.5 g/day to 10 g/day, 2.75 g/day to 10 g/day, 3.0 g/day to 10 g/day, 3.25 g/day to 10 g/day, 3.5 g/day to 10 g/day, 3.75 g/day to 10 g/day, 4.0 g/day to 10 g/day, 4.25 g/day to 10 g/day, 4.5 g/day to 10 g/day, 4.75 g/day to 10 g/day, 5.0 g/day to 10 g/day, 5.25 g/day to 10 g/day, 5.5 g/day to 10 g/day, 5.75 g/day to 10 g/day, 6.0 g/day to 10 g/day, 6.25 g/day to 10 g/day, 6.5 g/day to 10 g/day, 6.75 g/day to 10 g/day, 7.0 g/day to 10 g/day, 7.25 g/day to 10 g/day, 7.5 g/day to 10 g/day, 7.75 g/day to 10 g/day, 8.0 g/day to 10 g/day, 8.25 g/day to 10 g/day, 8.5 g/day to 10 g/day, 8.75 g/day to 10 g/day, 9.0 g/day to 10 g/day, 9.25 g/day to 10 g/day, 9.5 g/day to 10 g/day, 9.75 g/day to 10 g/day, 0.1 g/day to 7.5 g/day, 0.25 g/day to 7.5 g/day, 0.5 g/day to 7.5 g/day, 0.75 g/day to 7.5 g/day, 1.0 g/day to 7.5 g/day, 1.25 g/day to 7.5 g/day, 1.5 g/day to 7.5 g/day, 1.75 g/day to 7.5 g/day, 2.0 g/day to 7.5 g/day, 2.25 g/day to 7.51 g/day, 2.5 g/day to 7.5 g/day, 2.75 g/day to 7.5 g/day, 3.0 g/day to 7.5 g/day, 3.25 g/day to 7.5 g/day, 3.5 g/day to 7.5 g/day, 3.75 g/day to 7.5 g/day, 4.0 g/day to 7.5 g/day, 4.25 g/day to 7.5 g/day, 4.5 g/day to 7.5 g/day, 4.75 g/day to 7.5 g/day, 5.0 g/day to 7.5 g/day, 5.25 g/day to 7.5 g/day, 5.5 g/day to 7.5 g/day, 5.75 g/day to 7.5 g/day, 6.0 g/day to 7.5 g/day, 6.25 g/day to 7.5 g/day, 6.5 g/day to 7.5 g/day, 6.75 g/day to 7.5 g/day, 7.0 g/day to 7.5 g/day, 7.25 g/day to 7.5 g/day, 0.1 g/day to 5.0 g/day, 0.25 g/day to 5.0 g/day, 0.5 g/day to 5.0 g/day, 0.75 g/day to 5.0 g/day, 1.0 g/day to 5.0 g/day, 1.25 g/day to 5.0 g/day, 1.5 g/day to 5.0 g/day, 1.75 g/day to 5.0 g/day, 2.0 g/day to 5.0 g/day, 2.25 g/day to 5.0 g/day, 2.5 g/day to 5.0 g/day, 2.75 g/day to 5.0 g/day, 3.0 g/day to 5.0 g/day, 3.25 g/day to 5.0 g/day, 3.5 g/day to 5.0 g/day, 3.75 g/day to 5.0 g/day, 4.0 g/day to 5.0 g/day, 4.25 g/day to 5.0 g/day, 4.5 g/day to 5.0 g/day, 4.75 g/day to 5.0 g/day, 0.1 g/day to 2.5 g/day, 0.25 g/day to 2.5 g/day, 0.5 g/day to 2.5 g/day, 0.75 g/day to 2.5 g/day, 1.0 g/day to 2.5 g/day, 1.25 g/day to 2.5 g/day, 1.5 g/day to 2.5 g/day, 1.75 g/day to 2.5 g/day, 2.0 g/day to 2.5 g/day, or 2.25 g/day to 2.5 g/day. In some embodiments, acetaminophen is administered to the subject at a dose of 0.5 g/day to 10 g/day. In some embodiments, acetaminophen is administered to the subject at a dose of 1 g/day to 5 g/day. [0852] In some embodiments, acetaminophen is administered to the subject at a dose of about 0.5 g/day, about 1 g/day, about, about 1.5 g/day, about 2 g/day, about 2.5 g/day, about 3 g/day, about 3.5 g/day, about 4 g/day, about 4.5 g/day, or about 5 g/day. In some embodiments, acetaminophen is administered to the subject at a dose of about 4 g/day. B. Combinatorial Use with Vaccines [0853] In some embodiments, the present disclosure provides a method of treating or preventing HSV (HSV-1, HSV-2, or a combination thereof) infection. [0854] In some embodiments, the present disclosure provides a method of treating or preventing HSV (HSV-1, HSV-2, or a combination thereof) infection comprising administering to a subject in need thereof one or more doses of a therapeutically effective amount of a composition as disclosed herein, wherein the subject is further administered a medically indicated vaccine at least 1 day, at least 2 days, at least 3 days, at least 4 days, at least 5 days, at least 6 days, at least 7 days, at least 8 days, at least 9 days, at least 10 days, at least 12 days, at least 14 days, at least 16 days, at least 17 days, at least 18 days, at least 19 days, or at least 20 days before administration of the therapeutically acceptable amount of the composition. In some embodiments, the subject is further administered a medically indicated vaccine at least 14 days before the administration of the therapeutically acceptable amount of the composition. [0855] In some embodiments, the present disclosure provides a method of treating or preventing HSV (HSV-1, HSV-2, or a combination thereof) infection comprising administering to a subject in need thereof one or more doses of a therapeutically effective amount of a composition as disclosed herein, wherein the subject is further administered a medically indicated vaccine at least 1 day, at least 2 days, at least 3 days, at least 4 days, at least 5 days, at least 6 days, at least 7 days, at least 8 days, at least 9 days, at least 10 days, at least 12 days, at least 14 days, at least 16 days, at least 17 days, at least 18 days, at least 19 days, or at least 20 days after administration of the therapeutically acceptable amount of the composition. In some embodiments, the subject is further administered a medically indicated vaccine at least 14 days after the administration of the therapeutically acceptable amount of the composition. [0856] In some embodiments, a medically indicated vaccine includes, but is not limited to, a rabies vaccine, a tetanus vaccine, a hepatitis A vaccine, a hepatitis B vaccine, a measles mumps rubella (MMR) vaccine, a polio vaccine, a diphtheria vaccine, a varicella vaccine, a pertussis vaccine, a shingles vaccine, a pneumococcal vaccine, a human papillomavirus vaccine (HPV), a meningococcal vaccine, or a rotavirus vaccine. In some embodiments, a medically indicated vaccine is a rabies vaccine or a tetanus vaccine. XII. Monitoring Efficacy [0857] In some embodiments, the present disclosure provides a method of treating or preventing HSV (HSV-1, HSV-2, or a combination thereof) infection comprising administering to a subject in need thereof one or more doses of a therapeutically effective amount of a composition as disclosed herein, wherein the method further comprises collecting one or more samples from the subject after administration of the one or more doses of a therapeutically effective amount of the composition. In some embodiments, at least one, at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, at least ten, at least eleven, at least twelve, at least thirteen, at least fourteen, or at least fifteen samples are collected from the subject after administration of the one or more doses of a therapeutically effective amount of the composition. [0858] In some embodiments, the one or more samples collected from the subject is a blood volume draw. [0859] In some embodiments, a sample is collected from the subject about 120 minutes, about 90 minutes, about 60 minutes, about 45 minutes, about 30 minutes, about 20 minutes, about 15 minutes, about 10 minutes, about 7.5 minutes, about 5 minutes, about 4 minutes, about 3 minutes, about 2 minutes or about 1 minute before administration of a first dose of one or more doses of a therapeutically effective amount of the composition. In some embodiments, a sample is collected from the subject about 1 minute, about 2 minutes, about 3 minutes, about 4 minutes, about 5 minutes, about 7.5 minutes, about 10 minutes, about 15 minutes, about 20 minutes, about 30 minutes, about 45 minutes, about 60 minutes, about 90 minutes, or about 120 minutes after administration of a first dose of one or more doses of a therapeutically effective amount of the composition. [0860] In some embodiments, a sample is collected about 1 week after administration of a first dose of one or more doses of a therapeutically effective amount of the composition. In some embodiments, a sample is collected about 2 weeks after administration of a first dose of one or more doses of a therapeutically effective amount of the composition. In some embodiments, a sample is collected about 4 weeks after administration of a first dose of one or more doses of a therapeutically effective amount of the composition. In some embodiments, a sample is collected about 5 weeks after administration of a first dose of one or more doses of a therapeutically effective amount of the composition. In some embodiments, a sample is collected about 6 weeks after administration of a first dose of one or more doses of a therapeutically effective amount of the composition. In some embodiments, a sample is collected about 8 weeks after administration of a first dose of one or more doses of a therapeutically effective amount of the composition. In some embodiments, a sample is collected about 16 weeks after administration of a first dose of one or more doses of a therapeutically effective amount of the composition. In some embodiments, a sample is collected about 7 months after administration of a first dose of one or more doses of a therapeutically effective amount of the composition. In some embodiments, a sample is collected about 13 months after administration of a first dose of one or more doses of a therapeutically effective amount of the composition. [0861] In some embodiments, a sample is collected from the subject about 120 minutes, about 90 minutes, about 60 minutes, about 45 minutes, about 30 minutes, about 20 minutes, about 15 minutes, about 10 minutes, about 7.5 minutes, about 5 minutes, about 4 minutes, about 3 minutes, about 2 minutes or about 1 minute before administration of a second dose of two or more doses of a therapeutically effective amount of the composition. In some embodiments, a sample is collected from the subject about 1 minute, about 2 minutes, about 3 minutes, about 4 minutes, about 5 minutes, about 7.5 minutes, about 10 minutes, about 15 minutes, about 20 minutes, about 30 minutes, about 45 minutes, about 60 minutes, about 90 minutes, or about 120 minutes after administration of a second dose of two or more doses of a therapeutically effective amount of the composition. [0862] In some embodiments, a sample is collected about 1 week after administration of a second dose of two or more doses of a therapeutically effective amount of the composition. In some embodiments, a sample is collected about 2 weeks after administration of a second dose of two or more doses of a therapeutically effective amount of the composition. In some embodiments, a sample is collected about 1 month after administration of a second dose of two or more doses of a therapeutically effective amount of the composition. In some embodiments, a sample is collected about 3 months after administration of a second dose of two or more doses of a therapeutically effective amount of the composition. In some embodiments, a sample is collected about 6 months after administration of a second dose of two or more doses of a therapeutically effective amount of the composition. In some embodiments, a sample is collected about 12 months after administration of a second dose of two or more doses of a therapeutically effective amount of the composition. [0863] In some embodiments, methods of the present disclosure further comprise a step of measuring levels of HSV (HSV-1 and/or HSV-2) virus-specific neutralizing antibodies in one or more samples collected from the subject. In some embodiments, levels of HSV (HSV-1 and/or HSV-2) virus-specific neutralizing antibodies are measured using any one of a number of assays known to persons of ordinary skill in the art. For example, in some embodiments, levels of HSV (HSV-1 and/or HSV-2) virus-specific neutralizing antibodies are measured using a plaque reduction neutralization test (PRNT). In some embodiments, levels of HSV (HSV-1 and/or HSV-2) virus-specific neutralizing antibodies are measured using a pseudo-viral neutralization test. [0864] In some embodiments, methods of the present disclosure further comprise a step of measuring levels of neutralizing antibodies in one or more samples collected from a subject, where the neutralizing antibodies are specific for one or more HSV (HSV-1 and/or HSV-2) antigens or antigenic fragments thereof encoded by one or more polyribonucleotides in a composition disclosed herein. In some embodiments, levels of neutralizing antibodies are measured using any one of a number of assays known to persons of ordinary skill in the art. For example, in some embodiments, levels of neutralizing antibodies are measured using an enzyme-linked immunosorbent assay (ELISA). EXAMPLES [0865] The disclosure is further illustrated by the following examples. The examples are provided for illustrative purposes only. They are not to be construed as limiting the scope or content of the disclosure in any way. Example 1: Exemplary LNP Formulations [0866] The present Example describes certain preferred exemplary LNP formulations useful for
Figure imgf000401_0001
compositions (e.g., immunogenic compositions, e.g., vaccines) as described herein. [0867] In some embodiments, exemplary LNP formulations that are useful for compositions (e.g., immunogenic compositions, e.g., vaccines) as described herein can comprise at least one ionizable aminolipid. In some embodiments, exemplary LNP formulations that are useful for compositions (e.g., immunogenic compositions, e.g., vaccines) as described herein can further comprise a helper lipid, which in some embodiments may be or comprise a neutral helper lipid. In some embodiments, exemplary LNP formulations that are useful for compositions (e.g., immunogenic compositions, e.g., vaccines) as described herein can further comprise a polymer-conjugated lipid, for example in some embodiments PEG-conjugated lipids. In some embodiments, exemplary LNP formulations that are useful for compositions (e.g., immunogenic compositions, e.g., vaccines) as described herein can comprise at least one ionizable aminolipid, at least one helper lipid (e.g., a neutral helper lipid, which in some embodiments may comprise a phospholipid, a steroid, or combinations thereof), and at least one polymer-conjugated lipid (e.g., PEG-conjugated lipid). In some embodiments, an exemplary LNP formulation may comprise an ionizable aminolipid, a phospholipid, a steroid, and a PEG-conjugated lipid. [0868] In some embodiments, an ionizable aminolipid may be present in an exemplary LNP formulation within a range of 45 to 55 mol percent, 40 to 50 mol percent, 41 to 49 mol percent, 41 to 48 mol percent, 42 to 48 mol percent, 43 to 48 mol percent, 44 to 48 mol percent of total lipids. In some embodiments, an ionizable aminolipid is or comprises ((4-hydroxybutyl)azanediyl)bis(hexane-6,1-diyl)bis(2-hexyldecanoate) (also known as 6- [N-6-(2-hexyldecanoyloxy)hexyl-N-(4-hydroxybutyl)amino]hexyl 2-hexyldecanoate). In some embodiments, an ionizable aminolipid is or comprises SM-102 (heptadecan-9-yl 8 ((2 hydroxyethyl)(6 oxo 6- (undecyloxy)hexyl)amino)octanoate) or an aminolipid as described in Sabnis et al. “ A Novel Amino Lipid Series for mRNA Delivery: Improved Endosomal Escape and Sustained Pharmacology and Safety in Non-human Primates” Mol. Ther. (2018) 26:1509-1519. In some embodiments, an ionizable aminolipid is or comprises an ionizable aminolipid as disclosed in US2020/0163878 or WO2018/078053, the entire contents of each of which are incorporated herein by reference for the purposes described herein. [0869] In some embodiments, a phospholipid may be present in an exemplary LNP formulation within a range of 5 to 15 mol percent, 7 to 13 mol percent, or 9 to 11 mol percent of total lipids. In some embodiments, a phospholipid is or comprises 1,2-Distearoyl-sn-glycero- 3-phosphocholine (DSPC). [0870] In some embodiments, a sterol may be present in an exemplary LNP formulation within a range of 30 to 50 mol percent, 35 to 45 mol percent or 38 to 43 mol percent of total lipids. In some embodiments, a sterol is or comprises cholesterol. [0871] In some embodiments, a polymer conjugated lipid (e.g., PEG-conjugated lipid) may be present in an exemplary LNP formulation within a range of 1 to 10 mol percent, 1 to 5 mol percent, or 1 to 2.5 mol percent of total lipids. In some embodiments, a PEG-conjugated lipid is or comprises 2-[(polyethylene glycol)-2000]-N,N- ditetradecylacetamide (also known as 2-[2-(ω-methoxy (polyethyleneglycol2000) ethoxy]-N,N- ditetradecylacetamide). In some embodiments, a phospholipid is or comprises PEG2000-DMG (1- monomethoxypolyethyleneglycol-2,3- dimyristylglycerol with polyethylene glycol of average molecular weight 2000). In some embodiments, a PEG-conjugated lipid is or comprises a PEG-lipid as disclosed in US2020/0163878 or WO2018/078053, the entire contents of each of which are incorporated herein by reference for the purposes described herein. [0872] In some embodiments, an exemplary LNP formulation comprises (i) an ionizable aminolipid within a range of 45 to 55 mol percent of total lipids; (ii) a phospholipid within a range of 8 to 12 mol percent of total lipids; (iii) a steroid within a range of 35 to 45 mol percent of total lipids; and (iv) a polymer conjugated (e.g., PEG- conjugated polymer) within a range of 1 to 2 mol percent of total lipids; and RNA molecules as described herein that are encapsulated within or associated with the lipid nanoparticles. [0873] In some embodiments, an exemplary LNP formulation comprises (i) ionizable amino lipid within a range of 45 to 55 mol percent of total lipids; (ii) DSPC within a range of 5 to 15 mol percent of total lipids; (iii) cholesterol within a range of 35 to 45 mol percent of total lipids; and (iv) a PEG-conjugated lipid within a range of 1 to 2 mol percent of total lipids; and RNA molecules as described herein that are encapsulated within or associated with the lipid nanoparticles. [0874] In some embodiments, an exemplary LNP formulation comprises (i) an ionizable aminolipid within a range of 40 to 50 mol percent of total lipids; (ii) a phospholipid within a range of 5 to 15 mol percent of total lipids; (iii) a steroid within a range of 35 to 45 mol percent of total lipids; and (iv) a polymer conjugated (e.g., PEG- conjugated polymer) within a range of 1 to 10 mol percent of total lipids; and RNA molecules as described herein that are encapsulated within or associated with the lipid nanoparticles. In some such embodiments, an ionizable aminolipid is or comprises ((4-hydroxybutyl)azanediyl)bis(hexane-6,1-diyl)bis(2-hexyldecanoate) (also known as 6- [N-6-(2-hexyldecanoyloxy)hexyl-N-(4-hydroxybutyl)amino]hexyl 2-hexyldecanoate). In some such embodiments, a phospholipid is or comprises 1,2-Distearoyl-sn-glycero-3-phosphocholine (DSPC). In some such embodiments, a steroid is or comprises cholesterol. In some such embodiments, a polymer conjugated polymer is or comprises 2- [(polyethylene glycol)-2000]-N,N-ditetradecylacetamide (also known as 2-[2-(ω-methoxy (polyethyleneglycol2000) ethoxy]-N,N-ditetradecylacetamide). [0875] In some embodiments, an exemplary LNP formulation comprises the following lipids included in Table 23 below and RNA molecules as described herein. Table 23: Exemplary LNP Formulation Lipids Proportion (mol%) ((4-h drox b t l)azanedi l)bis(hexane-61-di l)bis(2-hex ldecanoate) 47-49 (e 475) [0876]
Figure imgf000403_0001
n some em o ments, an exempary ormuaton comprses an onza e amno pd, DSPC, cholesterol, and PEG-conjugated lipid at a molar ratio of approximately 50:10:38.5:1.5 or 47.5:10:40.8:1.7. In some embodiments, an ionizable amino lipid is or comprises ((4-hydroxybutyl)azanediyl)bis(hexane-6,1-diyl)bis(2- hexyldecanoate) (also known as 6-[N-6-(2-hexyldecanoyloxy)hexyl-N-(4-hydroxybutyl)amino]hexyl 2- hexyldecanoate). [0877] In some embodiments, an exemplary LNP formulation comprises (i) SM-102 (heptadecan-9-yl 8 ((2 hydroxyethyl)(6 oxo 6-(undecyloxy)hexyl)amino)octanoate) within a range of 45 to 55 mol percent of total lipids; (ii) DSPC within a range of 5 to 15 mol percent of total lipids; (iii) cholesterol within a range of 35 to 45 mol percent of total lipids; and (iv) PEG2000-DMG within a range of 1 to 2 mol percent of total lipids; and RNA molecules as described herein that are encapsulated within or associated with the lipid nanoparticles. [0878] In some embodiments, an exemplary LNP formulation comprises (i) ((4- hydroxybutyl)azanediyl)bis(hexane-6,1-diyl)bis(2-hexyldecanoate) (also known as 6-[N-6-(2-hexyldecanoyloxy)hexyl- N-(4-hydroxybutyl)amino]hexyl 2-hexyldecanoate) within a range of 45 to 55 mol percent of total lipids; (ii) DSPC within a range of 5 to 15 mol percent of total lipids; (iii) cholesterol within a range of 35 to 45 mol percent of total lipids; and (iv) a PEG-conjugated lipid within a range of 1 to 2 mol percent of total lipids; and RNA molecules as described herein that are encapsulated within or associated with the lipid nanoparticles. Example 2: Exemplary Characterization Studies [0879] The present Example describes certain potential characterization studies that may be utilized, for example, to identify, select, and/or characterize compositions (e.g., vaccines, e.g., manufacturing batches thereof), or components thereof as described herein. [0880] A potential immunization protocol that can be utilized to assess ability of a composition candidate, such as a trivalent composition, that comprises or delivers an HSV-2 antigen(s) or immunogenic fragment(s) thereof as described herein to induce B- and/or T-cells, e.g., after intramuscular immunization, directed to the antigen(s) and/or epitope(s) thereof. In some embodiments, level and/or diversity of response is determined. In some embodiments, presence and/or level of neutralizing antibodies is/are determined. In some embodiments, protection of the immunized subject from challenge with HSV is assessed. [0881] Alternatively or additionally, in some embodiments, one or more in vitro assessments may be performed, for example: (1) in vitro expression of an antigen encoded by an RNA included in a composition; and/or (2) in vitro potency of antigen expressed from an RNA included in a composition as described herein. Example 3: Exemplary Pre-clinical assessment [0882] The present Example describes certain pre-clinical assessments that may be performed of certain trivalent Composition described herein: [0883] In some embodiments, an exemplary trivalent composition candidate is assessed. In some embodiments, more than one different exemplary trivalent composition candidate may be assessed. In some such embodiments, different candidates may vary, for example, in: RNA platform (e.g., unmodified RNA, modified RNA, saRNA); Encoded antigen(s); Number of RNAs; Elements of RNA construct (e.g., cap and/or cap-adjacent sequences, 5’-UTR, 3’-UTR, and/or PolyA tail); and/or Lipid composition of LNP. [0884] In some embodiments, pre-clinical assessment of certain RNA compositions (e.g., LNP formulated RNA-based HSV compositions) comprises one or more of assessment in challenge experiments, assessment of level of protection, assessment of immunogenicity, and/or assessment of functional antibody responses. [0885] Exemplary LNP formulated RNA-based HSV compositions are tested in a challenge model. Non- human primate models, such as Rhesus macaques and Cynomolgus monkey, and/or rodent models, such as C57/B16 mice, Balb/c mice or NODscidIL2Rγnull mice; and/or guinea pig models, inoculated with HSV, are administered a first vaccination and can be administered an additional vaccination (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 additional vaccinations) following the first vaccination. Wherein more than one vaccination is administered, the vaccinations are administered at an interval of 1, 2, 3, 4, 5, 6, 7, or 8 week intervals). Following vaccination, animals are challenged by HSV. Alternatively or additionally, animals are challenged by intravenous, subcutaneous, and/or intramuscular injection of virus-infected lymphocytes. Lymphocytes can be infected with any suitable strain of HSV. Animals are then evaluated for reduced infection of neurons. In some embodiments, an animal model is challenged in a plurality of instances (e.g., before first vaccination and/or wherein additional vaccinations are administered, at any time point between or after vaccinations). Following challenge, animals subjected to the study may be assessed according to any method known in the art, including, for example, serology assessment, immunogenicity, level of protection, etc. [0886] In some embodiments, serum antibody characterization and/or serum transfer experiments (e.g., from one vaccinated species to a different non-vaccinated species, e.g., from vaccinated non-human primate to non- vaccinated mouse) are conducted (e.g., to assess protective antibody response). [0887] In some embodiments, certain exemplary RNA compositions (e.g., immunogenic compositions, e.g., vaccines) of the present disclosure are assessed for level of protection. Level of protection can be assessed according to any suitable method known in the art. [0888] In some embodiments, certain exemplary RNA compositions (e.g., immunogenic compositions, e.g., vaccines) of the present disclosure are assessed for immunogenicity. For example, ELISA can be used to determine IgG specific (and subclasses thereof) titers and/or avidity of antibodies generated in response to certain RNA compositions (e.g., immunogenic compositions, e.g., vaccines) of the present disclosure to HSV antigens. In some embodiments, serum antibody titers against HSV glycoprotein (e.g., gC, gD, and/or gE glycoprotein, etc.) is determined by ELISA using standard methods. In some embodiments, for example, ELISpot (e.g., for CD8+, CD4+ T cells and/or IFNγ) and assessment of pro-inflammatory cytokine responses with splenocytes from immunized and/or challenged animal models and peptide pools derived from composition (e.g., immunogenic composition, e.g., vaccine) targets can also be assessed. In some embodiments, for example, phenotyping of immune responses (e.g., by flow cytometry) are assessed. In some embodiments, for example, T cell depletion and/or protection assays are conducted to assess immunogenicity (e.g., according to any suitable known method in the art). [0889] In some embodiments, one or more functional responses of antibodies generated in response to certain RNA compositions (e.g., immunogenic compositions, e.g., vaccines) of the present disclosure are assessed. Functional antibody responses can be assessed, for example, using an HSV neutralization assay. In some embodiments, an HSV in vitro neutralization assay is performed to evaluate one or more anti-HSV glycoprotein (e.g., HSV gC, gD gE, or a combination hereof) antibodies in neutralizing HSV. For example, anti-HSV glycoprotein antibodies are obtained by collecting the sera of animals (e.g., mice) vaccinated with HSV RNA compositions. HSV virus are added to the diluted sera and neutralization is allowed to continue for 1 hour at room temperature. 3T3 cells are seeded in 96-wells one day before and the virus/serum mixtures are added to 3T3 monolayers. The cells are fixed on the next day and HSV-specific staining is performed. The plates are scanned and analyzed. A neutralization titer is expressed as the highest serum dilution required to achieve a 50% reduction in the number of plaques. [0890] In some embodiments, functional antibody responses can be assessed, for example, using passive transfer studies of sera from immunized animals to naïve animals that are challenged and assessing level of protection. Example 4: In Vitro testing [0891] The present Example provides in vitro transfection and expression data for a composition comprising three RNAs described herein: a first RNA encoding an exemplary gC2 antigen or immunogenic fragment thereof, a second RNA encoding an exemplary gD2 antigen or immunogenic fragment thereof, and a third RNA encoding an exemplary gE2 antigen or immunogenic fragment thereof. In the present Example, an exemplary combination comprising modRNAs comprising SEQ ID NOs: 106, 118, and 123 and encoding exemplary polypeptides having amino acid sequences according to SEQ ID NOs: 65, 70, and 73 was characterized. HEK293T cells were transfected in triplicates with 0.3 μg/mL modRNA using a commercial transfection reagent or 0.3 μg/mL LNP- formulated modRNA. After transfection, cells were incubated to express the modRNA for 18h. Afterwards, cells were harvested and probed with antigen-specific mouse monoclonal antibodies and a secondary fluorescence-tagged anti- mouse antibody to detect expression of composition (e.g., vaccine) antigens in HEK293T cells. The median fluorescence intensity (MFI) measured in FACS serves as a surrogate for antigen expression. [0892] Briefly, 0.4x106 HEK293T cells were seeded 6 hours prior to transfection in 12-well plates. Exemplary Trivalent modRNA (Drug Substance (DS)) encoding for exemplary HSV-2 gC, HSV-2 gD, and HSV-2 gE immunogenic fragments was formulated using Lipofectamine™ MessengerMAX™ (ThermoFisher Scientific) according to the manufacturers’ instructions prior to transfection, or LNP-formulated modRNA (Drug Product (DP)) encoding exemplary HSV-2 gC, HSV-2 gD, and HSV-2 gE immunogenic fragments was used to transfect cells in triplicates at a concentration of 0.3 μg/mL modRNA. Cells were incubated for 18 hours at 37°C and 5% CO2 prior to staining. Afterwards, cells were harvested and incubated with a viability dye (eBioscience™ Fixable Viability Dye eFluor™ 450, ThermoFisher Scientific), fixed (Fixation Buffer, BioLegend), permeabilized (Permeabilization Buffer, eBioscience) and incubated with a mouse anti-gC, gD or gE antibody and subsequently stained with an Alexa Fluor® 647 AffiniPure Donkey Anti-Mouse IgG (H+L) secondary antibody (Jackson ImmunoResearch). Cells were acquired using a BD FACSCelesta 2 (BD) and data was analyzed with FlowJo V10.8 (BD). [0893] As demonstrated in FIG.3, all exemplary composition (e.g., vaccine antigens) were delivered to and expressed in HEK293T cells. DP transfection led to slightly higher frequencies of transfected cells compared to DS transfections with Lipofectamine™ MessengerMAX™ (FIGS.3(A), (C), AND (D)), while DS transfection using Lipofectamine™ MessengerMAX™ led to higher total expression levels in HEK293T cells compared to those obtained with LNP-formulated modRNA (FIGS.3(B), (E), (F)), as seen in the percentage of antigen expressing cells and the MFI of the total HEK293T population. [0894] High transfection frequencies and strong expression was observed with both DP and DS transfections. Both transfection methods also showed comparable transfection frequencies and expression levels when comparing the three different antigens. These results demonstrate that (i) exemplary RNAs described herein can induce strong antigen expression, and (ii) exemplary RNAs described herein can be administered in combination without significantly interfering with transfection efficiency or antigen expression. Example 5: Confirmatory pharmacology study of exemplary trivalent composition in guinea pigs [0895] The objective of the study summarized in the present Example was to analyze the tolerability, immunogenicity, and disease protection of a combination comprising a pol
Figure imgf000407_0001
yrbonuceotde encodng an exemplary HSV-2 glycoprotein C (gC) immunogenic fragment, a polyribonucleotide encoding an exemplary HSV-2 glycoprotein D (gD) immunogenic fragment and a polyribonucleotide encoding an exemplary HSV-2 glycoprotein E (gE) immunogenic fragment and formulated in lipid nanoparticles comprising ALC-0315 (4-hydroxybutyl) azandiyl bis(hexane-6,1-diyl) bis (2-hexyl decanoate). In the present Example, the composition tested comprised three exemplary polyribonucleotides: a first comprising SEQ ID NO:106 (construct 1600) and encoding an exemplary polypeptide having an amino acid sequence according to SEQ ID NO: 65, a second comprising SEQ ID NO: 118 (construct 1601) and encoding an exemplary polypeptide having an amino acid sequence according to SEQ ID NO: 70, and a third comprising SEQ ID NO: 123 (construct 1602) and encoding an exemplary polypeptide having an amino acid sequence according to SEQ ID NO: 73. [0896] In guinea pigs, the composition induced potent antigen-specific immunoglobulin G (IgG) antibodies in serum and vaginal mucosa, as well as high titers of neutralizing antibodies in serum. 100% protection was provided against a highly lethal intravaginal HSV-2 challenge, while 70% of animals succumbed to viral challenge in the control group. Furthermore, days and severity of genital lesions as well as urinary retention were reduced in immunized animals and vaginal titers tend to be decreased on day 2 and 4 after challenge compared to the unvaccinated control group. Results of the study support the use of HSV-2 compositions tested herein as a clinical candidate for human trials. Background [0897] Genital herpes, caused by HSV, is a sexually transmitted infection. To date, a composition (e.g., immunogenic composition, e.g., vaccine) has not been licensed to prevent genital lesions resulting from herpes simplex virus-1 (HSV-1) or 2 (HSV-2) infection, despite extensive efforts by major pharmaceutical companies. A strategy behind the RNA composition (e.g., immunogenic composition, e.g., vaccine) candidate tested in the present Example is to induce immune responses that block HSV-2 cellular entry, inhibit cell-to-cell spread, and counteract HSV-2’s inhibitory effects on the complement system and virus clearance. The composition (e.g., immunogenic composition, e.g., vaccine) candidate characterized herein includes three exemplary HSV-2 immunogens, namely exemplary immunogenic fragments of HSV-2 glycoproteins C, D, and E (HSV-2 gC, HSV-2 gD, and HSV-2 gE), formulated with ALC-0315 lipid nanoparticles. [0898] An earlier generation of exemplary trivalent HSV-2 gC, HSV-2 gD, and HSV-2 gE immunogenic fragments composition (e.g., immunogenic composition, e.g., vaccine) containing recombinantly expressed immunogenic fragments of ectodomains of these glycoproteins in a baculovirus system, formulated with CpG oligonucleotides and an aluminum salt as adjuvant, demonstrated immunogenicity and efficacy in mice. However, guinea pigs immunized with the same trivalent protein composition (e.g., immunogenic composition, e.g., vaccine) still developed subclinical infection based on vaginal shedding of HSV-2 DNA during the recurrent phase of infection (Awasthi et al. 2017, Hum Vaccin Immunother. 2017 Dec 2;13(12):2785-2793). To improve composition (e.g., immunogenic composition, e.g., vaccine) efficacy, the same three glycoprotein antigens were produced by immunizing mice and guinea pigs with an RNA-LNP composition instead of recombinant protein. Superior immunogenicity and efficacy of the RNA-LNP composition compared to the baculovirus glycoprotein CpG/alum composition was shown in both mice and guinea pigs (Awasthi et al. 2019, Sci Immunol. 2019 Sep 20). [0899] Previous studies found that formulation of the RNA composition with ALC-0315 lipid nanoparticles (RNA-LNP315) increased immunogenicity compared to formulation with ALC-0307 (RNA-LNP307). Therefore, ALC- 0315 was chosen as a component of the exemplary LNP formulation used in the present Example. [0900] Additionally, as previous experiments showed complete protection in most guinea pigs, lower doses, and a prime boost regimen (without a second boost) were chosen in this study. This approach allows coverage of a broader range of disease severity when guinea pigs were challenged with a highly lethal dose of HSV-2 virus. Objective [0901] The objective of this study was to confirm tolerability, immunogenicity, and disease protection of an exemplary HSV-2 RNA composition candidate (a combination comprising a polyribonucleotide encoding an exemplary HSV-2 glycoprotein C (gC) immunogenic fragment, a polyribonucleotide encoding an exemplary HSV-2 glycoprotein D (gD) immunogenic fragment and a polyribonucleotide encoding an exemplary HSV-2 glycoprotein E (gE) immunogenic fragment) in guinea pigs. Study Design [0902] Thirty female Hartley guinea pigs, weighing 300 to 350 g, were divided into three experimental groups (10 animals each) and immunized on day 0 and day 28 (PBS as control, and 3 μg or 15 μg of Exemplary RNA composition, see Table 24, below). The study design is displayed in FIG.4. Table 24: Experimental groups and treatment Group Test article Dose Immunization Challenge -2 .
Figure imgf000408_0001
[0903] Blood samples were collected at day 56, 4 weeks after the second immunization, for analysis of HSV-2 specific antibody responses. Levels of IgG antibodies against HSV-2 gC, HSV-2 gD, and HSV-2 gE were measured by ELISA. Neutralizing antibody titers were determined using a serum HSV-2 plaque reduction assay and defined as highest dilution of serum with 5% human complement that reduced the number of HSV-2 plaques by 50%. Vaginal swabs were also collected 4 weeks after the second immunization for analysis of mucosal antibody titers. Vaginal swabs were placed into 100 μL PBS and a serial two-fold dilution was tested for HSV-2 gC, HSV-2 gD and HSV-2 gE IgG by ELISA. [0904] At day 60, animals were challenged with a lethal dose of 5 x 105 plaque forming units (PFU) of HSV-2 strain MS (25-fold LD50) and scored for survival and signs of genital disease for a period of 48 days. Body weights were recorded daily for the first 2 weeks post-infection. Day 2 and day 4 vaginal titers, and vaginal shedding of HSV-2 DNA between days 28 to 48 post-infection (p.i.) were also determined. Vaginal shedding of HSV-2 DNA was analyzed by a qPCR assay, together with HSV-2 DNA copy in DRG (dorsal root ganglia) and spinal cord at day 48. Materials and Methods [0905] Test items included three codon-optimized, 5’capped, modRNA constructs encoding for exemplary HSV-2 glycoprotein C (gC), D (gD) and E (gE) immunogenic fragments, encapsulated at a 1:1:1 ratio within ALC- 0315 (LNP315). Sterile PBS was used as control. Animal care [0906] Guinea pigs were pair housed under a 12:12-h light:dark cycle in 2000P Tecniplast cages (Buguggiate, Italy), on cellulose fiber animal bedding in an ABSL-2 vivarium. The room temperature was maintained between 68 to 79°F and humidity 30 to 70% with air exchange 10 to 15/h in the room. Guinea Pig Diet 5025 (LabDiet, Richmond, IN), Oxbow timothy hay, and reverse osmosis water were provided ad libitum and cages were changed twice weekly. All husbandry materials were autoclaved prior to use. [0907] Routine animal monitoring was carried out daily and included inspection for dead animals and control of food and water supplies. Each animal's health was monitored at least twice weekly prior to HSV-2 infection and daily after HSV-2 infection until the end of the experiment. The general physical condition was assessed with the following parameters: x Body weight loss ≥ 20% x Labored/gasping breathing x Signs of lethargy x Pain when handled x Genital disease x Hindlimb weakness x Urinary retention Treatment schedule, route of administration, and dose [0908] Test items were diluted in PBS and administered via IM injection on day 0 and day 28 (see Table 24, above). The control group was injected with sterile PBS. 50 μL was injected into the right hind muscle using an insulin syringe (29-gauge needle). [0909] Blood was sampled via the lateral saphenous vein in the left hind limb. In short, the animal was held face down with the left lateral aspect of the hind limb facing the person performing the bleed. The fur off the left hind leg was shaved up to the knee and Vaseline was applied. A 20-gauge needle was used to prick the vein and blood droplets (totaling between 100 to 1000 μL) were collected into an Eppendorf tube. To allow the blood to clot, the samples were placed on ice for at least 30 min. Blood samples were centrifuged at 8000 RPM for 10 min. The supernatant (serum) was collected and stored at -20oC. [0910] Immunized guinea pigs were challenged with a dose of 5 x 105 PFU HSV-2 MS at day 60, approximately 1 month after the last immunization. On the day of infections, viral stocks of HSV-2 (strain MS from the American Type Culture Collection [ATCC]) were thawed on ice. Thawed stocks were then diluted to the desired concentration (5 x 105 PFU/50 μL) with sterile Dulbecco’s Modified Eagle’s medium (DMEM) tissue culture media. A polyester swab (Puritan) dipped into phosphate buffered saline (PBS) was used to clear the mucus from the vaginal vault. The guinea pig was held with the head slightly down and 25 μL of HSV-2 virus was inoculated into the vaginal vault using a soft catheter attached to a pipette tip. The guinea pig was held in position for a further 30 to 60 second and then placed back into the cage. A second HSV-2 inoculation was performed 1 hour later to inoculate the remaining 25 μL. [0911] Guinea pigs were scored for signs of acute disease on days 1 to 14 after infection and for signs of recurrent disease on days 15 to 48. Naive non-immunized guinea pigs are expected to show clear indications of genital herpes disease 3 to 4 days after infection. [0912] For scoring the severity of genital disease, a score of 1 to 4 was assigned: x Score 1: 1 to 2 distinct genital lesions x Score 2: > 2 distinct genital lesions x Score 3: coalesced lesions x Score 4: ulcerated lesions x Animals are evaluated daily for urinary retention (recorded when present) x Animals are observed daily for evidence of hindlimb weakness (recorded when present) [0913] On day 2 and 4 post-infection, vaginal swabs were performed to document infection in the vaginal canal. Replicating virus was titrated using plaque assay. The cotton swab was inserted into the vaginal cavity and rotated six times to collect the tissue. The swab was then removed and placed in 1 ml of swab media (DMEM with 5% heat-inactivated FBS, 1% L-Glut, 1% Antibiotic/antimycotic, 0.4% Gentamycin, 0.025% Vancomycin and 1.5% HEPES). The aluminum handle was clipped, and the swab was left in the tube. The tubes were stored at -80°C. Vaginal wash procedure to obtain IgG titers in ELISA [0914] Vaginal secretions for neutralizing titers were obtained using an eye spear swab (BVI). The eye spear was trimmed 1 mm from each side and inserted into the guinea pig vaginal cavity for 45-60 s. A small hole was pricked in the bottom of a 0.5 ml reaction tube using a 20-gauge needle. Next, the spear was placed into this tube, and both together were placed into a 1.5 ml reaction tube. 100 μl of PBS were added directly onto the spear and the tubes were centrifuged for 3 min at 8000 RPM. The vaginal fluid and PBS were collected in the 1.5 mL tube and stored at -80°C. Isolation of dorsal root ganglia (DRG) and spinal cord [0915] Spinal cord and DRG were isolated at the time of termination of experiment. Guinea pigs were anesthetized with Euthasol (IP) at 150 mg/kg of body weight. Once they lost consciousness (confirmed by toe pinch), cardiac bleed (~10ml) was performed. The animal’s back was shaved, and the skin lifted to expose the spinal column. The spinal column was cut out along with muscle on either side from mid-back to the tail. Next, the spinal column was opened longitudinally with sharp surgical scissors to expose the spinal cord. A portion of the spinal cord (about 5/8” from the lower end) was collected into a 1.5 ml tube containing 1 ml DMEM media and stored at -80°C. [0916] Next, the remaining spinal cord was gently removed to expose the DRG on either side of vertebral column. The entire DRG root (not just the DRG) was pulled from the vertebral canals of lumbosacral region by going deep into the vertebral canals. Each ganglion was placed in 1 ml DMEM media in a 1.5 ml tube. Tubes were stored at -80°C until further use. Endpoint of experiment / termination criteria [0917] Animals were euthanized in accordance with the Panel on Euthanasia of the American Veterinary Medical Association by intraperitoneal injection of Euthasol euthanasia solution (1 mL for up to a 1 kg guinea pig). Additionally, termination criteria applied according to the recommendation of Panel on Euthanasia of the American Veterinary Medical Association. Body weight losses exceeding 20%, or a high severity level in any of the other categories described below were on their own sufficient reason for immediate euthanasia. ELISA protocol to detect IgG antibody responses to exemplary HSV-2 gC, HSV-2 gD, and HSV-2 gE immunogenic fragments in guinea pig sera and vaginal wash samples [0918] Serum and vaginal wash samples were tested in 96-well plates to detect HSV-2 gC-, HSV-2 gD-, and HSV-2 gE-specific antibody concentrations. Briefly, 4 weeks after the last injection, serum and vaginal wash samples were analyzed by endpoint titration. HSV antigens (recombinant exemplary immunogenic fragments of HSV- 2 gC, HSV-2 gD, and HSV-2 gE from a Baculovirus-expression system) were diluted to a final concentration of 1 μg/mL in 50 mM sodium bicarbonate binding buffer, pH 8.5-9.0. Wells of a 96-well ELISA plate were coated with 100 μl of HSV antigen (100 ng/well). Thereafter, the plate was covered with adhesive plastic and incubated overnight at 4oC. After aspirating the coating solution, the plate was washed three times using the Wellwash with 300 μL PBS/0.05% Tween-20 (PBST). Remaining drops were removed by blotting the plate on a paper towel. Wells were blocked by adding 250 μL of 5% milk in PBST per well. The plate was covered with adhesive plastic and incubated for 2 hours at room temperature with gentle agitation. [0919] During the blocking step, guinea pig sera was diluted 1:500 in PBST. Thereafter, two-fold serial dilutions were performed in PBST so that the following dilutions were available to add to the plate: 1:500, 1:1,000, 1:2,000, 1:4,000, 1:8,000, 1:16,000, 1:32,000, 1:64,000, 1:128,000, 1:256,000, 1:512,000, and 1:1,024,000. Vaginal wash was diluted 1:50 in PBS. Once blocking was complete, the plate was washed three times with 300 μL PBST and blotted dry on a paper towel. Then, 100 μL of the diluted sera or vaginal wash was added to each well. The plate was covered with an adhesive sheet and incubated for 1 hour at room temperature with gentle agitation. Secondary antibody (rabbit anti-guinea pig IgG) was prepared by diluting 1:2,000 in PBST. The plate was washed three times with 300 μL PBST, blotted dry on a paper towel, and 100 μL of the secondary antibody was added to each well of the ELISA plate. After covering the plate with an adhesive sheet, it was incubated for 30 min at room temperature with gentle agitation. The plate was washed three times with 300 μL PBST, blotted dry on a paper towel, and 100 μL of ABTS (1-Component Microwell Peroxidase Substrate) was added to each well of the ELISA plate. Again, the plate was covered with foil and incubated for 30 min at room temperature with gentle agitation. Subsequently, 100 μL of stop solution (1x) were added per well. [0920] The absorbance was measured within 30 minutes (MRX Revelation) at 405 to 410 nm (405 nm). Endpoint titers were calculated as the serum dilution giving an optical density (OD) reading >0.1 and at least 2-fold higher than background OD. Background OD - wells were treated with everything but using PBS instead of guinea pig sera or vaginal wash. Neutralization antibody titer using complement by plaque reduction assay [0921] The aim of this analysis was to determine the dilution of serum from vaccinated and mock vaccinated guinea pigs required for 50% neutralization of the HSV-2 virus in the presence of human complement. The source of complement was whole serum from an HSV-1/HSV-2 negative human individual. [0922] For this assay, Vero cells were seeded on 24-well plates at a dilution of 2 x 105 cells per well overnight. The following day, 2-fold serum dilutions ranging from 1:10 to 1:10,240 were prepared in DMEM in a volume of 50 μL and mixed with 45 μL of virus solution containing 100 PFU and 5 μL of human complement. For the initial 1:10 serum dilution, 10 μL of undiluted sera was mixed with 40 μL of media, 45 μL of virus and 5 μL of human complement. The sera, virus and complement were mixed in a 96-well u-bottom plate by gentle rocking at 37°C for 1 hour. As a negative control for neutralization, PBS was mixed with HSV-2 and human complement. [0923] Then, the media from the Vero cells was replaced with 100 μL of the mixture containing serum (or PBS), virus, and complement, and incubated at 37°C for 1 hour with rocking every 10 minutes to ensure that the cells did not dry out. [0924] After 1 hour, the 100 μL virus / serum mix was aspirated and replaced with 1 mL of 1.5% CMC- DMEM per well. The plate was then incubated at 37°C for 68 to 72 hours. [0925] For staining of the plaques, the CMC-DMEM overlay was gently aspirated and 0.5 mL/well crystal violet dye (0.05% w/v) was added. The plate was incubated at room temperature for 30 min to 1h. The crystal violet dye was aspirated and 1 mL water/well was added to remove traces of the dye. Thereafter, the water was aspirated, and the remaining water was removed by inverting plate on a blotting paper and allowing the plate to air-dry. Plaques were counted under the inverted light microscope. [0926] Dilution of serum required to reduce virus plaques by 50% was considered as the neutralization titer for that serum sample. Virus titers of vaginal swabs from guinea pig by plaque assay [0927] This protocol assumed that at least 1 x 75 cm2 flask of Vero cells ready for splitting were available before the plaque assay. Briefly, swab sample dilutions were prepared in a 96-well plate such that one 96-well plate corresponded to four 24-well plates (i.e., A1-A6 = plate 1, A7-A12 = plate 2, E1-E6 = plate 3, and E7-E12 = plate 4). 225 μL of swab medium was aliquoted to each 24-well plate. Undiluted stock swab virus was prepared by adding 250 μL of the thawed swab samples to 225 μL of swab medium in the corresponding well. The -1 dilution was prepared by adding 25 μL of stock swab virus to 225 μL of swab medium. This procedure was repeated to obtain the -2, and -3 dilutions of the swab virus. Vero cells in a 24-well plate (~80% confluent) were infected with the swab virus samples starting with the -3 dilution through undiluted stock swab virus. The plate was rocked 10 x and placed in an incubator. Rocking was repeated every 5 min for the first 15 min followed by every 15 min for a total of 1 h. Next, 0.5 mL of CMC-DMEM overlaying medium pre-warmed to 37°C was added to each well. Plates were incubated at 37°C for 72 hours. [0928] Following this, the incubating cells were fixed and stained by adding enough 0.05% crystal violet such that the bottom of the plate was covered and allowed to stain for 3 hours. The next day plaques were counted by scoring the plate in a grid-pattern like using a hemocytometer (i.e., cells touching top and left grid lines) under a microscope. The dilution at which 10-30 plaques per well were counted was the dilution factor (e.g., dilution factor = 10-2 if 10 to 30 plaques were counted in the -2 dilution). Swab titer was calculated by taking the plaque count and multiplying by the 1 x 10n to get PFU/200 μL where n is the dilution factor e.g., 14 plaques at 10-2 dilution is equal to 14 x 10-2 = 1400 PFU/200 μL. Finally, swab titer was reported PFU/mL by multiplying PFU/200 μL by five. qPCR on DNA in vaginal swabs and isolated DRG and SP [0929] qPCR assays on guinea pig vaginal swab samples, as well as in samples from isolated DRG and SP were performed in duplicate for HSV-2 DNA. Separate reactions were used to amplify HSV-2 DNA and the guinea pig GAPDH. The DRG HSV-2 DNA copy number was expressed as log10 DNA copies per 106 guinea pig GAPDH genes. The HSV-2 qPCR assay was designed to detect low levels of HSV-2 DNA in swab samples from guinea pig in vivo studies. The primers and probes used in this assay were designed from published sequences to type-specific and type- common genes of HSV-2. Specific primers and probes were also designed from published sequences to host housekeeping genes to monitor consistency of sampling and to allow for normalizing of final results. (See Table 25, below, for primer and probe sequences). [0930] The quantification method used a standard curve that was prepared from a dilution series of control template of known concentration. The standard curve was generated by plotting the log10 of the initial template copy number against the cycle threshold (Ct) value generated at each dilution. Ct values from samples were then compared to the template controls of the standard curve and a template quantity was then estimated. Table 25: Primer/Probe sets SEQ ID NO Primer/probe ’ ’
Figure imgf000413_0001
[0931] First, the master mix was prepared using the ratio shown in Table 26, below. 20 μL master mix was added to appropriate wells and stored covered at 4°C until use. Table 26: Ratio for master mix preparation Reagent Vol/Rxn (μL) Primer/Probe Mix Prep
Figure imgf000413_0002
Reagent Vol/Rxn (μL) Primer/Probe Mix Prep Primer/Probe Mix 1.5 Combine 60 μL each: forward and
Figure imgf000414_0001
0.6 mL tubes with nuclease-free water were prepared in the PCR laminar flow cabinet (see below for volume) and dilution series were completed for appropriate DNA type. [0933] Then, 5 μL of test samples or standard concentration were added to the wells containing 20 μL of the Master Mix. The plate was covered with an optical film and centrifuged for 1 min at 1000 RPM (190 x g). [0934] PCR procedure was performed on Applied Biosystems 7500 Fast PCR Machine with the following setup: x Holding state: step 1: 95ºC 20 sec x Cycling state: step 1: 95ºC 03 sec; step 2: 60ºC 45 sec: x Number of cycles: 40 x Analyze results using Applied Biosystems’ software [0935] Limit of quantitation was established based on the total volume of media holding the swab, amount of swab media used for DNA isolation and the amount of DNA used for the PCR amplification. In this case, the total volume of swab media was approximately 1 mL, 200 μL was used to isolate DNA, that was resuspended in 200 μL, and 5 μL of DNA was used for DNA amplification. One copy of viral genome could be detected in this assay; therefore, the limit of quantitation was less than 200 HSV genome copy per swab. [0936] Limit of detection was established based on the volume of DNA used per reaction. In this assay limit of detection was less than one HSV genome per amplification reaction. Statistical analysis [0937] GraphPad Prism 9 Software (La Jolla, USA) was used for statistical analysis and figure generation. As data did not pass test for normal distribution, Kruskal-Wallis-Test with uncorrected Dunn’s test was performed for analysis of multiple data sets, and non-parametric Mann-Whitney for testing two data sets. For analysis of survival, log-rank (Mantel-Cox) test was performed. Results Immunogenicity Serum and vaginal IgG titers after second immunization [0938] Serum and vaginal IgG antibodies against gC2, gD2, and gE2 were assessed by ELISA in samples obtained at day 56, 4 weeks after the second immunization. Results are shown in FIGS.5 AND 6. [0939] The PBS control had no detectable serum or vaginal IgG antibodies. IgG titers could be detected for both doses for all three antigens with the exception of vaginal titers against gE2 at a 3 μg dose (gC2, serum 3 μg p:0.0068 and 15 μg p:<0.0001, vaginal 3 μg p:0.0018 and 15 μg p:<0.0001; gD2, serum 3 μg p:0.0013 and 15 μg p:<0.0001, vaginal 3 μg p:0.0003 and 15 μg p:0.0007, gE2, serum 3 μg p:0.0085 and 15 μg p:<0.0001, vaginal 15 μg p:0.0001). A dose-response was observed between the 3 μg and 15 μg groups for gC2 serum titers, and gE2 serum and vaginal titers (gC2, serum p: 0.0420; gE2, serum p: 0.0409, vaginal p:0.0002). Neutralizing antibodies by plaque reduction assay [0940] Virus-neutralizing antibodies in guinea pig serum were measured by plaque reduction assay using samples obtained one month after the second immunization. Both doses led to dose-dependent serum neutralizing antibody titers (FIG.7; p: 0.0234). HSV-2 challenge [0941] At day 60, approximately one month after the second (final) immunization, animals in all groups were challenged with a lethal intravaginal dose of HSV-2 MS strain and scored for survival and signs of genital disease and vaginal virus titers for a period of up to 48 days. Weight loss [0942] A comparable minor transient decrease in body weights was observed from day 5 post-challenge onwards for some animals in both groups receiving exemplary HSV-2 modRNA composition after the first and second immunization (FIG.8). Body weight loss was more evident and not transient for most animals in the PBS group and for one animal dosed with 3 μg of modRNA composition. Survival [0943] All animals dosed with 15 μg a combination comprising a polynucleotide survived viral challenge with HSV-2 MS strain at a dose of 25-fold LD50, while one animal in the 3 μg dose group and seven animals in the PBS group succumbed to disease within 15 days (FIG.9). Animals dosed with 3 μg or 15 μg showed a significantly higher probability of survival (3 μg, p:0.0092; 15 μg, p:0.0012). Disease scores [0944] Disease severity, incidence, and duration of genital lesions as well as urinary retention were reduced in all immunized groups (FIG.10). As most animals in the PBS group succumbed to viral challenge within the first 2 weeks after challenge (red bordered symbols), scoring genital lesions and urinary retention by days leads to an underrepresentation in disease scores in this group. Therefore, only immunized groups were included in the statistical analysis. Animals dosed with 15 μg showed a tendency towards decrease in days with genital lesion (FIG. 10A, p:0.0664) and a significant decrease in severity (FIG.10B, p:0.0444) compared to 3 μg dosed animals, while no significant difference was detected in urinary retention (FIG.10C). [0945] In addition to individual scores, mean cumulative days of disease up to 48 days after challenge were analyzed (FIG.11). No scoring for days after death was assigned to animals that succumbed to viral disease. In addition to a reduction of days with disease directly after challenge, immunized animals showed a decrease in reoccurring genital disease over the course of 48 days. Vaginal virus titers [0946] Two days and four days after viral challenge, vaginal virus titers were determined by plaque assay (FIG.12). Although results were not significant, a clear tendency of reduction in vaginal virus titers was overserved in vaccinated groups for day 2 and day 4 compared to the PBS control. Appreciable differences were not apparent when comparing the 3 μg dose group with the 15 μg dose group on day 2 or 4, nor on mean days of shedding between day 28 to 48. Mean days of shedding of vaccinated animals were not compared to PBS controls, as a reasonable comparison was not possible since only three out of ten guinea pigs survived viral challenge. HSV-2 DNA copy in DRG and spinal cord [0947] At day 48 (end of the study) post viral challenge, guinea pigs were euthanized and DRG (FIG. 13(A)) and spinal cord tissue (FIG.13(B)) were collected. Both were analyzed for HSV-2 genomic copy number by quantitative PCR (qPCR) to assess latent infection. Separate reactions were used to amplify HSV-2 DNA and the GAPDH gene was used as control. HSV-2 DNA copy number was expressed as log10 DNA copies per 106 GAPDH genes. HSV-2 DNA copy numbers of vaccinated animals were not compared to PBS controls, as a reasonable comparison was not possible because only 3 out of 10 guinea pigs survived viral challenge. No appreciable differences were apparent when comparing the 3 μg dose group with the 15 μg dose group regarding HSV-2 DNA copy numbers in DRG and spinal cord. Conclusion [0948] The exemplary HSV-2 composition characterized in the present Example induced potent antigen- specific IgG antibodies in serum and vaginal mucosa, as well as high titers of neutralizing antibodies in serum. The clinical candidate provided protection against a highly lethal intravaginal challenge with HSV-2 in guinea pigs (for 90% at 3 μg dose and for 100% of animals at 15 μg dose), while only 30% of animals survived viral challenge in the control group. A significant dose-dependent response was shown in the induction of neutralizing antibody titers. The same tendency was observed with induction of serum and vaginal antibody responses upon immunization. [0949] Furthermore, days and severity of genital lesions as well as urinary retention were reduced in immunized animals and vaginal titers tended to be decreased on day 2 and 4 after challenge compared to the unvaccinated control group. Overall, the results of this study support the HSV-2 composition as a clinical composition (e.g., immunogenic composition, e.g., vaccine) candidate. [0950] References Cited in Example 5: x Awasthi S, Hook LM, Shaw CE, Friedman HM. A trivalent subunit antigen glycoprotein vaccine as immunotherapy for genital herpes in the guinea pig genital infection model. Hum Vaccin. Immunother. 2017 Dec 2;13(12):2785-2793 x Awasthi S, Hook LM, Pardi N, Wang F, Myles A, Cancro MP, Cohen GH, Weissman D, Friedman HM. Nucleoside-modified mRNA encoding HSV-2 glycoproteins C, D, and E prevents clinical and subclinical genital herpes. Sci Immunol. 2019 Sep 20 Example 6: In Vitro Testing of Further Polyribonucleotides Encoding Exemplary HSV-2 Antigens [0951] The present Example demonstrates high transfection rates and expression levels in HEK293T cells transfected with exemplary nucleoside-modified RNA (modRNAs) encoding exemplary gC, gD, or gE immunogenic fragment variants. [0952] HEK293T cells were transfected with 0.2 μg/mL modRNA encoding an exemplary gC2 antigen immunogenic fragment, an exemplary gE2 antigen immunogenic fragment or an exemplary gD2 antigen immunogenic fragment using a commercial transfection reagent. In the present Example, the following modRNAs were transfected into HEK293T cells: x For the exemplary gC2 antigen immunogenic fragment: a modRNA comprising a ribonucleic acid sequence according to SEQ ID NO:106 (construct 1600) encoding a polypeptide that comprises an exemplary IL2 secretory signal and an exemplary gC2 antigen immunogenic fragment; a modRNA comprising a ribonucleic acid sequence according to SEQ ID NO: 204 (construct 2138) encoding a polypeptide that comprises an exemplary gE2 secretory signal and an exemplary gC2 antigen immunogenic fragment; a modRNA comprising a ribonucleic acid sequence according to SEQ ID NO: 192 (construct 2140) encoding an exemplary polypeptide that comprises an exemplary gD1 secretory signal and an exemplary gC2 antigen immunogenic fragment; a modRNA comprising a ribonucleic acid sequence according to SEQ ID NO: 196 (construct 2141) encoding a polypeptide that comprises an exemplary gB1 secretory signal and an exemplary gC2 antigen immunogenic fragment; or SEQ ID NO: 107 (construct 1876) encoding a polypeptide that comprises an exemplary IL2 secretory signal and an exemplary gC2 antigen immunogenic fragment; x for the exemplary gD2 antigen immunogenic fragment: a modRNA comprising a ribonucleic acid sequence according to SEQ ID NO: 118 (construct 1601) encoding an exemplary polypeptide that comprises an exemplary gD2 secretory signal and an exemplary gD2 antigen immunogenic fragment; a modRNA comprising a ribonucleic acid sequence according to SEQ ID NO: 117 (construct 1874) encoding an exemplary polypeptide that comprises an exemplary gD2 secretory signal and an exemplary gD2 antigen immunogenic fragment; a modRNA comprising a ribonucleic acid sequence according to SEQ ID NO: 120 (construct 1877) encoding an exemplary gD2 secretory signal and an exemplary gD2 antigen immunogenic fragment; or a modRNA comprising a ribonucleic acid sequence according to SEQ ID NO: 119 (construct 1659) encoding an exemplary gD2 secretory signal and an exemplary gD2 antigen immunogenic fragment; or x for the exemplary gE2 antigen immunogenic fragment: a modRNA comprising a ribonucleic acid sequence according to SEQ ID NO: 123 (construct 1602) encoding an exemplary polypeptide that comprises an exemplary IL2 secretory signal and an exemplary gE2 antigen immunogenic fragment; a modRNA comprising a ribonucleic acid sequence according to SEQ ID NO: 127 (construct 1660) encoding an exemplary polypeptide that comprises an exemplary gD2 secretory signal and an exemplary gE2 antigen immunogenic fragment; or a modRNA comprising a ribonucleic acid sequence according to SEQ ID NO: 212 (construct 2143) encoding an exemplary polypeptide that comprises an exemplary gD1 secretory signal and an exemplary gE2 antigen immunogenic fragment. [0953] Briefly, 0.4x106 HEK293T cells were seeded 6^h prior to transfection in 12-well plates. modRNA encoding for exemplary HSV-2 gC (gC2), gD (gD2) or gE (gE2) immunogenic fragment variants were formulated using Lipofectamine™ MessengerMAX™ (ThermoFisher Scientific) according to the manufacturers’ instructions prior to transfection in triplicates at a concentration of 0.2 μg/mL modRNA. After transfection, cells were incubated to express the modRNA for 18^h at 37°C and 5% CO2 prior to staining. Afterwards, cells were harvested and incubated with a viability dye (eBioscience™ Fixable Viability Dye eFluor™ 450, ThermoFisher Scientific), fixed (Fixation Buffer, BioLegend), permeabilized (Permeabilization Buffer, eBioScience) and incubated with a mouse anti-gC2, anti-gD2 or anti-gE2 antibody and subsequently stained with an Alexa Fluor® 647 AffiniPure Donkey Anti-Mouse IgG (H+L) secondary antibody (Jackson ImmunoResearch) to detect expression of the antigens in HEK293T cells. The median fluorescence intensity (MFI) measured in FACS serves as a surrogate for antigen expression. Cells were acquired using a BD FACSCelesta 2 (BD) and data was analyzed with FlowJo V10.8 (BD). [0954] Percentage of exemplary gC2, gD2, and dE2 antigen immunogenic fragment expressing cells is shown in FIGS.14A, D, AND G. All antigen variants showed high transfection rate. Median fluorescence intensity (MFI) of the total HEK293T population depicted per antigen is shown in FIGS.14B, E AND H . Antigen variants showed comparable or enhanced expression (FIGS.14C, F, and I) as compared to RNA comprising SEQ ID NOs: 106 (construct 1600), 118 (construct 1601), or 123 (construct 1602), which combined have been shown in Example 5 to induce a potent immune response in animal subjects. As in Example 5, these results indicate that the modRNA constructs characterized in the present Example would produce a strong immune response. Example 7: Example Clinical Studies of Exemplary RNA Compositions [0955] The present Example describes certain clinical assessments that may be performed of certain
Figure imgf000418_0001
trivalent compositions described herein. [0956] In some embodiments, more than one different exemplary trivalent composition candidate may be assessed. In some such embodiments, different candidates may vary, for example in: (1) RNA platform (e.g., unmodified RNA, nucleoside-modified RNA, self-amplifying RNA (saRNA), trans- amplifying RNA); (2) encoded antigen – e.g.: - which HSV (HSV-1 and/or HSV-2) protein(s) utilized, - full length protein antigen vs portion vs plurality of portions vs fusion with one or more heterologous sequences (e.g., membrane tether, secretion, linker(s)), and/or - epitopes from different (and/or multiple) phases of HSV life cycle; (3) number of RNAs; (4) elements of RNA construct; - cap and/or cap-adjacent sequences, - 5’ UTR, - 3’ UTR, and/or - polyA tail; (5) lipid composition of LNP. [0957] In one particular embodiment, a candidate exemplary trivalent composition comprises three RNAs described herein: a first RNA encoding an exemplary HSV-2 gC antigen immunogenic fragment, a second RNA encoding an exemplary HSV-2 gD antigen immunogenic fragment, and a third RNA encoding an exemplary HSV-2 gE antigen immunogenic fragment. In this particular exemplary embodiment, composition candidates may be evaluated by intramuscular administration, for example, based on a dose-escalation scheme. Example 8: In vitro Testing of Further Polyribonucleotides Encoding Exemplary HSV-2 Antigens [0958] The present Example demonstrates high transfection rates in HEK293T cells of modRNAs encoding exemplary gC2 and gE2 immunogenic frag
Figure imgf000419_0001
as high expression levels in HEK293T cells of exemplary gC2, gD2, or gE2 immunogenic fragment variants. [0959] HEK293T cells were transfected with 0.2 μg/mL modRNA encoding exemplary gC2, gE2 or gD2 antigen immunogenic fragments using a commercial transfection reagent. In the present Example, the following modRNAs were transfected into HEK293T cells: x modRNAs comprising SEQ ID NO: 104 (construct 1597) or SEQ ID NO: 106 (construct 1600) encoding an exemplary gC2 antigen immunogenic fragment with an exemplary IL2 secretory signal; x modRNAs comprising SEQ ID NO: 116 (construct 1598), or SEQ ID NO: 118 (construct 1601) encoding an exemplary gD2 antigen immunogenic fragment with an exemplary gD2 secretory signal; or x modRNAs comprising SEQ ID NO: 121 (construct 1599) or SEQ ID NO: 123 (construct 1602) encoding an exemplary gE2 antigen immunogenic fragment with an exemplary IL2 secretory signal. [0960] After transfection, cells were incubated to express the modRNA. Afterwards, cells were harvested and probed with antigen-specific mouse monoclonal antibodies and a secondary fluorescence-tagged anti-mouse antibody to detect expression of composition (e.g., immunogenic composition, e.g., vaccine) antigens in HEK293T cells. The median fluorescence intensity (MFI) measured in FACS serves as a surrogate for antigen expression. [0961] As shown in FIG.15, gC and gE variants were expressed in HEK293T cells (FIG.15A and FIG. 15B). Exemplary gC immunogenic fragment, exemplary gD immunogenic fragment, and exemplary gE immunogenic fragment showed comparable or enhanced protein expression (C , D, and E respectively) compared to gD variant SEQ ID NO: 118 (construct 1601) or gE variant SEQ ID NO: 123 (construct 1602), which have been shown in Example 5 to induce a potent immune response in animal subjects. [0962] Cumulative total HEK293T expression data from up to n=7 (F) experiments are shown in relation to the exemplary gC2 immunogenic fragment (construct 1600, SEQ ID NO: 106), gD2 (construct 1601, SEQ ID NO: 118) or gE2 (construct 1602, SEQ ID NO: 123) constructs, respectively. [0963] As in Example 5, these results indicate that the modRNA constructs characterized in the present Example would produce a strong immune response. Example 9: In-vitro Expression of Example Polyribonucleotides Encoding Exemplary HSV gB Antigen Immunogenic Fragment Constructs [0964]
Figure imgf000419_0002
sent Example demonstrates that exemplary polyribonucleotides encoding different exempalry HSV gB antigen immunogenic fragment constructs, as described herein, can be assessed for in-vitro expression (e.g., intracellular, surface) in mammalian cells (HEK293T cells). [0965] In vitro expression assays are used to assess expression and localization of different exemplary HSV gB antigen immunogenic fragment constructs. Polynucleotides encoding various exemplary HSV gB antigen immunogenic fragment constructs provided herein are generated. Assays are initially performed with non-formulated DNA constructs to determine functionality. HEK293T cells are transfected with DNA constructs. HEK293T cells transfected with DNA constructs are assessed for protein expression by antibody staining and FACS. Expression from the DNA constructs is also assessed by Western blot and/or mass spectrometry. Antigenicity of polypeptides expressed from DNA constructs is assessed by antibody binding assays. Purified protein is also used to assess antigenicity. [0966] Based on results from assays using DNA constructs, corresponding RNA constructs are selected for further assessment. In vitro expression assays are used to assess expression and localization of polyribonucleotides encoding exemplary HSV gB antigen immunogenic fragment constructs. Polyribonucleotides encoding various exemplary HSV gB antigen immunogenic fragment constructs provided herein are generated. Assays are initially performed with non-formulated RNA constructs to determine functionality. Formulated RNA constructs are also assessed. HEK293T cells are transfected with (i) exemplary RNA constructs or (ii) exemplary LNP formulated RNA constructs. HEK293T cells transfected with RNA constructs or with formulated RNA constructs are assessed for protein expression by antibody staining and FACS. A transfection rate is determined by measuring percentage of positive cells, and total expression is determined by measuring median fluorescence of the total HEK population. HEK293T cells transfected with formulated RNA constructs are assessed for protein secretion by detecting protein in the culture supernatant of transfected cells. Example 10: Immunogenicity Studies of Example Polyribonucleotides Encoding Exemplary HSV gB antigen Immunogenic Fragment Constructs [0967] The present Example describes the ability of certain polyribonucleotides encoding exemplary HSV gB antigen immunogenic fragment constructs, provided by the present disclosure, to induce immune responses, as assessed in mice. [0968] C57BL6 female mice (10-12-weeks old) are immunized intramuscularly (IM) twice, generally on days 0 and 21, with a formulated RNA construct (described in Example 1) or injected with phosphate buffer saline (vehicle). Blood samples are collected pre-immunization (day 0) and after the first dose (on days 7, 14, 21, 28 and 35) to generate serum samples at various time points. At the end of the experiment (e.g., day 35), splenocytes are harvested and cryopreserved. Animals are divided into multiple groups receiving treatment as indicated in Table 27 below: Table 27: Study plan for certain example immunogenicity studies with RNA constructs. Group Treatment Example Dose D0 D7,14,21 D21 D28 D35
Figure imgf000420_0001
4 F3 1 μg Intramuscular (FBC) (IM) injection 5 F4 1 g
Figure imgf000421_0001
BC = Blood collection / Serum generation (amount of blood that can be collected: 14 days interval between collections: 150μl/20g mouse; 7 days interval between collections: 110μl/20g mouse) * pre-immune serum: blood sample is collected from 10% of all animals = 4 animals. FBC = Final blood collection/ Serum generation; Spleen, serum or cell collection (for, e.g., Fluorospot analysis) [0969] Serum samples obtained from each group of immunized animals are analyzed by one or more of the following method(s): (1) Enzyme-linked Immunosorbent Assay (ELISA), (2) multiplex assay, and/or (3) Fluorospot assay. (1) Enzyme-linked Immunosorbent Assay (ELISA) [0970] Provided polyribonucleotides can be assessed for their ability to induce production of antibodies that may bind to an exempalry HSV gB antigen or antigenic fragment thereof. In some embodiments, a provided polyribonucleotide is determined to induce a useful immune response if serum from a subject (e.g., a mouse) immunized with such construct is shown to bind an exemplary HSV gB antigen or antigenic fragment thereof. [0971] RNA constructs (as described in Example 1) are assessed for their ability to induce production of antibodies that bind to an HSV gB antigen or antigenic fragment thereof using an ELISA assay. [0972] Briefly, MaxiSorp 96-well plates are coated with 100 ng/well an exemplary HSV gB antigen or antigenic fragment thereof in coating buffer (50mM sodium carbonate, pH 9.6) and incubated overnight at 4°C, or 250 ng/well of an HSV gB antigen overlapping peptides in PBS and incubated 1h at 37°C. Plates are then blocked with 1% BSA in PBS for 1h at 37°C (for an HSV gB antigen or antigenic fragment thereof) or overnight at 4°C (for an HSV gB antigen or antigenic fragment thereof overlapping peptides). Bound IgG is detected using horseradish peroxidase (HRP)-conjugated goat anti-mouse IgG. Signal is detected after adding the substrate 3,3',5,5'- Tetramethylbenzidine (TMB) and 25% sulfuric acid to stop the reaction. Optical densities (OD) are read at 450 nm. [0973] Reciprocal end titers at day 35, after 2 immunizations, are used as a representative as they showed the highest antibody response. (2) Multiplex Assay [0974] Provided polyribonucleotides can be assessed for their ability to induce production of antibodies that bind to specificexemplary HSV gB antigens or antigenic fragments thereof. In some embodiments, a provided polyribonucleotide is determined to induce a useful immune response if serum from a subject (e.g., a mouse) immunized with such construct is shown to target peptides from an exemplary HSV gB antigen or antigenic fragment thereof in a multiplex assay, as described herein. [0975] RNA constructs (as described in Example 1) are assessed for their ability to induce production of antibodies that bind to specific exemplary HSV gB antigens or antigenic fragments thereof by performing a multiplex analysis (Meso Scale Discovery) according to the manufacturer’s instructions. Briefly, exemplary HSV gB antigens or immunogenic fragments thereof are conjugated with bovine serum albumin (BSA) and then bound to the wells of a 96-well plate, in a specific spot on the well. After incubation with serum from immunized mice, antibodies bound to each specific peptide are detected with a “Sulfo-Tag” conjugated secondary antibody. A multiplex reader instrument (MESO QuickPlex SQ 120) is used to quantify the light emitted from the Sulfo-Tag. (3) FluoroSpot Assay [0976] Provided polyribonucleotides can be assessed for their ability to induce production of antibodies responsive to an exemplary HSV gB antigen or immunogenic fragment thereof. In some embodiments, a polyribonucleotide is determined to induce a useful immune response if splenocytes from a subject (e.g., a mouse) immunized with such construct, following incubation with peptide(s) as described herein, exhibit T-cell secretion of one or more pro-inflammatory cytokines (e.g., IFN-γ, TNF-α, or IL-2) in a FluoroSpot Assay, as described herein. FluoroSpot assays are performed with mouse IFN-γ/IL-2/TNF-α FluoroSpotPLUS kit according to the manufacturer’s instructions (Mabtech). Example 11: Protection Studies of Example Polyribonucleotides Encoding Exemplary HSV gB Antigen Immunogenic Fragment Constructs [0977] The present Example documents the ability of certain polyribonucleotides, provided by the present disclosure, to induce immune responses, as assessed in mice. [0978] Provided polyribonucleotides can be assessed for their ability to protect a subject from HSV challenge. In some embodiments, a provided polyribonucleotide is determined to induce a useful immune response if a subject (e.g., a mouse) immunized with a polyribonucleotide and exposed to HSV demonstrate a reduced level of infection in a challenge assay as described herein. [0979] A challenge assay is performed in which C57BL/6 female mice (7-week-old, ~20 g at day 0) were immunized intramuscularly (IM) twice, on days 0 and 21, with 1 μg of a RNA construct (as described in Example 1) or injected with vehicle (n=7 mice/group). Blood samples are collected on days 7, 14, 20, 28, 35, 42, and 49 for analysis of antibody titers and functionality in serum. HSV challenge was performed at day 50. Protection against infection was assessed at experimental day 60 (day 10 after challenge). [0980] Antibody titers are assessed by ELISA. Pre-boost and pre-challenge samples (days 20 and 49) are also used in functionality assays. Example 12: Identification of Immunogenic T Cell Antigens ings, results of systematic analyses that produced
Figure imgf000422_0001
an unexpected list or set of T cell antigens (e.g., CD4 and/or CD8 T cell antigens) for HSV. T cell antigens of the present Examples are useful, individually or in combination, for the production of HSV antigen constructs (e.g., for use in HSV composition (e.g., immunogenic composition, e.g., vaccine) agents). The present Example provides, among other things, discussion of the systematic analysis applied, which included selection of source data and stepwise evaluations of source data to generate an unpredicted list of T cell antigens for HSV. [0982] A first step in the systematic identification of T cell antigens for HSV included analysis of publicly available datasets relating to T cell induction in HSV-infected individuals. T cell antigens for HSV preferably demonstrate immunogenicity as a target of T cells in HSV-infected human subjects, and still more preferably elicit T cell responses in a large proportion of infected human subjects in cohort studies. Many studies of T cell responses to HSV have been conducted. However, the present inventors appreciate that many studies are narrow (e.g., explored immune responses to one or a few antigens at a time in animal models or human subjects) or subject to bias. Comprehensive evaluation of every HSV gene with minimal bias is difficult to achieve by combining data from independent studies narrowly focused on a few targets. Moreover, systematic analysis of data from diverse studies is challenging due to use of unique methodologies, reagents and patient cohorts by different groups. The analysis of the present Example therefore included selection of studies that measured T cell responses (e.g., CD8 T cell responses) to the majority of HSV proteins simultaneously in multiple subjects. Three publications met these criteria and were further evaluated as sources of pan-genome immunogenicity data for HSV T cell targets: Hosken (J Virol 2006 Jun;80(11):5509-152006-15; PMID: 16699031; hereafter “Hosken” or “Hosken 2006”), Jing (J Clin Invest 2012 Feb;122(2):654-73; PMID: 22214845; hereafter “Jing” or “Jing 2012”), and Long (Virology 2014 Sep;464-465:296- 311; PMID: 25108380; hereafter “Long” or “Long 2014”). [0983] Having identified the above sources of pan-genome immunogenicity data for HSV T cell targets, the next step was to determine how to weight the three studies. To do so, we first determined if the reported frequencies of individuals with T cell responses (percent responders) to each antigen correlated between studies. We assumed that broad trends in immunogenicity should correspond between studies, and a lack of correlation would suggest possible methodological deficits. A strong correlation (r = 0.70, Spearman correlation) was observed between the percent responders reported in Jing 2012 and Hosken 2006, but no significant correlation was observed between the results in Long 2014 and either other dataset. We therefore deemed Long 2014 to be a low reliability dataset and excluded it from further analysis. [0984] Selection of T cell antigens for HSV based on the studies of Hosken 2006 and Jing 2012, from the set of proteins encoded by genes of the HSV genome, was undertaken in two steps. Each step was designed to reflect the form of data available in the respective publications. First, all HSV proteins below the median T cell percent responder frequency in Hosken 2006 were eliminated from consideration as T cell antigens (except that, as certain HSV genes were not represented in the data from Hosken 2006, genes for which no data were available were not eliminated from consideration). Second, genes for which fewer than 3/7 subjects had measurable T cell responses in Jing 2012 were eliminated from consideration as T cell antigens. A total of 14 genes remained after these elimination steps, which were further refined based on expression level as described below. [0985] A further analysis was conducted to identify candidate T cell antigens characterized by expression above a median level. In this further analysis, RNA and protein expression studies were evaluated to identify HSV genes encoding the most highly expressed T cell antigens during early viral replication. For this analysis, study data were derived from multiple sources. [0986] Some of the data sources comprised scRNA-Seq data. For these studies, the gene expression profiles of individual cells were analyzed, and by quantifying the expression of well-characterized immediate early (IE) genes (US1, UL54, UL50, UL23, UL30) and late genes (UL48, UL45, UL44, US11, UL36), the cells were placed in an approximate temporal ordering from early activation too late. Specifically, expression of late genes was averaged (termed as ‘late gene score’) for each single cell, and cells were arranged in order of increasing late gene score. Cells were then divided into 5 equal bins, and expression profiles of cells belonging to the same bin were averaged to generate a 5 distinct expression profiles representing different timepoints along the activation continuum. [0987] All expression profiles (from bulk RNA-Seq, scRNA-Seq, and proteomics) were normalized by scaling each sample/stage to have a maximum expression of 1 and minimum expression of 0. The resulting normalized profiles were used for visualizing overall expression and calculating Spearman’s correlation coefficient between pairs of expression profiles. The expression profiles were also sorted according to the expression of well- characterized immediate early (IE) genes (US1, UL54, UL50, UL23, UL30) and late genes (UL48, UL45, UL44, US11, UL36) to help determine which profiles more likely represented early stages of activation. Specifically, expression of late genes was averaged (termed as ‘late gene score’) for each single cell and cells were arranged in order of increasing late gene score (in precisely the manner used to order singles cells in the scRNA-Seq data analysis). By jointly analyzing the correlation coefficients and the “late gene score”, we selected 12 expression profiles from 11 studies that appeared to reliably represent expression during early activation. For each gene, the median expression level was determined across the 12 studies. Finally, across all genes, median expression was calculated, and genes with expression above the median were considered for vaccine inclusion. Based upon these analyses, a set of 10 T cell antigens (e.g., CD4 and/or CD8 T cell antigens) for HSV were identified. [0988] Source literature is listed below with brief comments on methodology: x Fox (mBio.2017 May-Jun; 8(3): e00745-17; PMID: 28611249) o Data relating to HSV1 KOS in MRC5 fibroblasts, MOI of 5, from an expression table containing raw reads per gene (supplementary files in GSE109420) was normalized by number of reads per sample and by gene length. We took 4hpi sample for downstream analysis. x Krenn (Cell Stem Cell.2021 Aug 5;28(8):1362-1379.e7; PMID: 33838105) o Data relate to RNA-seq on HSV1/mock infected hIPSC-derived 3D-brain organoid with or without IFNa2 treatment (which rescues HSV1 infection). Condition-specific sample subsets (designated by their SRR IDs and condition labels in the sheet) were acquired from GEO (accession ID: GSE145496). These samples were aligned to the concatenated GRCh38 and HSV-1 strain F genome using the STAR alignment (--runMode alignReads --outSAMattributes All --outSAMtype BAM-- outMultimapperOrder Random). Reads overlapping HSV-1 genes were then aggregated using HTSeq package and HSV1 strain F reference annotation (HTSeq parameters: -t CDS -i gene). Gene-specific feature counts were TPM-normalized. x Wang (Virol J.2020 Jul 8;17(1):95; PMID: 32641145) o Data relate to Bulk RNA-seq on trigeminal ganglia of mouse and tree shrew, with HSV-1. Aligned reads from HSV-1 in BAM format were acquired from BIG Data center (accession number CRA001750) and quantified using featureCount function from Rsubread R package. Reads were normalized by gene length. We took 7dpi time point samples for mouse and tree shrew. x Walter (2021. Herpesviral induction of germline transcription factor DUX4 is critical for viral gene expression. bioRxiv doi: 10.1101/2021.03.24.436599) o Data relate to RNA-seq from HSV-1 infected HET293T cells with and without DUX4 KO. HSV-1 gene-level counts across the WT and DUX4KO conditions at 18hpi were downloaded using the GEO (GEO accession: GSE174759). Gene RS1-r had two rows with similar counts profile across the samples and was de-duplicated. The counts were then normalized by the total library size for the sample and scaled by 10^4. We took 18hpi time point for downstream analysis. x Tokuyama (2021. Endogenous retroviruses mediate IFN-independent protection against HSV-2 infection. Query DataSets for GSE185281 available at NCBI Gene Expression Ominibus (GEO)) o Data relate to RNA-seq from HSV-2 infected mice (vaginal samples). Raw read counts for two WT B6 mice infected with HSV-2 were downloaded from SRA archive under PRJNA768446. Reads were aligned to a concatenated human (hg38) and HSV-2 (HGS2) genome using the STAR software, and HSV-2 gene expression was quantified using htseq-count. Data is shown as average of log2(counts/total library size*1E4 +1) values from 2 replicates. x Khoury-Hanold (Cell Host Microbe.2016 Jun 8;19(6):788-99; PMID: 27281569) o Data relate to bulk RNA-seq from in vivo HSV-1 infection: vaginal route, DRG (dorsal root ganglia) and LIM (large intestinal musculara) sequences. Processed and normalized expression table was acquired from GSE74215. For downstream analysis, expression was additionally scaled to account for gene length bias. x Tombácz (Front Microbiol.2017 Jun 20;8:1079; PMID: 28676792) o Data relate to Pacbio seq of kidney epithelial cells infected with HSV1 (MOI=1). Expression data was extracted from Supplementary Files in the GEO record GSE97785 and gene names were unified using NCBI nomenclature. Data is in the form of percentage of HSV2 gene reads in each sample, and those mapping to the same gene are aggregated. x Wyler (Nature Communications volume 10, Article number: 4878 (2019); PMID: 31653857) o Data relate to scRNA-seq (host + viral) of HSV-1 infected fibroblasts GSE123782. Read counts mapping to host and viral genes were obtained from supplementary files in the publication. To generate expression profiles for samples from each time point, reads from cells belonging to the same time point were aggregated and transcripts per million (TPM) values are calculated for each HSV-1 gene. x Drayman (Elife.2019 May 15;8:e46339. doi: 10.7554/eLife.46339; PMID 31090537) o Data relate to scRNA-seq from HSV-1 infected fibroblasts, MOI2, 5hpi. Read counts were obtained from GSE126042. Reads were normalized by the depth of each sample, multiplied by 10000 and log2-normalized. Only cells with more than 50 reads aligned to HSV genome were considered HSV1+ and used for the analysis. x Kulej (Mol Cell Proteomics .2017 Apr;16(4 suppl 1):S92-S107; PMID: 28179408) o Proteomics expression data generated from primary human foreskin fibroblast (HFF) cells infected with HSV-1 strain 17 syn+ was extracted from Supplementary Table S2A from Kulej et al., MCP 2017;16(4 suppl 1):S92-S107, doi:10.1074/mcp.M116.065987 . We took 6hpi timepoint for downstream analysis x Soh (Cell Rep .2020 Oct 6;33(1):108235; PMID: 33027661) o Proteomics expression data generated from human keratinocyte cell line (HaCaT) cells infected with HSV-1 KOS strain was extracted from Supplementary Table S1 from Soh et al., Cell Reports 2020;33 (1), doi.org/10.1016/j.celrep.2020.108235. We took4hpi timepoint for downstream analysis. x Saiz-Ros (Cells .2019 Feb 3;8(2):120; PMID: 30717447) o Proteomics expression data generated from HeLa cells infected with HSV-1 strain 17 + was extracted from Table S1 from Saiz-Ros et al., Cells. 2019 Feb 3;8(2):120, doi: 10.3390/cells8020120. We took 8hpi timepoint for downstream analysis. [0989] Additional T cell antigens for HSV were identified from inspection of source literature, which additional antigens could be used individually or in combination with others described herein. One group of additional antigens was selected from subunit composition (e.g., immunogenic composition, e.g., vaccine)s that have progressed to clinical trials (Krishnan and Stuart. Front Microbiol. 2021 Dec 7;12:798927; PMID: 34950127). Another group of additional antigens was selected from promising prophylactic vaccination data in small animal models of HSV-2 infection (Morello et al. J Virol .2011 Apr;85(7):3461-72; PMID: 21270160; Platt et al. Cells. 2013 Jan 4;2(1):19-42; PMID: 24709642). A third group of additional antigens that were identified as candidate T cell antigens for HSV from Hosken 2006 and Jing 2012 (see above in this Example) but did not meet the expression threshold discussed above. These analyses inspecting source literature produced a set of 9 additional T cell antigens (e.g., CD4 and/or CD8 T cell antigens) for HSV. Example 13: Exemplary RNA Constructs Encoding HSV Antigens SV antigens, and sequences encoding them,
Figure imgf000426_0001
t at may be ut zed n certan embodments o t e present dscosure. Exemplary HSV (e.g., HSV-1 and/or HSV-2) antigens can be found in Tables 7-8. [0991] In some particular embodiments, an administered RNA has a structure: Structure 1: m2 7,3’-OGppp(m1 2’-O)ApG-hAg-Kozak-SEC-Immunogen -FI-A30L70, wherein m2 7,3’-OGppp(m1 2’-O)ApG = 5’ cap; hAg = 5’ UTR human alpha-globin; SEC = signal peptide (SP); Immunogen = a nucleotide sequence comprising a sequence that encodes an antigen described herein; FI = a 3’-UTR that is or comprises a sequence (e.g., 3’ UTR) from the “amino terminal enhancer of split” (AES) messenger RNA and a sequence (e.g., a non-coding region) from the mitochondrial encoded 12S ribosomal RNA (MT-RNR1); and A30L70 = a polyA sequence comprising 30 adenine nucleotides followed by 70 adenine nucleotides, wherein the 30 adenine nucleotides and 70 adenine nucleotides are separated by a linker sequence. [0992] In some embodiments, an administered RNA has a structure: Structure 2: m2 7,3’-OGppp(m1 2’-O)ApG-hAg-Kozak-SEC-Immunogen–MITD-FI-A30L70, wherein m2 7,3’-OGppp(m1 2’-O)ApG = 5’ cap; hAg = 5’ UTR human alpha-globin; SEC = signal peptide (SP); Immunogen = a nucleotide sequence comprising a sequence that encodes one or more antigens described herein; MITD = MHC Class I trafficking signal (MITD); FI = a 3’-UTR that is or comprises a sequence (e.g., 3’ UTR) from the “amino terminal enhancer of split” (AES) messenger RNA and a sequence (e.g., a non-coding region) from the mitochondrial encoded 12S ribosomal RNA (MT-RNR1); and A30L70 = a polyA sequence comprising 30 adenine nucleotides followed by 70 adenine nucleotides, wherein the 30 adenine nucleotides and 70 adenine nucleotides are separated by a linker sequence. [0993] In some embodiments, an Immunogen sequence encodes a plurality of immunogenic fragments (e.g., comprising epitopes) from an antigen. In some embodiments, an Immunogen sequence encodes a plurality of immunogenic fragments (e.g., epitopes) from two or more antigens. In some embodiments. In some embodiments, such immunogenic fragments are linked together to form an Immunogen sequence by linkers (e.g., in some embodiments a linker that is enriched in G and/or S amino acid residues). In some embodiments, a linker may be or comprise an amino acid sequence of GGSGGGGSGG (SEQ ID NO: 165). In some embodiments, a linker may be or comprise an amino acid sequence of GGSLGGGGSG (SEQ ID NO: 166). Example 14: Exemplary RNA Constructs Encoding Multiepitope HSV Antigens [0994] The present Example describes certain exemplary HSV multiepitope antigens, and sequences encoding them, that may be utilized in certain embodiments of the present disclosure. [0995] Exemplary Construct Encoding an HSV multi-epitope polypeptide: Structure m2 7,3’-OGppp(m1 2’-O)ApG-hAg-Kozak-SEC-CD8 string- MITD-FI-A30L70 [0996] In some embodiments, a T cell antigen string can include at least 2 (including, e.g., at least 1, at least 2, at least 3, at least 4, at least 5, at least 10, at least 15, at least 20, or more) T cell epitopes and/or HLA-I epitopes as listed in Tables 7-8. Example 15: Exemplary Prediction and/or Characterization of MHC Presentation HC presentation which may
Figure imgf000427_0001
be used in accordance with the present disclosure to select and/or characterize antigenic peptides as described herein. [0998] In some embodiments, an antigenic peptide is selected and/or characterized through analysis of its amino acid sequence using an MHC-peptide presentation prediction algorithm or MHC-peptide presentation predictor, for example implemented in a computer processor (e.g., a computer processor that has been trained by a machine learning software), which determines a likelihood of binding and presentation of an peptide by an MHC class I or an MHC class II antigen. [0999] In some embodiments, an MHC-peptide presentation prediction algorithm or MHC-peptide presentation predictor is or comprises neonmhc 1 and/or neonmhc2, which predict and/or characterize likelihood of MHC class I and MHC class II binding, respectively. Alternatively or additionally, in some embodiments, an MHC- peptide presentation prediction algorithm or MHC-peptide presentation predictor is or comprises NetMHCpan or NetMHCIIpan. In some embodiments, a hidden Markov model approach may be utilized for MHC-peptide presentation prediction and/or characterization. In some embodiments, the peptide prediction model MARIA may be utilized. In some embodiments, NetMHCpan is not utilized to predict or characterize likelihood of MHC binding for peptides as described herein. In some embodiments, the peptide prediction model MARIA may be utilized. In some embodiments, NetMHCIIpan is not utilized to predict or characterize likelihood of MHC binding for peptides as described herein. In some embodiments, neither NetMHCpan nor NetMHCIIpan is utilized to predict or characterize likelihood of MHC binding for peptides as described herein. In some embodiments, an MHC-peptide presentation prediction algorithm or MHC-peptide presentation predictor is or comprises RECON, which offers high quality MHC- peptide presentation prediction based on expression, processing and binding capabilities. [1000] In some embodiments, multiple MHC-peptide presentation prediction algorithms or MHC-peptide presentation predictors may be utilized; in some such embodiments, results obtained with different strategies are compared with one another. In some embodiments, a determination that a particular peptide is likely to be or is significantly likely to be presented by MHC class I or MHC class II may be considered to be better established if two or more algorithms or predictors agree. [1001] Alternatively or additionally, identification and/or characterization of MHC binding (e.g., of MHC class I and/or MHC class II binding) may involve experimental assessment, or reports thereof, which may involve presentation in one or more in vitro systems and/or in one or more organisms. In some embodiments, such assessment utilizes mammalian cells or systems; in some embodiments such assessment utilizes primate (e.g., in some embodiments, human and/or in some embodiments, non-human primate) cells or systems. Example 16: Exemplary HLA Class I and Class II Binding Assays [1002] The present Example describes exemplary techniques for assessing peptide binding to HLA molecules. In some embodiments, exemplified technologies may determine and/or characterize (e.g., quantify) binding affinities for HLA class I and HLA class II. [1003] In general, binding assays can be performed with peptides that are either motif-bearing or not motif-bearing. A detailed description of an exemplary protocol that can be utilized to measure the binding of peptides to Class I and Class II MHC has been published (Sette et al., Mol. Immunol. 31:813, 1994; Sidney et al., in Current Protocols in Immunology, Margulies, Ed., John Wiley & Sons, New York, Section 18.3, 1998). Briefly, purified MHC molecules (5 to 500nM) are incubated with various unlabeled peptide inhibitors and 1-10nM 1251-radiolabeled probe peptides for 48h in PBS containing 0.05% Nonidet P40 (NP40) (or 20% w/v digitonin for H-2 IA assays) in the presence of a protease inhibitor cocktail. Assays are typically performed at pH 7.0, though in some embodiments a lower pH (typically above about pH 4.0) may be performed. [1004] Following incubation, MHC-peptide complexes are separated from free peptide, for example by gel filtration, e.g., on 7.8 mm x 15 cm TSK200 columns (TosoHaas 16215, Montgomeryville, PA), though those skilled in the art will appreciate that column size can be adjusted, if desired, for example to improve separation of bound vs unbound peptides of a particular size or characteristic. The eluate from the TSK columns is passed through a Beckman 170 radioisotope detector, and radioactivity is plotted and integrated using a Hewlett-Packard 3396A integrator, and the fraction of peptide bound is determined. [1005] Radiolabeled peptides can be iodinated using the chloramine-T method. Typically, in preliminary experiments, each MHC preparation is titered in the presence of fixed amounts of radiolabeled peptides to determine the concentration of HLA molecules necessary to bind 10-20% of the total radioactivity. Subsequent inhibition and direct binding assays can be performed using these HLA concentrations. [1006] Since under these conditions [label]<[HLA] and IC50>[HLA], the measured IC50 values are often reasonable approximations of the true KD values. Peptide inhibitors are typically tested at concentrations ranging from 120 μg/ml to 1.2 ng/ml, and are tested in two to four completely independent experiments. To allow comparison of the data obtained in different experiments, a relative binding figure is typically calculated for each peptide by dividing the IC50 of a positive control for inhibition by the IC50 for each tested peptide (typically unlabeled versions of the radiolabeled probe peptide). For database purposes, and inter-experiment comparisons, relative binding values can be compiled. Such values can subsequently be converted back into IC50 nM values, for example by dividing the IC50 nM of the positive controls for inhibition by the relative binding of the peptide of interest. This method of data compilation has proven to provide accurate and consistent comparison for peptides that have been tested on different days, or with different lots of purified MHC. [1007] Alternatively or additionally, live cell/flow cytometry-based assays can also be used. This is a well- established assay utilizing the TAP-deficient hybridoma cell line T2 (American Type Culture Collection (ATCC Accession No. CRL-1992), Manassas, Va.). The TAP deficiency in this cell line leads to inefficient loading of MHCI in the ER and an excess of empty MHCIs. Salter and Cresswell, EMBO J. 5:94349 (1986); Salter, Immunogenetics 21:235-46 (1985). Empty MHCIs are highly unstable, and are therefore short-lived. When T2 cells are cultured at reduced temperatures, empty MHCIs appear transiently on the cell surface, where they can be stabilized by the exogenous addition of MHCI-binding peptides. To perform this binding assay, peptide-receptive MHCIs are induced by culturing aliquots of 107 T2 cells overnight at 26°C in serum free AIM-V medium alone, or in medium containing escalating concentrations (0.1 to 100 PM) of peptide. Cells are then washed twice with PBS, and subsequently incubated with a fluorescent tagged HLA allele-specific monoclonal antibody (e.g., HLA-A02:01-specific monoclonal antibody, BB7.2), to quantify cell surface expression. Samples are acquired on a FACS Calibur instrument (Becton Dickinson) and the mean fluorescence intensity (MFI) determined using the accompanying Cellquest software. Example 17: Confirmation of Immunogenicity [1008] The present Example describes an exemplary method for confirmation of immunogenicity, in particular by utilizing in vitro expansion (IVE) assays to test the ability of one or more exemplary antigens or peptides to expand CD8+ T cells. [1009] Mature professional APCs are prepared for these assays in the following way. 80-90x106 PBMCs from a healthy human donor are plated in 20 ml of RPMI media containing 2% human AB serum, and incubated at 37°C for 2 hours to allow for plastic adherence by monocytes. Non-adherent cells are removed and the adherent cells are cultured in RPMI, 2% human AB serum, 800 IU/ml of GM-CSF and 500 IU/ml of IL-4. After 6 days, TNF-alpha is added to a final concentration of 10 ng/ml. On day 7, the dendritic cells (DC) are matured either by the addition of 12.5 mg/ml poly I:C or 0.3 μg/ml of CD4OL. The mature dendritic cells (mDC) are harvested on day 8, washed, and either used directly or cryopreserved for future use. [1010] For the IVE of CD8+ T cells, aliquots of 2x105 mDCs are pulsed with each exemplary peptide at a final concentration of 100 micromole, incubated for 4 hours at 37°C, and then irradiated (2500 rads). The peptide- pulsed mDCs are washed twice in RPMI containing 2% human AB serum. 2x105 mDCs and 2x106 autologous CD8+ cells are plated per well of a 24-well plate in 2 ml of RPMI containing 2% human AB, 20 ng/ml IL-7 and 100 pg/ml of IL-12, and incubated for 12 days. The CD8+ T cells are then re-stimulated with peptide-pulsed, irradiated mDCs. Two to three days later, 20 IU/ml IL-2 and 20 ng/IL7 are added. Expanding CD8+ T cells are re-stimulated every 8- 10 days, and are maintained in media containing IL-2 and IL-7. Cultures are monitored for peptide-specific T cells using a combination of functional assays and/or tetramer staining. Parallel IVES with the modified and parent peptides allowed for comparisons of the relative efficiency with which the peptides expanded peptide-specific T cells. Example 18: Quantitative and Functional Assessment of CD8+ and CD4+ T cells Tetramer Staining [1011] MHC tetramers are purchased or manufactured on-site, and are used to measure peptide-specific T cell expansion in the IVE assays. For the assessment, tetramer is added to lx105 cells in PBS containing 1% FCS and 0.1% sodium azide (FACS buffer) according to manufacturer's instructions. Cells are incubated in the dark for 20 minutes at room temperature. Antibodies specific for T cell markers, such as CD8, are then added to a final concentration suggested by the manufacturer, and the cells are incubated in the dark at 4°C for 20 minutes. Cells are washed with cold FACS buffer and resuspended in buffer containing 1% formaldehyde. Cells are acquired on a FACS Calibur (Becton Dickinson) instrument, and are analyzed by use of Cellquest software (Becton Dickinson). For analysis of tetramer positive cells, the lymphocyte gate is taken from the forward and side-scatter plots. Data are reported as the percentage of cells that are CD8+/Tetramer+. [1012] CD4+ T cell responses towards exemplary antigens or peptides can be tested using the ex vivo induction protocol. In this example, CD4+ T cell responses are identified by monitoring IFNγ and/or TNFα production in an antigen specific manner. Evaluation of Antigen Presentation: [1013] For a subset of exemplary antigens or peptides (e.g., that are or comprise predicted or selected epitope(s) as described herein), affinity for the indicated HLA alleles and/or stability with the HLA alleles can be determined. [1014] An exemplary detailed description of a protocol that can be utilized to measure the binding affinity of peptides to Class I MHC has been published (Sette et al, Mol. Immunol. 31(11):813-22, 1994). In brief, MHCI complexes are prepared and bound to radiolabeled reference peptides. Peptides are incubated at varying concentrations with these complexes for 2 days, and the amount of remaining radiolabeled peptide bound to MHCI is measured using size exclusion gel-filtration. The lower the concentration of exemplary test peptide needed to displace the reference radiolabeled peptide demonstrates a stronger affinity of the peptide for MHCI. Peptides with affinities to MHCI <50nM are generally considered strong binders while those with affinities <150nM are considered intermediate binders and those <500nM are considered weak binders (Fritsch et al, 2014). [1015] An exemplary detailed description of a protocol that can be utilized to measure binding stability of peptides to Class I MHC has been published (Harndahl et al. J Immunol Methods. 374:5-12, 2011). Briefly, synthetic genes encoding biotinylated MHC-I heavy and light chains are expressed in E. coli and purified from inclusion bodies using standard methods. The light chain (β2m) is radio-labeled with iodine (125I), and combined with the purified MHC-I heavy chain and peptide of interest at 18°C to initiate pMHC-I complex formation. These reactions are carried out in streptavidin coated microplates to bind the biotinylated MHC-I heavy chains to the surface and allow measurement of radiolabeled light chain to monitor complex formation. Dissociation is initiated by addition of higher concentrations of unlabeled light-chain and incubation at 37°C. Stability is defined as the length of time in hours it takes for half of the complexes to dissociate, as measured by scintillation counts. MHC-II binding affinity with peptides is measured following the same general procedure as with measuring MHCI-peptide binding affinity. Prediction algorithms utilized for predicting MHCII alleles for binding to a given peptide are described herein. Alternatively or additionally, NetMHCIIpan may be utilized for prediction of binding. [1016] To assess whether particular exemplary antigens or peptides or epitopes could be processed and presented from a larger polypeptide context, peptides eluted from HLA (class I or class II) molecules isolated from cells expressing the genes of interest can be analyzed by tandem mass spectrometry (MS/MS). ELISPOT [1017] Peptide-specific T cells are functionally enumerated, for example, using the ELISPOT assay (BD Biosciences), which measures the release of IFNgamma from T cells on a single cell basis. Target cells (T2 or specific HLA-transfected C1Rs (e.g., HLA-A0201 transfected C1Rs)) are pulsed with 10 uM exemplary peptide for 1 hour at 37°C, and washed three times. 1x105 peptide-pulsed targets are co-cultured in the ELISPOT plate wells with varying concentrations of T cells (5x102 to 2x103) taken from the IVE culture. Plates are developed according to the manufacturer's protocol, and analyzed on an ELISPOT reader (Cellular Technology Ltd.) with accompanying software. Spots corresponding to the number of IFNgamma-producing T cells are reported as the absolute number of spots per number of T cells plated. T cells expanded on exemplary modified peptides are tested not only for their ability to recognize targets pulsed with the exemplary modified peptide, but also for their ability to recognize targets pulsed with the parent peptide. CD107 Staining [1018] CD107a and b are expressed on the cell surface of CD8+ T cells following activation with cognate peptide. The lytic granules of T cells have a lipid bilayer that contains lysosomal-associated membrane glycoproteins (“LAMPs”), which include the molecules CD107a and b. Without wishing to be bound by any one theory, it is proposed that, when cytotoxic T cells are activated through the T cell receptor, the membranes of these lytic granules mobilize and fuse with the plasma membrane of the T cell. The granule contents are released, and this leads to the death of the target cell. As the granule membrane fuses with the plasma membrane, C107a and b are exposed on the cell surface, and therefore are markers of degranulation. Because degranulation as measured by CD107 a and b staining is reported on a single cell basis, the exemplary assay is used to functionally enumerate peptide-specific T cells. To perform the assay, exemplary peptide is added to specific HLA-transfected cells C1Rs (e.g., HLA-A0201 transfected C1Rs) to a final concentration of 20 PM, the cells are incubated for 1 hour at 37°C, and washed three times. 1x105 of the exemplary peptide-pulsed C1R cells are aliquoted into tubes, and antibodies specific for CD107 a and b are added to a final concentration suggested by the manufacturer (Becton Dickinson). Antibodies are added prior to the addition of T cells in order to “capture” the CD107 molecules as they transiently appear on the surface during the course of the assay. 1x105 T cells from the culture are added next, and the samples are incubated for 4 hours at 37°C. The T cells are further stained for additional cell surface molecules such as CD8 and acquired on a FACS Calibur instrument (Becton Dickinson). Data is analyzed using the accompanying Cellquest software, and results are reported as the percentage of CD8+ CD107 a and b+ cells. CTL Lysis [1019] Cytotoxic activity can be measured, for example, using a chromium release assay. Target T2 cells are labeled for 1 hour at 37°C with Na51Cr and washed 5x103 target T2 cells are then added to varying numbers of T cells from the IVE culture. Chromium release is measured in supernatant harvested after 4 hours of incubation at 37°C. The percentage of specific lysis is calculated as: Experimental release-spontaneous release/Total release-spontaneous release x100. Example 19: Administration of Exemplary Polyepitopic Compositions [1020] The present Example describes exemplary administration of compositions that comprise or deliver a plurality of epitopes. [1021] For example, an exemplary polyepitopic composition (e.g., immunogenic composition, e.g., vaccine, e.g., that comprises or delivers a collection of epitopes – e.g., as individual discrete exemplary peptides or as one or more exemplary polyepitopic peptides such as one or more string constructs as described herein). [1022] In some embodiments, an exemplary polyepitopic composition (e.g., immunogenic composition, e.g., vaccine) comprises or delivers multiple exemplary cytotoxic T lymphocyte (CTL) and/or helper T lymphocyte (HTL) epitopes. In some embodiments, such a composition (e.g., immunogenic composition, e.g., vaccine) is administered to subject(s) at risk of or having experienced exposure to infection. [1023] In some embodiments, an exemplary polyepitopic composition (e.g., immunogenic composition, e.g., vaccine) as described herein comprises or delivers one or more polypeptides, each of which encompasses multiple exemplary epitopes. In some embodiments, one or more exemplary monoepitopic peptides or exemplary polyepitopic antigens is delivered to a subject by administration of one or more a nucleic acid (e.g., DNA or and RNA) constructs. In some embodiments, a single nucleic acid construct (e.g., a DNA or RNA encoding an exemplary polyepitopic antigen) is administered. In some embodiments, a plurality of nucleic acids (e.g., each encoding a different exemplary monoepitopic or exemplary polyepitopic antigens) is administered. In some embodiments, an administered nucleic acid is an RNA (e.g., an mRNA); in some embodiments, a nucleic acid (e.g., an RNA) is administered in an LNP composition. [1024] In some embodiments, an administered composition includes an aqueous carrier and/or alum. [1025] In some embodiments, an initial administration is followed by one or more booster doses. In some embodiments, a booster dose includes the same amount of an exemplary polyepitopic construct as the initial dose. In some embodiments, a booster dose include more or less of an exemplary polyepitopic construct than was provided in the initial dose. In some embodiments, a booster dose is administered after an interval of at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 12, 11, 12, 13, 14, 15, 16, 17, 18, 29, 20, 21, 22, 23, 24 weeks or more after an initial dose. In some embodiments, multiple booster doses are administered. In some embodiments, each subsequent booster is administered at an interval that is the same as or longer than that between its immediate predecessor dose and the dose before that. In some embodiments, 2, 3, 4, 5, 6, or more boosters are administered. In some embodiments, not more than 4 doses total are administered. In some embodiments, not more than 3 doses total are administered. In some embodiments, not more than two doses total are administered. In some embodiments, not more than 1, 2, 3 or 4 doses are administered within a particular 12 month period. In some embodiments, not more than 3, not more than 2, or not more than 1 dose(s) is/are administered within a particular 12 month period (e.g., within 12 months of the initial dose). [1026] In some embodiments, evaluation of an induced immune response (e.g., of magnitude, character, and/or diversity of immune response, such as antibody and/or T cell response) is performed before and/or after one or more doses (e.g., 1, 2, 3, 4, or more weeks after administration of a particular dose and/or within 6, 5, 4, 3, 2, or 1 month of administration of a particular dose) and may, for example be considered in determination of whether one or more booster doses should be administered and/or timing of such booster dose administration. In some embodiments, assessment of an immune response may utilize, for example, techniques that determine presence and/or level of epitope-specific CTL populations in a PBMC sample. Example 20: Administration of Dendritic Cells [1027] The present Example describes exemplary dendritic cell compositions that comprise or deliver exemplary antigens as described herein. [1028] In this Example, exemplary peptides comprising epitopes as described herein (e.g., identified, designed, selected and/or characterized as described herein) are loaded onto dendritic cells. Exemplary Peptide- pulsed dendritic cells can be administered to a subject. In some embodiments, such administration may stimulate a CTL response in vivo. [1029] In this Example, dendritic cells (e.g., autologous dendritic cells) are isolated, expanded, and pulsed with exemplary peptide CTL and/or HTL epitopes as described herein. Dendritic cells may then be infused back into the patient. Such infusion can elicit CTL and/or HTL responses in vivo. The induced CTL and HTL then destroy (CTL) or facilitate destruction (HTL) of target cells (e.g., liver cells) that bear the proteins from which the exemplary epitopes in the composition (e.g., immunogenic composition, e.g., vaccine) are derived. [1030] Ex vivo CTL or HTL responses to a particular exemplary antigen can be induced by incubating in tissue culture the patient's, or genetically compatible, CTL or HTL precursor cells together with a source of antigen- presenting cells, such as dendritic cells, and the appropriate immunogenic peptides. [1031] After an appropriate incubation time (typically about 7-28 days), in which the precursor cells are activated and expanded into effector cells, the cells are infused back into the patient, where they will destroy (CTL) or facilitate destruction (HTL) of their specific target cells, i.e., cells displaying relevant epitopes. Example 21: Administration of Epitope Binding Agents [1032] The present Example describes administration of epitope binding agents, as an alternative or complement to vaccination strategies describes herein. [1033] For example, among other things, the present disclosure provides technologies for identification and/or characterization of HSV antigens that are particularly amenable to targeting in order to disrupt one or more features of infection. In many embodiments described herein, the present disclosure provides technologies that involve administration or delivery of antigens that are or comprise such epitopes, for example, in order to induce an immune response targeting such epitope(s) in a recipient. [1034] Alternatively or additionally, the present disclosure provides and encompasses technologies for developing, characterizing, and/or administering agents that bind to such epitopes. In some embodiments, such strategies may provide or represent therapeutic interventions, for example useful in addition or as an alternative to vaccination strategies. [1035] Epitope binding agents, such as antibody agents, TCR agents, CAR agents, and/or cells expressing any of the foregoing can be administered in accordance with methodologies known in the art and/or described herein. [1036] In some embodiments, a relevant epitope binding agent can be delivered by administration of a composition that is or comprises the polypeptide binding agent. Alternatively or additionally, in some embodiments, a relevant epitope binding agent can be delivered by administration of a composition that is or comprises a polynucleotide (e.g., a DNA or RNA and, in many favored embodiments, an RNA) encoding the polypeptide binding agent. In some embodiments, a polypeptide binding agent is delivered by administered by a cell (or population thereof) that comprises or expresses the polypeptide and/or a polynucleotide that encodes it. Example 22: Exemplary identification and/or characterization of variant sequences with immunogenic potential [1037] The present Example describes technologies for identification and/or characterization of peptide sequences that differ from a relevant reference, for assessment of immunogenic potential. [1038] The full-length amino acid sequence of a variant protein (e.g., as observed in a circulating strain or developed through a predictive model) was derived. [1039] Constituent 9-mer and 10-mer peptide fragments of the variant protein are each scored for binding potential on common HLA alleles (including, e.g., but not limited to HLA-A01:01, HLA-A02:01. HLA-A03:01, HLA- A24:02, HLA-B07:02, and HLA-B08:01) using available algorithms. Peptides scoring better than 1000 nM are noted as potential candidates. [1040] Alternatively or additionally, constituent 9-mer or 10-mer peptide sequences not found in the reference protein sequence are flagged and scored for binding potential on common HLA alleles (including, e.g., but not limited to HLA-A01:01, HLA-A02:01. HLA-A03:01, HLA-A24:02, HLA-B07:02, and HLA-B08:01) using available algorithms. Example 23: Exemplary HSV peptide string construct designs [1041] The present Example exemplifies certain constructs (referred to herein as “strings”) of multiple exemplary HSV antigens or immunogenic fragments thereof and/or epitopes present in, and/or linked to one another in, compositions (e.g., immunogenic compositions, e.g., vaccines) or otherwise as described herein. [1042] Strings described in the present Example are designed to contain specific exemplary epitopes of HSV, each of which is disclosed herein and, e.g., is predicted and/or selected as described here, for example through use of an MHC-binding algorithm as described herein. The strings presented in the present Example are designed for therapeutic use in preventing and/or treating HSV infection and can be administered as polynucleotide constructs, e.g., mRNA encapsulated in a lipid nanoparticle. [1043] In some embodiments, strings described throughout this disclosure are encoded in an RNA that includes a 5’-UTR and 3’-UTR. Epitopes are interconnected by peptide linkers, encoded by their respective polynucleotide sequences. In some embodiments, one or more linkers may have a specific cleavage site. Example 24: Exemplary Antigen Identification, Selection and/or Characterization [1044] The present Example describes identification, selection and/or characterization of certain
Figure imgf000435_0001
V protein sequences useful as or in (i.e., as part of) exemplary antigens or immunogenic fragments thereof as described herein (see Example 1). [1045] Degree of conservation of candidate proteins across relevant HSV strains (e.g., in relevant geographic region) can be considered, e.g., in selection of sequences and/or epitopes to include in, or encode in, a composition (e.g., immunogenic composition, e.g., vaccine) and/or antigen construct. Various lab and field isolate strains can be considered for assessing conserved proteins and T cell epitopes. To evaluate conservation of HSV genes identified as a T cell epitopes for HSV, HSV1 and HSV2 genomes were downloaded from VIPR database, using “Genome search -> Herpesviridae family -> Alphaherpesvirinae subfamily -> Simplexvirus Genus -> Human Alphaherpesvirus 1 & 2 species”, complete genomes only. All downloaded genomes were checked to have consistent gene names. HSV-1 strain 17 and HSV-2 strain HG52 were used as reference strains for HSV-1 and HSV-2 respectively. Multiple sequence alignment (MSA) was performed for all sequences belonging to each of the HSV proteins using the mafft-linsi program with default parameters. Two measurements of conservation were computed: 1) protein-level conservation was computed as the percentage of sequence similarity to the reference strains (HSV-1 strain 17 and HSV-2 strain HG52) according to the MSA profiles, and 2) conservation at each position along each protein was quantified as the frequency of the predominant 9-mer that starts at each amino acid position. [1046] Immunogenicity of conserved proteins can also be considered, for example by review of literature and/or application of predictive algorithms as described herein. [1047] In accordance with the present Example, in some embodiments, an exemplary antigen may be or comprise one or more, and specifically may comprise a plurality, of distinct fragments (e.g., epitope-containing fragments) of one or more of these proteins, for example in a string construct as described herein. [1048] In some embodiments, a secretory signal (“Sec”) or a signal peptide (SP) domain present in exemplary string candidates described herein (e.g., a CD8 T cell antigen string and/or a CD4 T cell antigen string as described herein) may be that from HSV-2 gD SP MGRLTSGVGTAALLVVAVGLRVVCA (SEQ ID NO: 29); in alternative embodiments, a different secretory signal, e.g., from HSV-1 gD SP, is used. In some embodiments, a secretory signal peptide may be or comprise an IgE signal peptide. In some embodiments, a secretory signal peptide may be or comprise an IgE HC (Ig heavy chain epsilon -1) secretory signal peptide. In some embodiments, a secretory signal peptide that may be useful in accordance with the present disclosure may comprise one of the following sequences: MDSKGSSQKGSRLLLLLVVSNLLLPQGVVG (SEQ ID NO: 405); MDWTWILFLVAAATRVHS (SEQ ID NO: 391); METPAQLLFLLLLWLPDTTG (SEQ ID NO:390); MLGSNSGQRVVFTILLLLVAPAYS (Japanese encephalitis PRM secretory signal; SEQ ID NO: 392); MKCLLYLAFLFIGVNCA (VSVg protein secretory signal; SEQ ID NO: 393); MWLVSLAIVTACAGA (Japanese encephalitis JEV secretory signal; SEQ ID NO: 406); or MFVFLVLLPLVSSQC (SEQ ID NO: 408). [1049] In some embodiments, certain chunk boundary considerations are incorporated into the string constructs, for example establishing chunk boundaries to minimize presence of sequences (e.g., epitopes) that may overlap with the human proteome. Example 25: Exemplary Composition [1050] The present Example describes certain exemplary compositions (e.g., immunogenic compositions, e.g., vaccines). [1051] In some embodiments, a provided composition (e.g., immunogenic composition, e.g., vaccine) candidate will contain at least 2 RNAs, at least one of which encodes an exemplary HSV antigen or immunogenic fragment thereof described herein (e.g., a full length HSV protein or one or more immunogenic fragments or epitopes thereof, such as a string construct described herein), and optionally at least one of which encodes at least one other conserved exemplary HSV protein (or immunogenic fragment(s) or epitope(s) thereof, such as in a string construct as described herein; in some embodiments such a string construct may include fragments or epitopes from two or more different exemplary HSV proteins). [1052] In some embodiments, two or more (e.g., 3, for example 2 of which are/encode HSV antigen string constructs and one of which is/encodes a string construct of a plurality of CD8 and/or CD4 epitopes from other conserved HSV proteins) are formulated together in a single exemplary LNP formulation; in other embodiments, individual RNAs may be separately formulated in (the same or different) exemplary LNP formulations and such may be mixed together (e.g., in a 1:1 ratio of each RNA, or alternatively in a 1:1 ratio of HSV-antigen-encoding-RNA to “other” RNA so that, for example, for a composition comprising 2 HSV-antigen-encoding RNAs and one other RNA, the ratios would be 0.5:0.5:1). Example 26: Exemplary LNP Formulations [1053] The present Example describes certain preferred exemplary LNP formulations useful for compositions (e.g., immunogenic compositions, e.g., vaccines) as described herein. [1054] In some embodiments, exemplary LNP formulations that are useful for compositions (e.g., immunogenic compositions, e.g., vaccines) as described herein can comprise at least one ionizable aminolipid. In some embodiments, exemplary LNP formulations that are useful for compositions (e.g., immunogenic compositions, e.g., vaccines) as described herein can further comprise a helper lipid, which in some embodiments may be or comprise a neutral helper lipid. In some embodiments, exemplary LNP formulations that are useful for compositions (e.g., immunogenic compositions, e.g., vaccines) as described herein can further comprise a polymer-conjugated lipid, for example in some embodiments PEG-conjugated lipids. In some embodiments, exemplary LNP formulations that are useful for compositions (e.g., immunogenic compositions, e.g., vaccines) as described herein can comprise at least one ionizable aminolipid, at least one helper lipid (e.g., a neutral helper lipid, which in some embodiments may comprise a phospholipid, a steroid, or combinations thereof), and at least one polymer-conjugated lipid (e.g., PEG-conjugated lipid). In some embodiments, an exemplary LNP formulation may comprise an ionizable aminolipid, a phospholipid, a steroid, and a PEG-conjugated lipid. [1055] In some embodiments, an ionizable aminolipid may be present in an exemplary LNP formulation within a range of 45 to 55 mol percent, 40 to 50 mol percent, 41 to 49 mol percent, 41 to 48 mol percent, 42 to 48 mol percent, 43 to 48 mol percent, 44 to 48 mol percent of total lipids. In some embodiments, an exemplary ionizable aminolipid is or comprises ((4-hydroxybutyl)azanediyl)bis(hexane-6,1-diyl)bis(2-hexyldecanoate) (also known as 6-[N-6-(2-hexyldecanoyloxy)hexyl-N-(4-hydroxybutyl)amino]hexyl 2-hexyldecanoate). In some embodiments, an exemplary ionizable aminolipid is or comprises SM-102 (heptadecan-9-yl 8 ((2 hydroxyethyl)(6 oxo 6-(undecyloxy)hexyl)amino)octanoate) or an aminolipid as described in Sabnis et al. “ A Novel Amino Lipid Series for mRNA Delivery: Improved Endosomal Escape and Sustained Pharmacology and Safety in Non-human Primates” Mol. Ther. (2018) 26:1509-1519. In some embodiments, an exemplary ionizable aminolipid is or comprises an ionizable aminolipid as disclosed in US2020/0163878 or WO2018/078053, the entire contents of each of which are incorporated herein by reference for the purposes described herein. [1056] In some embodiments, a phospholipid may be present in an exemplary LNP formulation within a range of 5 to 15 mol percent, 7 to 13 mol percent, or 9 to 11 mol percent of total lipids. In some embodiments, an exemplary phospholipid is or comprises 1,2-Distearoyl-sn-glycero- 3-phosphocholine (DSPC). [1057] In some embodiments, a sterool may be present in an exemplary LNP formulation within a range of 30 to 50 mol percent, 35 to 45 mol percent or 38 to 43 mol percent of total lipids. In some embodiments, an exemplary sterol is or comprises cholesterol. [1058] In some embodiments, a polymer conjugated lipid (e.g., PEG-conjugated lipid) may be present in an exemplary LNP formulation within a range of 1 to 10 mol percent, 1 to 5 mol percent, or 1 to 2.5 mol percent of total lipids. In some embodiments, an exemplary PEG-conjugated lipid is or comprises 2-[(polyethylene glycol)-2000]- N,N-ditetradecylacetamide (also known as 2-[2-(ω-methoxy (polyethyleneglycol2000) ethoxy]-N,N- ditetradecylacetamide). In some embodiments, an exemplary phospholipid is or comprises PEG2000-DMG (1- monomethoxypolyethyleneglycol-2,3- dimyristylglycerol with polyethylene glycol of average molecular weight 2000). In some embodiments, an exemplary PEG-conjugated lipid is or comprises a PEG-lipid as disclosed in US2020/0163878 or WO2018/078053, the entire contents of each of which are incorporated herein by reference for the purposes described herein. [1059] In some embodiments, an exemplary LNP formulation comprises (i) an ionizable aminolipid within a range of 45 to 55 mol percent of total lipids; (ii) a phospholipid within a range of 8 to 12 mol percent of total lipids; (iii) a steroid within a range of 35 to 45 mol percent of total lipids; and (iv) a polymer conjugated (e.g., PEG- conjugated polymer) within a range of 1 to 2 mol percent of total lipids; and RNA molecules as described herein that are encapsulated within or associated with the lipid nanoparticles. [1060] In some embodiments, an exemplary LNP formulation comprises (i) ionizable amino lipid within a range of 45 to 55 mol percent of total lipids; (ii) DSPC within a range of 5 to 15 mol percent of total lipids; (iii) cholesterol within a range of 35 to 45 mol percent of total lipids; and (iv) a PEG-conjugated lipid within a range of 1 to 2 mol percent of total lipids; and RNA molecules as described herein that are encapsulated within or associated with the lipid nanoparticles. [1061] In some embodiments, an exemplary LNP formulation comprises (i) an ionizable aminolipid within a range of 40 to 50 mol percent of total lipids; (ii) a phospholipid within a range of 5 to 15 mol percent of total lipids; (iii) a steroid within a range of 35 to 45 mol percent of total lipids; and (iv) a polymer conjugated (e.g., PEG- conjugated polymer) within a range of 1 to 10 mol percent of total lipids; and RNA molecules as described herein that are encapsulated within or associated with the lipid nanoparticles. In some such embodiments, an ionizable aminolipid is or comprises ((4-hydroxybutyl)azanediyl)bis(hexane-6,1-diyl)bis(2-hexyldecanoate) (also known as 6- [N-6-(2-hexyldecanoyloxy)hexyl-N-(4-hydroxybutyl)amino]hexyl 2-hexyldecanoate). In some such embodiments, a phospholipid is or comprises 1,2-Distearoyl-sn-glycero-3-phosphocholine (DSPC). In some such embodiments, a steroid is or comprises cholesterol. In some such embodiments, a polymer conjugated polymer is or comprises 2- [(polyethylene glycol)-2000]-N,N-ditetradecylacetamide (also known as 2-[2-(ω-methoxy (polyethyleneglycol2000) ethoxy]-N,N-ditetradecylacetamide). [1062] In one embodiment, an exemplary LNP formulation comprises the following lipids included in Table 28 below and RNA molecules as described herein. Table 28: Exemplary LNP Formulation Lipids Proportion (mol%)
Figure imgf000438_0001
[1063] In some embodiments, an exemplary LNP formulation comprises an ionizable aminolipid, DSPC, cholesterol, and PEG-conjugated lipid at a molar ratio of approximately 50:10:38.5:1.5 or 47.5:10:40.8:1.7. In some embodiments, an ionizable amino lipid is or comprises ((4-hydroxybutyl)azanediyl)bis(hexane-6,1-diyl)bis(2- hexyldecanoate) (also known as 6-[N-6-(2-hexyldecanoyloxy)hexyl-N-(4-hydroxybutyl)amino]hexyl 2- hexyldecanoate). [1064] In some embodiments, an exemplary LNP formulation comprises (i) SM-102 (heptadecan-9-yl 8 ((2 hydroxyethyl)(6 oxo 6-(undecyloxy)hexyl)amino)octanoate) within a range of 45 to 55 mol percent of total lipids; (ii) DSPC within a range of 5 to 15 mol percent of total lipids; (iii) cholesterol within a range of 35 to 45 mol percent of total lipids; and (iv) PEG2000-DMG within a range of 1 to 2 mol percent of total lipids; and RNA molecules as described herein that are encapsulated within or associated with the lipid nanoparticles. [1065] In some embodiments, an exemplary LNP formulation comprises (i) ((4- hydroxybutyl)azanediyl)bis(hexane-6,1-diyl)bis(2-hexyldecanoate) (also known as 6-[N-6-(2-hexyldecanoyloxy)hexyl- N-(4-hydroxybutyl)amino]hexyl 2-hexyldecanoate) within a range of 45 to 55 mol percent of total lipids; (ii) DSPC within a range of 5 to 15 mol percent of total lipids; (iii) cholesterol within a range of 35 to 45 mol percent of total lipids; and (iv) a PEG-conjugated lipid within a range of 1 to 2 mol percent of total lipids; and RNA molecules as described herein that are encapsulated within or associated with the lipid nanoparticles. Example 27: Exemplary Pre-clinical assessment [1066] The present Example describes certain pre-clinical assessments that may be performed of certain exemplary RNA compositions (e.g., immunogenic compositions, e.g., vaccines) described herein: [1067] In some embodiments, one exemplary composition (e.g., immunogenic composition, e.g., vaccine) candidate is assessed. In some embodiments, more than one different exemplary composition (e.g., immunogenic composition, e.g., vaccine) candidate may be assessed. In some such embodiments, different candidates may vary, for example, in: RNA platform (e.g., unmodified RNA, modified RNA, saRNA); Encoded antigen(s); Number of RNAs; Elements of RNA construct (e.g., cap and/or cap-adjacent sequences, 5’-UTR, 3’-UTR, and/or PolyA tail); and/or Lipid composition of LNP. [1068] In some embodiments, pre-clinical assessment of certain RNA compositions (e.g., LNP formulated RNA-based HSV vaccines) comprises one or more of assessment in challenge experiments, assessment of level of protection, assessment of immunogenicity, and/or assessment of functional antibody responses. [1069] LNP formulated RNA-based HSV compositions are tested in a challenge model. Non-human primate models, such as Rhesus macaques and Cynomolgus monkey, and/or rodent models, such as C57/B16 mice, Balb/c mice or NODscidIL2Rγnull mice; and/or guinea pig models, inoculated with HSV, are administered a first vaccination and can be administered an additional vaccination (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 additional vaccinations) following the first vaccination. Wherein more than one vaccination is administered, the vaccinations are administered at an interval of 1, 2, 3, 4, 5, 6, 7, or 8 week intervals). Following vaccination, animals are challenged by HSV. Alternatively or additionally, animals are challenged by intravenous, subcutaneous, and/or intramuscular injection of virus-infected lymphocytes. Lymphocytes can be infected with any suitable strain of HSV. Animals are then evaluated for reduced infection of neurons. In some embodiments, an animal model is challenged in a plurality of instances (e.g., before first vaccination and/or wherein additional vaccinations are administered, at any time point between or after vaccinations). Following challenge, animals subjected to the study may be assessed according to any method known in the art, including, for example, serology assessment, immunogenicity, level of protection, etc. [1070] In some embodiments, serum antibody characterization and/or serum transfer experiments (e.g., from one vaccinated species to a different non-vaccinated species, e.g., from vaccinated non-human primate to non- vaccinated mouse) are conducted (e.g., to assess protective antibody response). [1071] In some embodiments, certain RNA compositions (e.g., immunogenic compositions, e.g., vaccines) of the present disclosure are assessed for level of protection. Level of protection can be assessed according to any suitable method known in the art. [1072] In some embodiments, certain RNA compositions (e.g., immunogenic compositions, e.g., vaccines) of the present disclosure are assessed for immunogenicity. For example, ELISA can be used to determine IgG specific (and subclasses thereof) titers and/or avidity of antibodies generated in response to certain exemplary RNA compositions (e.g., immunogenic compositions, e.g., vaccines) of the present disclosure to HSV antigens. In some embodiments, serum antibody titers against HSV glycoprotein (e.g., gH and/or gL glycoprotein, etc.) is determined by ELISA using standard methods. In some embodiments, for example, ELISpot (e.g., for CD8+, CD4+ T cells and/or IFNγ) and assessment of pro-inflammatory cytokine responses with splenocytes from immunized and/or challenged animal models and peptide pools derived from composition (e.g., immunogenic composition, e.g., vaccine) targets can also be assessed. In some embodiments, for example, phenotyping of immune responses (e.g., by flow cytometry) are assessed. In some embodiments, for example, T cell depletion and/or protection assays are conducted to assess immunogenicity (e.g., according to any suitable known method in the art). [1073] In some embodiments, one or more functional responses of antibodies generated in response to certain RNA compositions (e.g., immunogenic compositions, e.g., vaccines) of the present disclosure are assessed. Functional antibody responses can be assessed, for example, using an HSV neutralization assay. In some embodiments, an HSV in vitro neutralization assay is performed to evaluate one or more anti-HSV glycoprotein (e.g., HSV gB, gD gH, gL) antibodies in neutralizing HSV. For example, anti-HSV glycoprotein antibodies are obtained by collecting the sera of animals (e.g., mice) vaccinated with exemplary HSV RNA compositions. HSV virus are added to the diluted sera and neutralization is allowed to continue for 1 hour at room temperature. 3T3 cells are seeded in 96- wells one day before and the virus/serum mixtures are added to 3T3 monolayers. The cells are fixed on the next day and HSV-specific staining is performed. The plates are scanned and analyzed. A neutralization titer is expressed as the highest serum dilution required to achieve a 50% reduction in the number of plaques. [1074] In some embodiments, functional antibody responses can be assessed, for example, using passive transfer studies of sera from immunized animals to naïve animals that are challenged and assessing level of protection. Example 28: Exemplary Characterization Studies [1075] The present Example describes certain potential characterization studies that may be utilized, for
Figure imgf000441_0001
ize candidate compositions (e.g., immunogenic compositions, e.g., vaccines, e.g., manufacturing batches thereof), or components thereof as described herein. [1076] A immunization protocol can be utilized to assess ability of a composition (e.g., immunogenic composition, e.g., vaccine) candidate that comprises or delivers an antigen(s) as described herein to induce B- and/or T-cells, e.g., after intramuscular immunization, directed to the antigen(s) and/or epitope(s) thereof. In some embodiments, level and/or diversity of response is determined. In some embodiments, presence and/or level of neutralizing antibodies is/are determined. In some embodiments, protection of the immunized subject from challenge with HSV is assessed. [1077] Alternatively or additionally, in some embodiments, one or more in vitro assessments may be performed, for example: (1) in vitro expression of an antigen encoded by an RNA included in a composition; and/or (2) in vitro potency of antigen expressed from an RNA included in a composition as described herein. Example 29: Exemplary Clinical Studies of RNA Compositions essments that may be performed of certain RNA
Figure imgf000441_0002
compositions described herein. [1079] In some embodiments, more than one different composition (e.g., immunogenic composition, e.g., vaccine) candidate may be assessed. In some such embodiments, different candidates may vary, for example in: (1) RNA platform (e.g., unmodified RNA, nucleoside-modified RNA, self-amplifying RNA (saRNA), trans- amplifying RNA); (2) encoded antigen – e.g.: - which HSV (HSV-1 and/or HSV-2) protein(s) utilized, - full length protein antigen vs fragment vs plurality of fragments vs fusion with one or more heterologous sequences (e.g., membrane tether, secretion, linker(s)), and/or - epitopes from different (and/or multiple) phases of HSV life cycle; (3) number of RNAs; (4) elements of RNA construct; - cap and/or cap-adjacent sequences, - 5’ UTR, - 3’ UTR, and/or - polyA tail; (5) lipid composition of LNP. [1080] In one particular embodiment, up to three candidate compositions (e.g., immunogenic compositions, e.g., vaccines) that have only 1 RNA encoding for an exemplary HSV glycoprotein (e.g., HSV gB, gD gH, gL) or immunogenic fragment thereof, or an exemplary HSV protein or variant or immunogenic fragments thereof are evaluated and/or up to three candidates that contain 2 RNAs, one encoding for an exemplary HSV glycoprotein (e.g., HSV gB, gD gH, gL) or immunogenic fragment thereof or an HSV protein or variant or immunogenic fragment thereof and another one encoding for exemplary CD8 and/or CD4 epitopes from conserved antigens (and optionally considering conserved T-cell epitopes from various stages of the HSV life cycle are evaluated. In this particular exemplary embodiment, composition (e.g., immunogenic composition, e.g., vaccine) candidates may be evaluated by intramuscular administration, for example, based on a dose-escalation scheme. Example 30: Exemplary Production, Characterization, and or Use of Certain Exemplary Polyepitopic Compositions [1081] In some embodiments, immunogenicity of a multi-epitopic RNA or polypeptide
Figure imgf000442_0001
odents (e.g., mice, e.g., transgenic mice) and/or larger animals such as non-human primates to evaluate the magnitude of immune response induced against the epitopes tested. In some embodiments, immunogenicity of encoded exemplary epitopes in vivo can be correlated with in vitro responses of specific CTL lines against target cells expressing the multi-epitope polypeptides. Thus, in some embodiments, such exemplary experiments can show that a multi-epitopic construct serves to both: 1) generate a cell mediated and/or humoral response and 2) that the induced immune cells recognized cells expressing the encoded exemplary epitopes. [1082] In some embodiments, for example, to create a DNA sequence encoding the selected exemplary multi-epitope construct (e.g., DNA or RNA) for expression in human cells, amino acid sequences of epitopes to be included can be reverse translated. A human codon usage table can be used to guide the codon choice for each amino acid. [1083] In some embodiments, epitope-encoding DNA sequences are directly adjoined, so that when transcribed and translated, a continuous polypeptide sequence is created. [1084] In some embodiments, expression and/or immunogenicity is optimized. In some such embodiments, expression and/or immunogenicity is optimized by incorporating additional elements into an encoding construct. Without limitation, examples of amino acid sequences that can be reverse translated and included in a multi-epitopic construct sequence include, for example: HLA class I epitopes, HLA class II epitopes, an ubiquitination signal sequence, and/or an endoplasmic reticulum targeting signal. In some embodiments, HLA presentation of CTL and HTL epitopes can be improved by including synthetic (e.g., poly-alanine) or naturally-occurring flanking sequences adjacent to the CTL or HTL epitopes; larger peptides comprising the epitope(s) are within the scope of the present disclosure. [1085] In some embodiments, a multi-epitope-encoding DNA sequence can be produced by assembling oligonucleotides that encode the plus and minus strands of the construct. In some embodiments, overlapping oligonucleotides (e.g., 30-100 bases long) can be synthesized, phosphorylated, purified and annealed under appropriate conditions using suitable technique known in the art. In some embodiments, ends of utilized oligonucleotides can be joined, for example, using ligation (e.g., T4 DNA ligation). In some embodiments, synthetic constructs encoding a multi-epitopic can then be cloned into a desired expression vector (e.g., using a suitable cloning technique known in the art). [1086] In some embodiments, standard regulatory sequences well known to those of skill in the art (e.g., promoters, enhancers, etc.) can be included to ensure expression of a polypeitopic construct in target cells. In some embodiments, for example, a promoter with a down-stream cloning site for insertion of the polyepitopic construct coding sequence; a polyadenylation signal for efficient transcription termination; an E. coli origin of replication; and an E. coli selectable marker (e.g., ampicillin or kanamycin resistance). In some embodiments, a utilized promoter or promoters is not limited and can be used for this purpose, e.g., the human herpes simplex virus (hHSV) promoter. See, e.g., U.S. Patent Nos. 5,580,859 and 5,589,466 for other suitable promoter sequences. [1087] In some embodiments, vector modifications are used to optimize expression and/or immunogenicity. In some embodiments, introns are utilized for efficient gene expression, and one or more synthetic or naturally-occurring introns are incorporated into the transcribed region. In some embodiments, inclusion of stabilization sequences (e.g., mRNA stabilization sequences) and/or sequences for replication in mammalian cells are used for increasing expression. [1088] In some embodiments, once an expression vector is selected, a multi-epitopic construct coding sequence is cloned into a polylinker region downstream of a promoter (e.g., generating a “plasmid”). In some embodiments, such a plasmid is transformed into an appropriate E. coli strain, and DNA is prepared using any suitable technique known in the art. In some embodiments, the orientation and DNA sequence of the multi-epitopic encoding sequence, as well as all other elements included in the vector, are confirmed using, for example, restriction mapping and/or DNA sequence analysis. In some embodiments, bacterial cells comprising a desired plasmid can be stored, for example, as a master cell bank and/or a working cell bank. [1089] In some embodiments, immunomodulatory sequences contribute to the immunogenicity, e.g., of nucleic acid composition (e.g., immunogenic composition, e.g., vaccine) constructs. In some embodiments, such sequences are included in a vector, outside the coding sequence, if desired to enhance immunogenicity. In some embodiments, such sequences are immunostimulatory. In some embodiments, such sequences are ISSs or CpGs. [1090] In some embodiments, a bi-cistronic expression vector which allows production of both exemplary multi-epitopic construct and a second protein (e.g., included to enhance or decrease immunogenicity) are used. Without limitation, examples of proteins or polypeptides that can enhance the immune response if co-expressed with a multi-epitopic construct include cytokines (e.g., IL-2, IL-12, GM-CSF), cytokine-inducing molecules (e.g., LeIF), costimulatory molecules, or for HTL responses, pan-DR binding proteins. In some embodiments, helper (HTL) epitopes are fused and/or linked to intracellular targeting signals and expressed separately from expressed CTL epitopes; this allows direction of the HTL epitopes to a cell compartment different than that of the CTL epitopes. If required, this could facilitate more efficient entry of HTL epitopes into the HLA class II pathway, thereby improving HTL induction. In contrast to HTL or CTL induction, specifically decreasing the immune response by co-expression of immunosuppressive molecules (e.g. TGF-E) can be beneficial in certain diseases. [1091] In some embodiments, commercially-relevant quantities of plasmid DNA (e.g., for administration or for production of exemplary RNA and/or exemplary protein for administration) can be produced, for example, by fermentation in E. coli, followed by purification. In some embodiments, aliquots from a working cell bank are used to inoculate growth medium, and grown to a predetermined level (e.g., saturation) in flasks (e.g., shaker flasks) or a bioreactor according to well-known techniques. In some embodiments, plasmid DNA is purified using standard bioseparation technologies such as, for example, solid phase anion-exchange resins supplied by QIAGEN, Inc. (Valencia, California). In some embodiments, supercoiled DNA is separated from open circular and linear forms using gel electrophoresis or other suitable methods known in the art. [1092] In some embodiments, purified plasmid DNA is prepared for injection into a subject using a variety of formulations. In some embodiments, lyophilized DNA is reconstitution in sterile phosphate-buffer saline (PBS). This approach, known as “naked DNA,” and is currently being used for intramuscular (IM) administration in clinical trials. In some embodiments, to maximize the immunotherapeutic effects of polyepitopic compositions (e.g., immunogenic compositions, e.g., vaccines), an alternative method for formulating nucleic acids (e.g., purified plasmid DNA, in vitro transcribed RNA, etc) can be used. A variety of methods have been described, and new techniques can become available. In some embodiments, cationic lipids are used in the formulation (see, e.g., as described by WO 93/24640; Mannino & Gould-Fogerite, BioTechniques 6(7): 682 (1988); U.S. Pat No. 5,279,833; WO 91/06309; and Felgner, et al., Proc. Nat'l Acad. Sci. USA 84:7413 (1987). In some embodiments, glycolipids, fusogenic liposomes, peptides and compounds referred to collectively as protective, interactive, non-condensing compounds (PINC) are complexed to purified plasmid DNA to influence variables such as stability, intramuscular dispersion, or trafficking to specific organs or cell types. [1093] In some embodiments, an exemplary polynucleotide is introduced into cells by use of high-speed cell deformation. During high-speed deformation, cells are squeezed such that temporary disruptions occur in the cell membrane, thus allowing the nucleic acid to enter the cell. In some embodiments, polypeptides are produced from expression vectors, e.g., in a bacterial expression vector, for example, and the proteins can then be delivered to the cell. [1094] In some embodiments, target cell sensitization is used as a functional assay for expression and HLA class I presentation of encoded CTL epitopes. For example, in some embodiments, a polynucleotide is introduced into a mammalian cell line that is suitable as a target for standard CTL chromium release assays. In some embodiments, a transfection method used is dependent on the final formulation. In some embodiments, electroporation is used, e.g., for “naked” polynucleotides (e.g., DNA). In some embodiments, wherein cationic lipids are utilized, direct in vitro transfection is utilized as a transfection method. In some embodiments, a plasmid expressing marker protein or polypeptide (e.g., green fluorescent protein (GFP)) is co-transfected to allow enrichment of transfected cells (e.g., using fluorescence activated cell sorting (FACS)). In some embodiments, cells are then chromium-51 (51-Cr) labeled and used as target cells for epitope-specific CTL lines; cytolysis, detected by 51Cr release, indicates both production of, and HLA presentation of encoded CTL epitopes. In some such embodiments, expression of HTL epitopes can be evaluated in an analogous manner using assays to assess HTL activity. [1095] In some embodiments, in vivo immunogenicity is utilized for functional testing. In some embodiments, rodents (e.g., mice, e.g., transgenic mice expressing appropriate human HLA proteins) are immunized with a polyepitopic composition (e.g., immunogenic composition, e.g., vaccine) composition (e.g., comprising a DNA or RNA active agent). In some embodiments, dose and route of administration are formulation dependent (e.g., IM for DNA in PBS or LNP-formulated DNA or RNA, intraperitoneal (IP) for lipid-complexed DNA). In some embodiments, for example, twenty-one days after immunization, splenocytes are harvested and restimulated for 1 week in the presence of peptides encoding each epitope being tested. Thereafter, for CTL effector cells, assays are conducted for cytolysis of peptide-loaded, 51Cr-labeled target cells using standard techniques. Lysis of target cells that are sensitized by HLA loaded with peptide epitopes, corresponding to minigene-encoded epitopes, demonstrates composition (e.g., immunogenic composition, e.g., vaccine) function for in vivo induction of CTLs. Immunogenicity of HTL epitopes is evaluated in transgenic mice in an analogous manner. Example 31: Exemplary Guinea pig T cell assay – Phase 1 Uninfected Guinea pigs Step 1. Measure CD4 and CD8 T cells in naïve guinea pigs (n=10) [1096] Naïve guinea pigs will be used to standardize reagents. Two animals will be used per attempt to standardize. PMA will serve as a positive control. Pan-T cell (CD3), CD4, CD8, IFN gamma, TNF alpha, CD45 (pan leukocyte), CD1B3 (B cell marker to exclude B cells) will be standardized. Step 2. Measure CD4 and CD8 T cells in immunized guinea pigs (n=16) [1097] Three groups of guinea pigs: PBS (n=4), gE2 RNA-LNP315 (n=4), test ‘T cell string’ according to the present disclosure (n=4) will be tested. Group 1. PBS as control. Group 2. Immunize guinea pigs x 2 separated by 4 weeks with exemplary gE2 immunogenic fragment RNA (15 ug) and measure CD4 and CD8 T cell responses 10- 14 days after 2nd immunization (n=4). Group 3. Immunize guinea pigs x 2 separated by 4 weeks with test ‘T cell string’ (15 ug) according to the present disclosure and measure CD4 and CD8 T cells 10-14 days after 2nd immunization (n=4). All splenocytes will be harvested at same time. Two extra animals in group 2 will be immunized and 2 extra in group 3 for additional studies if needed (n=4). Example 32: Exemplary proteomics (HFF/HeLa) and ligandomics studies (HeLa) [1098] Cells were infected with HSV-2. HSV-2 protein expression was uently evaluated. In
Figure imgf000445_0001
particular, early, middle and late expressed HSV-2 proteins were identified. [1099] The results showed that all proteins, except for immediate-early (IE) are accumulating, and expression is rising from one time point to another (FIGS.54A-55B). IE proteins peak early (and lower than Late on 6-18 time points) and plateau after 9hr. RL2 (IE gene) and US10 are the most abundant among early time points (FIG.54B). US10 furthermore shows low immunogenicity in Hosken 2006 and Jing 2012. [1100] Example 33: Exemplary Trivalent HSV-2 Composition The present Example provides an exemplary trivalent HSV-2 composition, BNT163, that comprises a polyribonucleotide encoding a glycoprotein C (gC) antigenic fragment, a polyribonucleotide encoding a glycoprotein D (gD) antigenic fragment and a polyribonucleotide encoding glycoprotein E (gE) antigenic fragment. BNT163 is a preservative-free, fixed dose combination, sterile suspension, for dilution. In some embodiments, a compositioncontains three RNAs formulated as lipid nanoparticles (LNP), in an aqueous cryoprotectant buffer and is, after dilution with sterile normal saline, suitable for intramuscular (IM) injection. The total concentration of the three RNAs is 0.5 mg/mL. The LNP formulation can protect the RNA from extracellular RNases after administration. [1101] The composition of BNT163 is provided in Table 29. Table 29: Composition of drug product. Component Function Concentration Quantity per vial 1 (mg/mL) (mg)
Figure imgf000446_0001
ydroxybuty)azanedy)bs( exane-6, -dy)bs( - exydecanoate). ALC-0159 = 2-[(polyethylene glycol)-2000]-N,N-ditetradecylacetamide. DSPC = 1,2-Distearoyl-sn-glycero-3-phosphocholine. q.s. = quantum satis (as much as may suffice) BNT163 is filled into a vial with a fill volume of 0.2 mL. BNT163 is stored frozen at -80 to -60°C. The primary container closure system consists of a glass vial closed with a rubber stopper and sealed with an aluminum cap with a plastic flip-off disk. BNT163 is diluted, with sterile Normal Saline (0.9% sodium chloride) prior to administration. Example 34: Characterization of safety and immunogenicity of exemplary HSV Composition Candidates Objectives [1102] The present Example describes a Phase I randomized, observer-blinded, placebo-controlled, 3-part, dose escalation and expanded safety evaluation trial to evaluate the safety, tolerability, and immunogenicity of an exemplary Trivalent HSV2 compositions, BNT163, for the prevention or treatment of genital lesions caused by HSV-2 and/or HSV-1. [1103] Safety, tolerability, and immune responses after vaccination with an exemplary BNT163 composition are studied in healthy volunteers and volunteers with recurrent genital herpes. [1104] This example includes three parts: Part A, Part B and Part C. Part A includes a safety and dose escalation study in healthy individuals. Part B includes an expanded safety selection and evaluation study in healthy individuals. Part C includes a safety and immunogenicity evaluation study in individuals with recurrent HSV-2 genital herpes. BNT163 investigational medicinal product (IMP) is shown in Table 30. Table 30: Study treatments Name BNT163 T B T1 l i l i L P i i l i ifi l i l i
Figure imgf000447_0001
IMP = investigational medicinal product; LNP = lipid nanoparticle. [1105] Part A: Part A will focus on safety evaluations. Composition-induced immune responses (specifically neutralizing antibodies) will also be analyzed to assess if there is a dose-response. [1106] Part A will enroll healthy subjects aged 18 to 55 years without current or history of symptomatic genital herpes infections. Groups of six subjects will be randomized observer-blinded, placebo controlled, 5:1 to composition:placebo. Part A will evaluate four planned dose levels (DLs): 3, 10, 30 and 60 μg. In each DL, subjects will receive three administrations of a BNT163 composition or placebo at Visit 1 (Day 0), Visit 4 (Day 56±2 from Visit 1) and Visit 7 (Day 112±5 of Visit 4). [1107] Starting at 3 μg, the first two subjects will be dosed and complete 7 days (d) of follow-up post- Dose 1. If there is no trial treatment pausing, four more subjects will receive Dose 1. Once the first six subjects have completed the 7 days follow-up post-Dose 1, the following applies: x If there is no trial treatment pausing rule met in the first six subjects, six further subjects will be dosed at the same DL (thus, in total 12 subjects will be dosed in the respective DL). x If no safety findings of clinical concern are observed, dosing at 10 μg will be assessed. [1108] Enrollment of subsequent DL cohorts, 10, 30 and 60 μg, will be conducted sequentially, e.g., with two subjects completing 7 days of follow-up post Dose 1, followed by enrollment of four more subjects. If no safety findings of clinical concern are observed, dosing at 30 μg and 60 μg will be assessed. [1109] Part B: Part B will enroll healthy subjects aged 18 to 55 years without current or history of symptomatic genital herpes infections. Part B of the trial will expand the safety characterization for two BNT163 dose levels (30 μg and 60 μg) selected based on Part A data and also enable a more comprehensive assessment of the impact of pre-existing immunity to HSV-1 and HSV-2 on the safety and BNT163-induced immune responses than could be assessed in Part A. [1110] Part B may be initiated once the last cohort of 12 subjects of Part A have completed 7 days of follow-up after receiving their second trial vaccination. [1111] The enrolled subjects will be randomized 1:1 to one of two dose levels (30 μg and 60 μg) of BNT163 selected based on data from Part A. Subjects enrolled into Part B will be stratified by HSV-1/HSV-2 baseline serostatus as follows: 1. HSV-1 negative/HSV-2 negative; 100 subjects, randomized 1:1 into each of the two selected BNT163 dose groups. OR 2. Any seropositive status (HSV-1 positive/HSV-2 positive or HSV-1 positive/HSV-2 negative or HSV-1 negative/HSV 2 positive); 100 subjects, further stratified based on baseline HSV-2 serostatus (HSV-1 positive/HSV-2 positive or HSV-1 negative/HSV-2 positive vs. HSV-1 positive/HSV-2 negative) with a minimum of 40 (maximum of 60) HSV-2 seropositive subjects and a minimum of 40 (maximum of 60) HSV-2 seronegative subjects. Subjects in each statum will be randomized 1:1 into each of the two selected BNT163 dose groups. [1112] Part C: will evaluate safety and immunogenicity of 60 μg BNT163 composition in a two-dose regimen compared to a placebo injection in subjects with a history of HSV-2 recurrent genital herpes and allow for a comparative assessment of the baseline and BNT163-induced immune responses in individuals with recurrent genital herpes (such as Part C subjects) and without (such as Part B HSV-2 seropositive subjects). See FIG.21 for a schematic overview. [1113] In Part C, subjects will receive two doses of either BNT163 or placebo. The highest dose used in Parts A and B will be utilized to potentially optimize the immune response in individuals with recurrent genital herpes. Two doses of BNT163 will be given to assure composition-related memory responses and to assess the differences in individuals without recurrent genital herpes compared to individuals with recurrent genital herpes following immune challenge (Dose 1) and boost (Dose 2) with BNT163. In addition, comparisons between seronegative composition responses compared to the responses in individuals with recurrent genital herpes will be analyzed. Comparison measurements of immune response will be conducted until 28 days post-Dose 2 to match the planned pivotal trial primary endpoint. [1114] Part C will enroll ~40 subjects with a history of HSV-2 recurrent genital herpes. Subjects will be randomized 3:1 to BNT163 composition or placebo, and will receive 60 μg BNT163 composition (~30 subjects) or placebo (~10 subjects) administered as a two-dose regimen at Visits 3 and 6 (Day 56±2 of Visit 3). Outcome measures Part A and Part B [1115] Outcome measures for assessing the safety and tolerability of BNT163 in healthy adults during the dose escalating in Part A and during the expanded safety and dose evaluation in Part B include frequency of solicited local reactions (e.g., pain, erythema/redness, induration/swelling) at the injection site recorded up to 7 d after each dose, frequency of solicited systemic reactions (e.g., vomiting, diarrhea, headache, fatigue/tiredness, myalgia, arthralgia, chills, and fever) recorded up to 7 d after each dose, proportion of subjects with at least one unsolicited adverse event (AE) occurring up to 28 d after each dose, proportion of subjects in each cohort with at least one serious adverse event (SAE), or adverse events of special interest (AESI), or medically attended adverse event (MAAE) occurring up to 24 weeks post-Dose 3, and/or number and proportion of unsolicited AEs occurring up to 28 d after each dose. [1116] Outcome measures for assessing immune response and changes in HSV-2 neutralizing and binding antibody titers from baseline for composition (Part A and Part B) and in relationship to placebo (Part A) include geometric mean titers (GMTs) at each time point, geometric mean fold (GMF) change from baseline of neutralizing and binding antibody titers to each time point after vaccination, and/or proportion of subjects with seroconversion defined as a minimum of 4-fold increase from baseline of neutralizing and binding antibody titers to each subsequent time point after vaccination. [1117] An exploratory objective for Study A and Study B includes assessing the effects on pre-existing HSV-2 and/or HSV-1 serology status on immunogenic responses (e.g., neutralizing and binding antibody titers and/or number and proportion of subjects with seroconversion defined as a minimum of 4-fold increase of neutralizing antibody titers) per the following serology groups: (1) HSV-1-, HSV-2- subjects; (2) HSV-1+, HSV-2- subjects; (3) HSV-1±, HSV-2+ subjects. [1118] An additional exploratory objective for Study A and Study B includes assessing T-cell responses (e.g., absolute and fold-change from baseline for CD4 and CD8 T-cell responses to HSV-1 and HSV-2 gC2, gD2, and gE2; frequency and phenotypic characterization of cell mediated immune (CMI) responses such as CD4 and CD8 T- cell responses to HSV-1 and HSV-2 gC, gD, and gE antigenic fragments; and/or functional activity of the responses) to HSV antigens after BNT163 vaccination per different dose levels (DLs) and schedules). Assessment of B-cell responses is also contemplated, but is optional. [1119] An additional exploratory objective includes assessing cross-reactivity to HSV-1, including cross- binding of composition-induced antibodies to HSV-1 gD, gC, and gE, and cross-neutralization to HSV-1 (e.g., GMTs at each time point; GMF change from baseline of neutralizing and binding antibody titers to each time point after vaccination; and/or proportion of subjects with seroconversion defined as a minimum of 4-fold increase from baseline of neutralizing antibody titers to each subsequent time point after vaccination). [1120] An additional exploratory objective includes assessing the number of breakthrough lesions/complications attributed to HSV infection (e.g., total number and type (oral vs. genital) of breakthrough lesions/complications reported until 52 weeks post-Dose 3). [1121] FIG.20 and FIG.22 are schematic diagrams of study design for Part A and Part B. FIG.20 shows a flow diagram of Part A and Part B. FIG.22 shows the Part A dose escalation schema. Outcome measures Part C [1122] Outcome measures for assessing the safety and tolerability of BNT163 or placebo vaccination in subjects with recurrent HSV-2 genital herpes include frequency of solicited local reactions (e.g., pain, erythema/redness, induration/swelling) at the injection site recorded up to 7 d after each dose, frequency of solicited systemic reactions (e.g., vomiting, diarrhea, headache, fatigue/tiredness, myalgia, arthralgia, chills, and fever) recorded up to 7 d after each dose, proportion of subjects with at least one unsolicited adverse event (AE) occurring up to 28 d after each dose, proportion of subjects in each cohort with at least one serious adverse event (SAE), adverse events of special interest (AESI), or medically attended adverse event (MAAE) occurring up to 24 weeks post-Dose 2, and/or number and proportion of unsolicited AEs occurring up to 28 d after each dose. [1123] Outcome measures for assessing immune response and changes in HSV-2 neutralizing and binding antibody titers from baseline after BNT163 or placebo vaccination in subjects with recurrent HSV-2 genital herpes include geometric mean titers (GMTs) at each time point, e.g., at baseline, 7 days and 28 days after each dose and at 24 weeks post-dose 2, and/or geometric mean fold (GMF) change from baseline of neutralizing and binding antibody titers to each time point after vaccination, e.g., 7 days and 28 days after each dose and at 24 weeks post- dose 2. [1124] An exploratory objective for Study C includes assessing B- and/or T-cell responses (e.g., absolute and fold-change from baseline for CD4 and CD8 T- and B-cell responses to HSV-2 gC2, gD2, and gE2 comparable to e.g., 7 days and 28 days after each dose and at 24 weeks post-dose 2; frequency and phenotypic characterization of cell mediated immune (CMI) responses such as CD4 and CD8 T- and B-cell responses to HSV-2 gC, gD, and gE; and/or functional activity of the responses) to HSV-2 antigens after BNT163 or placebo vaccination in subjects with recurrent HSV-2 genital herpes. [1125] An additional exploratory objective includes assessing cross-reactivity to HSV-1, including cross- binding of composition-induced antibodies to HSV-1 gD, gC, and gE, and cross-neutralization to HSV-1 (e.g., GMTs at each time point e.g., at baseline, 7 days and 28 days after each dose and at 24 weeks post-dose 2; GMF change from baseline of neutralizing and binding antibody titers to each time point after vaccination, e.g., 7 days and 28 days after each dose and at 24 weeks post-dose 2; and/or proportion of subjects with seroconversion defined as a minimum of 4-fold increase from baseline of neutralizing antibody titers to each subsequent time point after vaccination, e.g., 7 days and 28 days after each dose and at 24 weeks post-dose 2) after BNT163 or placebo vaccination in subjects with recurrent HSV 2 genital herpes. [1126] An additional exploratory objective includes assessing changes from baseline to month 2 post-dose 2 in herpes lesion rate and HSV-2 genital shedding rate (e.g., proportion of HSV-2 PCR-positive anogenital swabs) after BNT163 or placebo vaccination in subjects with recurrent HSV-2 genital herpes. Days with HSV-2 is detected by PCR. [1127] FIG.21 is a schematic diagram of the study design for Part C. Exemplary Schedule of activities [1128] The schedule of activities for Part A and Part B includes: Visit 0 (screening), Visit 1 (Dose 1), Visit 2 (7+3 days follow-up from Visit 1), Visit 3 (28±2 days follow-up from Visit 1), Visit 4 (56±2 days from Visit 1: Dose 2), Visit 5 (7 days follow-up from Visit 4), Visit 6 (28±2 days follow-up from Visit 4), Visit 7 (112±5 days from Visit 4: Dose 3), Visit 8 (7+3 days follow-up from Visit 7), Visit 9 (28±2 days follow-up from Visit 7), Visit (Call) 10 (90±10 days follow-up from Visit 7), Visit 11 (167±10 days follow-up from Visit 7), and Visit 12 (365±10 days follow-up from Visit 7). [1129] The schedule of activities for Part C includes: Visit 0 (screening), Visit 1 (Pre-Dose visit 1; 35±2 days prior to Dose 1), Visit 2 (Pre-Dose visit 2; 28±2 days prior to Dose 1), Visit 3 (Dose 1), Visit 4 (7±3 days from Visit 3), Visit 5 (28±2 days from Visit 3), Visit 6 (Dose 2; 56±2 days from Visit 3), Visit 7 (7+3 days from Visit 6), Visit 8 (28±2 days from Visit 6), Visit 9 (168±10 days from Visit 6), and Visit 10 (365±10 days from Visit 6). Trial population [1130] Part A and Part B of this study include volunteers aged 18 to 55 years without current or history of symptomatic genital herpes infections. Part C of this trial will enroll volunteers aged 18 to 55 years with a history of recurrent genital herpes. A balanced enrollment of males and females is not required. Inclusion criteria Part A and Part B [1131] Volunteers are eligible to be included in the trial only if all of the following criteria apply: 1. Have given informed consent by signing and dating the informed consent form (ICF) before initiation of any trial-specific procedures. 2. Are aged 18 to 55 years, have a body mass index over 18.5 kg/m2 and under 35 kg/m2 and weigh at least 50 kg at Visit 0. 3. Are willing and able to comply with scheduled visits, treatment schedule, laboratory tests, and other requirements of the trial. This includes that they are able to understand and follow trial-related instructions. 4. Are overall healthy in the clinical judgment of the investigator based on medical history, physical examination, 12 lead electrocardiogram (ECG), vital signs, and screening laboratory tests (blood clinical laboratory) at Visit 0. 5. Negative HIV-1 and -2 blood test: Sites may use locally available Clinical Laboratory Improvement Amendments (CLIA)-certified assays at Visit 0. 6. Negative Hepatitis B surface antigen at Visit 0. 7. Negative anti-Hepatitis C virus (HCV) antibodies (anti-HCV), or undetectable HCV viral load if the anti-HCV is positive at Visit 0. 8. Negative syphilis test at Visit 0. 9. Volunteers of childbearing potential (VOCBP): negative serum beta human chorionic gonadotropin (β-HCG) pregnancy test at Visit 0 and negative urine pregnancy test prior to each IMP administration and at end of the trial. Volunteers born female that are postmenopausal (verified by follicle stimulating hormone [FSH] level) or permanently sterilized (verified by medical records) will not be considered VOBCP. 10. VOCBP who agree to practice a highly effective form of contraception (for guidance on highly effective forms of contraception) and to require their male partners to use condoms with a spermicidal agent, starting at Visit 0 and continuously until 60 d after receiving the last trial treatment. 11. VOCBP who agree not to donate eggs (ova, oocytes) for the purposes of assisted reproduction during trial, starting at Visit 0 and continuously until 60 d after receiving the last trial treatment. 12. Men who are sexually active with a VOCBP and have not had a vasectomy who agree to use condoms coated with a spermicidal agent and to practice a highly effective form of contraception with their partners of childbearing potential (for guidance on highly effective forms of contraception) during the trial, starting at Visit 0 and continuously until 90 d after receiving the last trial treatment. 13. Men who are willing to refrain from sperm donation, starting at Visit 0 and continuously until 90 d after receiving the last trial treatment. Inclusion criteria - Part C [1132] Volunteers are not eligible to be included in the trial only if any of the following criteria apply: 1. Have given informed consent by signing and dating the ICF before initiation of any trial-specific procedures. 2. Are aged 18 to 55 years, have a body mass index over 18.5 kg/m2 and under 35 kg/m2 and weigh at least 50 kg at Visit 0. 3. Are willing and able to comply with scheduled visits, treatment schedule, laboratory tests, and other requirements of the trial. This includes that they are able to understand and follow trial-related instructions. 4. Are overall healthy in the clinical judgment of the investigator based on medical history, physical examination, 12 lead ECG, vital signs, and screening laboratory tests (blood clinical laboratory) at Visit 0. 5. Have had a diagnosis of recurrent HSV-2 genital herpes defined as at least 3 and no more than 9 subject-reported genital herpes recurrences either in the 1-year preceding Visit 0, or, if currently on suppressive therapy, in the 1 year preceding the start of suppressive therapy. 6. Are seropositive for HSV-2 as determined by Western Blot prior to Visit 2. 7. Are willing to refrain from taking suppressive antiviral therapy from Visit 1 until the end of trial. 8. Are willing to refrain from the use of episodic antiviral therapy during the two 28-day anogenital swabbing periods. Episodic therapy may be used outside the swabbing periods. 9. Negative HIV-1 and HIV-2 blood test: Sites may use locally available CLIA-certified assays at Visit 0. 10 Negative Hepatitis B surface antigen at Visit 0. 11. Negative anti-HCV, or undetectable HCV viral load if the anti-HCV is positive at Visit 0. 12. Negative syphilis test at Visit 0. 13. VOCBP: negative serum β-HCG pregnancy test at Visit 0 and negative urine pregnancy test prior to each IMP administration and at end of the trial. Volunteers born female that are postmenopausal (e.g., verified by FSH level) or permanently sterilized (e.g., verified by medical records) will not be considered VOBCP. 14. VOCBP who agree to practice a highly effective form of contraception and to require their male partners to use condoms coated with a spermicidal agent, starting at Visit 0 and continuously until 60 d after receiving the last trial treatment. 15. VOCBP who agree not to donate eggs (ova, oocytes) for the purposes of assisted reproduction during trial, starting at Visit 0 and continuously until 60 d after receiving the last trial treatment. 16. Men who are sexually active with a VOCBP and have not had a vasectomy who agree to use condoms coated with a spermicidal agent and to practice a highly effective form of contraception with their partners of childbearing potential during the trial, starting at Visit 0 and continuously until 90 d after receiving the last trial treatment. 17. Men who are willing to refrain from sperm donation, starting at Visit 0 and continuously until 90 d after receiving the last trial treatment. Exclusion criteria Part A and Part B [1133] Volunteers are not eligible to be included in the trial only if any of the following criteria apply: 1. Breastfeeding or intending to become pregnant within the projected duration of the trial starting with Visit 0 until 60 d after receiving the last trial treatment or intending to father children within the projected duration of the trial starting with Visit 0 until 90 d after receiving the last trial treatment. 2. Current or history of symptomatic genital herpes infections. Volunteers with oral herpes or herpetic whitlow will not be excluded. 3. Current or history of any form of ocular HSV infection or HSV-related central nervous system disease or complication. 4. History of any serious adverse reactions to composition (e.g., immunogenic compositions, e.g., vaccines) or to composition (e.g., immunogenic composition, e.g., vaccine) components such as lipids, and including history of anaphylaxis and related symptoms such as hives, respiratory difficulty, angioedema, and/or abdominal pain. 5. Current or history of the following medical conditions: a. Uncontrolled or moderate or severe respiratory diseases (e.g., asthma, chronic obstructive pulmonary disease); symptoms of asthma severity as defined in the most recent National Asthma Education and Prevention Program Expert Panel report - e.g., exclude a volunteer who: i. Uses a short-acting rescue inhaler (typically a beta 2 agonist) daily, or ii. Uses high dose inhaled corticosteroids (per American Academy of Allergy Asthma & Immunology), or iii. In the past year has either of the following: 1. Had greater than one exacerbation of symptoms treated with oral/parenteral corticosteroids; 2. Needed emergency care, urgent care, hospitalization, or intubation for asthma. b. History of thyroidectomy, or thyroid disease requiring medication during the last 12 months; c. History of diabetes mellitus type 1 or type 2, including cases controlled with diet alone (not excluded: history of isolated gestational diabetes); d. Hypertension: i. If a person has been found to have elevated blood pressure or hypertension during screening or previously, exclude for blood pressure that is not well controlled. Well controlled blood pressure is defined as consistently ≤140 mm Hg systolic and ≤90 mm Hg diastolic, with or without medication, with only isolated, brief instances of higher readings, which must be ≤150 mm Hg systolic and ≤90 mm Hg diastolic at enrollment. ii. If a person does not have a history of elevated blood pressure or hypertension, also exclude for systolic blood pressure ≥150 mm Hg at enrollment or diastolic blood pressure ≥100 mm Hg confirmed by two measurements prior to enrollment. e. Malignancy within 5 years of Visit 0, excluding localized basal or squamous cell cancer; f. Current or history of cardiovascular diseases, e.g., myocardial infarction, congestive heart failure, cardiomyopathy, or clinically significant arrhythmias, myocarditis, or pericarditis; g. Bleeding disorder diagnosed by a doctor (e.g., factor deficiency, coagulopathy, or platelet disorder requiring special precautions); h. Seizure disorder: History of seizure(s) within past 3 years. Also exclude if volunteer has used medications in order to prevent or treat seizure(s) at any time within the past 3 years. 6. History of psychiatric illness, including alcohol abuse or drug addiction within 1 year before Visit 0, or a history (within the past 5 years) of substance abuse or known medical, psychological, or social conditions which, in the opinion of the investigator, could compromise their wellbeing if they participate as subjects in the trial, or that could prevent, limit, or confound the protocol specified assessments. 7. Any of the following associated with immune dysregulation: a. Primary immunodeficiencies. b. History of solid organ or bone marrow transplantation. c. Asplenia: any condition resulting in the absence of a functional spleen. d. Currently existing or history of autoimmune disease including and not limited to thyroid autoimmune disease, multiple sclerosis, psoriasis, etc. 8. Use of any non-trial IMP within 28 d before Dose 1 in this trial (Visit 1) or planned receipt continuously until Visit 12 in this trial, or participation in the active treatment phase of another interventional clinical trial. 9. Previous vaccination with an investigational herpes virus composition (e.g., immunogenic composition, e.g., vaccine) at any time. 10. Any non-trial vaccination within 28 d before Dose 1 and continuously until 28 d after receiving Dose 3. Note: Licensed inactivated and RNA vaccines (e.g., seasonal influenza and COVID 19 vaccines) are allowed to be given at least 14 d before or 14 d after each IMP administration. Licensed live attenuated vaccines (e.g., influenza vaccines) are allowed at least 28 d before or 28 d after any IMP administration. 11. Received allergy treatment with antigen injections within 28 d before first IMP administration or that are scheduled within 14 d after Visit 1. 12. Received blood/plasma products or immunoglobulin within 120 d before Visit 1 or planned administration starting at Visit 0 until completion of Visit 12. 13. Received chronic suppressive antiviral therapy for treatment of recurrent HSV 1 and/or HSV 2 genital herpes infections (i.e., oral acyclovir, oral valacyclovir, oral famciclovir, and/or intravenous ganciclovir) from 1 year prior to Visit 0 until completion of Visit 12. 14. Any existing condition which may affect composition (e.g., immunogenic composition, e.g., vaccine) injection and/or assessment of local reactions assessment at the injection site, e.g., tattoos, severe scars. 15. Vulnerable individuals as per ICH E6 definition, i.e., are individuals whose willingness to volunteer in a clinical trial may be unduly influenced by the expectation, whether justified or not, of benefits associated with participation, or of a retaliatory response from senior members of a hierarchy in case of refusal to participate. 16. Any screening hematology and/or blood chemistry laboratory value that meets the definition of a Grade ≥2 abnormality at Visit 0. For laboratory values for which toxicity grading guidance is not available or for Grade ≤1 abnormalities, subject eligibility will be determined at the discretion of the investigator. Part B only: 17. Applicable HSV serology stratum is already full or the HSV serostatus is reported as indeterminate. Exclusion criteria Part C [1134] Volunteers are not eligible to be included in the trial only if any of the following criteria apply: 1. Breastfeeding or intending to become pregnant within the projected duration of the trial starting with Visit 0 until 60 d after receiving the last trial treatment or intending to father children within the projected duration of the trial starting with Visit 0 until 90 d after receiving the last trial treatment. 2. Current or history of any form of ocular HSV infection or HSV-related central nervous system disease or complication. 3. History of any serious adverse reactions to composition (e.g., immunogenic composition, e.g., vaccine)s or to composition (e.g., immunogenic composition, e.g., vaccine) components such as lipids, and including history of anaphylaxis and related symptoms such as hives, respiratory difficulty, angioedema, and/or abdominal pain. 4. Current or history of the following medical conditions: a. Uncontrolled or moderate or severe respiratory diseases (e.g., asthma, chronic obstructive pulmonary disease); symptoms of asthma severity as defined in the most recent National Asthma Education and Prevention Program Expert Panel report – e.g., exclude a volunteer who: • Uses a short-acting rescue inhaler (typically a beta-2 agonist) daily, or • Uses high dose inhaled corticosteroids (per American Academy of Allergy Asthma & Immunology), or • In the past year has either of the following: o Had greater than one exacerbation of symptoms treated with oral/parenteral corticosteroids; o Needed emergency care, urgent care, hospitalization, or intubation for asthma. b. History of thyroidectomy, or thyroid disease requiring medication during the last 12 months; c. History of diabetes mellitus type 1 or type 2, including cases controlled with diet alone (not excluded: history of isolated gestational diabetes); d. Hypertension: • If a person has been found to have elevated blood pressure or hypertension during screening or previously, exclude for blood pressure that is not well controlled. Well controlled blood pressure is defined as consistently ≤140 mm Hg systolic and ≤90 mm Hg diastolic, with or without medication, with only isolated, brief instances of higher readings, which must be ≤150 mm Hg systolic and ≤90 mm Hg diastolic at enrollment. • If a person does not have a history of elevated blood pressure or hypertension, also exclude for systolic blood pressure ≥150 mm Hg at enrollment or diastolic blood pressure ≥100 mm Hg confirmed by two measurements prior to enrollment. e. Malignancy within 5 years of Visit 0, excluding localized basal or squamous cell cancer; f. Current or history of cardiovascular diseases, e.g., myocardial infarction, congestive heart failure, cardiomyopathy, or clinically significant arrhythmias, myocarditis, or pericarditis; g. Bleeding disorder diagnosed by a doctor (e.g., factor deficiency, coagulopathy, or platelet disorder requiring special precautions); h. Seizure disorder: History of seizure(s) within past 3 years. Also exclude if volunteer has used medications in order to prevent or treat seizure(s) at any time within the past 3 years. 5. History of psychiatric illness, including alcohol abuse or drug addiction within 1 year before Visit 0, or a history (within the past 5 years) of substance abuse or known medical, psychological, or social conditions which, in the opinion of the investigator, could compromise their wellbeing if they participate as subjects in the trial, or that could prevent, limit, or confound the protocol specified assessments. 6. Any of the following associated with immune dysregulation: • Primary immunodeficiencies. • History of solid organ or bone marrow transplantation. • Asplenia: any condition resulting in the absence of a functional spleen. • Currently existing or history of autoimmune disease including and not limited to thyroid autoimmune disease, multiple sclerosis, psoriasis, etc. 7. Use of any non-trial IMP within 28 d before Dose 1 in this trial (Visit 3) or planned receipt continuously until Visit 10 in this trial, or participation in the active treatment phase of another interventional clinical trial. 8. Previous vaccination with an investigational herpes virus vaccine at any time. 9. Any non-trial vaccination within 28 d before Dose 1 and continuously until 28 d after receiving Dose 2. Note: Inactivated and RNA vaccines (e.g., seasonal influenza and COVID 19 vaccines) are allowed to be given at least 14 d before or 14 d after each IMP administration. Live attenuated vaccines (e.g., influenza vaccines) are allowed at least 28 d before or 28 d after any IMP administration. 10. Received allergy treatment with antigen injections within 28 d before first IMP administration or that are scheduled within 14 d after Visit 3. 11. Received blood/plasma products or immunoglobulin within 120 d before Visit 3 or planned administration starting at Visit 0 until completion of Visit 10. 12. Any existing condition which may affect vaccine injection and/or assessment of local reactions assessment at the injection site, e.g., tattoos, severe scars. 13. Vulnerable individuals as per ICH E6 definition, i.e., are individuals whose willingness to volunteer in a clinical trial may be unduly influenced by the expectation, whether justified or not, of benefits associated with participation, or of a retaliatory response from senior members of a hierarchy in case of refusal to participate. This includes investigator site staff directly involved in the conduct of the trial and their family members, site staff otherwise supervised by the investigator, and sponsor and sponsor delegate employees directly involved in the conduct of the trial and their family members. 14. Any screening hematology and/or blood chemistry laboratory value that meets the definition of a Grade ≥2 abnormality at Visit 0. For laboratory values for which toxicity grading guidance is not available or for Grade ≤1 abnormalities, subject eligibility will be determined at the discretion of the investigator. Concomitant Therapies During Trial [1135] Trial subjects should not receive prophylactic antipyretics and other pain medication to prevent symptoms associated with IMP administration. However, if a trial subject is taking a medication for another condition, even if it may have antipyretic or pain-relieving properties, it should not be withheld prior to IMP administration in this trial. [1136] Administration of standard therapeutic dose of acetaminophen (preferable), or a non-steroidal anti- inflammatory drug (NSAID) if acetaminophen is contraindicated is permitted (paracetamol / acetaminophen at doses of up to 4 g/day). [1137] Inhaled, topical, or localized administration of corticosteroids (e.g., intra-articular or intrabursal administration) are permitted. [1138] The use of antipyretics and other pain medication to treat symptoms after IMP administration or for ongoing conditions is permitted, but not for prophylaxis (i.e., up to 12 h before each IMP administration). [1139] Other concomitant medication may be considered on a case by case basis by the investigator, if required after consultation with the trial Medical Monitor. Assessments [1140] HSV-1/HSV-2 serology is assessed by commercial HSV type-specific antibody (IgG) tests and a Western Blot test to detect antibody responses to HSV-1/HSV-2 specific viral proteins. [1141] Perceived pain at the injection site is assessed as absent, mild, moderate, or severe, according to the grading scale in Table 31. [1142] Subjects are provided with a ruler to measure erythema / redness and induration / swelling. Erythema / redness and induration / swelling are measured at the greatest single diameter and recorded and then categorized during analysis as absent, mild, moderate, or severe, based on the grading scale in Table 31. Table 31: Local Reaction Grading Scale Mild Moderate Severe Potentially life-threatening (Grade 1) (Grade 2) (Grade 3) (Grade 4) c tis s
Figure imgf000458_0001
measurement should be recorded as a continuous variable. Erythema/redness should be measured using an appropriate measuring device provided by the sponsor. [1144] b. Induration/swelling should be measured using an appropriate measuring device provided by the sponsor. [1145] c. Investigator or medically qualified person confirmation is required for all reactogenicity graded as Grade 3 or Grade 4. Modified from the US FDA guidance (US FDA 2007) [1146] Symptoms of systemic reactions are assessed as absent, mild, moderate, or severe, according to the grading scale in Table 32. Table 32: Systemic reaction grading scale Mild Moderate Severe Potentially life- thr t nin
Figure imgf000458_0002
Mild Moderate Severe Potentially life- (Grade 1) (Grade 2) (Grade 3) threatening G d 4 a in
Figure imgf000459_0001
graded as Grade 4. Grade 4 reactions will be documented in the EDC system only and not in the subject’s e-diary. Abbreviations: C = Celsius; EDC = electronic data capture; e-diary = electronic diary; F = Fahrenheit; h = hours. Modified from the US FDA guidance (US FDA 2007). Genital herpes assessment – Part C [1148] To support exploratory objectives assessing change from baseline in genital herpes lesions and anogenital HSV-2 shedding rates, the following assessments will be performed: • Symptom diary: Symptoms of genital herpes will be collected daily (on paper) during two predefined 28-day periods. • Anogenital swabs for HSV-1/2 DNA PCR: anogenital swabs will be collected by the subject twice daily (morning and evening) during two predefined 28-day periods. • Genital herpes episode diary: Starting at Dose 1, subjects will be asked to complete a symptom diary (on paper) for any episode of genital herpes lesions. • Unscheduled genital herpes episode visits: Up to two unscheduled genital herpes episode visits will be conducted, one visit between Visits 2–3 and one between Visits 8–10. A brief (symptom-directed) physical exam will be conducted and lesion swabs as well as blood samples for humoral and cell-mediated immune responses will be obtained. Immune Responses [1149] Immune responses such as neutralization and binding antibody titers are assessed at visits 0, 2, 5 and 8 for Parts A and Part B, and visit 0, 4, and 7 for Part C. Cell mediated immune (CMI) responses and explorative research are assessed at Visits 1-9 and 11-12 for Part A, Visits 1, 5, 6, 8, and 11 for Part B, and at Visits 1, 3, 7, 8, 9, and 10 for Part C. Neutralization and binding antibody titers [1150] Neutralizing antibody titers and binding antibody titers assessments to HSV comprise of: x A functional antibody titer, e.g., viral neutralization test or an equivalent assay (e.g., pseudo-viral neutralization test assay). x Seronegative is defined as titers below the starting dilution (i.e., below the limit of detection for the assay). x Seroconversion after immunization is defined as a 4-fold increase in titer. o For seronegative pre-immunization sera: a titer which is ≥ 4-fold the limit of detection. o For seropositive pre-immunization sera: a titer which is 4-fold the measured pre- immunization titer. x An antibody binding assay, e.g., ELISA or an equivalent assay. o For seronegative pre-immunization sera: a titer which is ≥ 4-fold the limit of detection. o For seropositive pre-immunization sera: a titer which is 4-fold the measured pre- immunization titer. Cell mediated immune (CMI) responses [1151] Blood samples will be used to describe CMI responses to HSV-2: x T-cell responses include responses mediated by immune cells such as CD4 and CD8 T cells and their functional phenotypic subset by, e.g., enzyme-linked immunospot, intracellular cytokine staining, multimer analyses, cytokine secretion assays, flow cytometry, etc. x These analyses may further include frequency of CD4 and CD8 T cells and responses, such as (but not limited to) IL-2, IFN-gamma and activation markers after in vitro stimulation with peptide pools from the BNT163 vaccine antigens. Additional Characterizations [1152] Further exploratory research analyses may be conducted using residual biological samples from subsets of subjects in order to further characterize the composition (e.g., immunogenic composition, e.g., vaccine) induced and/or innate immune responses, explorative biomarkers, and/or immunogenicity research, e.g., phenotypic or functional characterization of antigen-specific B cells and T cells (e.g., by flow cytometry-based phenotyping including multimer staining), analysis of B-cell receptor / T-cell receptor repertoire (e.g., by next generation sequencing, single cell RNA sequencing), multiplex-cytokine, and inflammatory protein analysis. Transcriptomic, epigenomic, metabolomic, or phenotypic or functional characterization of other immune cell populations (e.g., innate immune cells) that may be relevant to understand the composition (e.g., immunogenic composition, e.g., vaccine) induced immune responses or for the product development for BNT163 or related therapeutic areas may be included in this example. [1153] Blood samples for research may be used to describe the following immune responses to HSV-2: x T cell and B cell analyses may include characterization of composition (e.g., immunogenic composition, e.g., vaccine) induced plasma blasts and HSV antigen-specific B cells and T cells to identify B cells and T cells recognizing conserved and strain-specific epitopes which could include but is not limited to protein expression profile, gene expression or transcriptomic profile, epigenomic or open chromatin region profiling or metabolomic profiling. Further single cell variability diversity joining analysis and somatic hyper-mutation analysis may also be performed on B cells, and repertoire analysis on T cells. x Some of the sample may be used for sequencing of trial subjects’ antibody and/or B-cell receptor heavy- and light-chain genes, T-cell receptor genes, and/or gene expression and/or epigenetic profile, for further understanding the B-cell, T-cell, and antibody repertoires and for generating monoclonal antibodies for product development or for development of T cell-based therapies. x Exploratory research on antibody activity to investigate composition (e.g., immunogenic composition, e.g., vaccine)-induced immune responses, including blocking of complement (e.g, C3b) binding to gC, blocking of IgG Fc binding to gE and other Fc-mediated antibody functions, may be measured in assays including the following but not limited to: antibody-dependent cellular cytotoxicity, antibody dependent cell mediated viral inhibition assays, Fc-mediated phagocytic activity, complement binding and cell-to-cell virus spread. Investigation of antibody functionality and specificity to HSV antigens may also include antibody epitope mapping studies. In addition, in Part C of this trial, anogenital swabs will be collected in subjects with recurrent HSV-2 genital herpes as described in Section 13.4 to assess the mucosal antibody response. Adverse Events [1154] An adverse event (AE) is any untoward medical occurrence in a trial subject administered a pharmaceutical product and which does not necessarily have to have a causal relationship with this treatment. An AE can therefore be any unfavorable and unintended sign (including an abnormal laboratory finding), symptom, or disease (new or exacerbated) temporally associated with the use of a medicinal product, whether or not considered related to the medicinal product. A medically attended adverse event (MAAE) is an unsolicited AE for which the subjects received medical attention defined as hospitalization, or an otherwise unscheduled visit to or from medical personnel for any reason, including emergency room visits. [1155] Events meeting the AE definition: x Any abnormal laboratory test results (e.g., hematology and clinical chemistry) or other safety assessments (e.g., ECG, vital signs measurements), including those that worsen from baseline, considered clinically significant in the medical and scientific judgment of the investigator. x Exacerbation of a chronic or intermittent pre-existing condition including either an increase in frequency and/or severity of the condition. x Serious adverse event (SAE) and non-serious trial procedure-related AEs occurring after the date informed consent was given (ICF signed and dated) and AEs (non-serious AEs) occurring after first IMP administration. x Signs, symptoms, or the clinical sequelae of a suspected drug-drug interaction. x Signs, symptoms, or the clinical sequelae of a suspected overdose of either trial treatment or a concomitant medication. Overdose per se will not be reported as an AE/SAE (for actions in case of overdose or errors in drug administration, see Section 8.4). [1156] Events not meeting the AE definition: x Any clinically significant abnormal laboratory findings or other abnormal safety assessments which are associated with the underlying disease, unless judged by the investigator to be more severe than expected for the trial subject’s condition. x Medical or surgical procedure (e.g., endoscopy, appendectomy): the condition that leads to the procedure is the AE. x Situations in which an untoward medical occurrence did not occur or continue (social and/or convenience admission to a hospital). Serious Adverse Events [1157] A serious adverse event (SAE) is defined as any untoward medical occurrence that, at any dose, results in death or is life threatening. [1158] The assessment of AE and/or SAE intensity is consistently done for all trial subjects treated with the same treatment and dose. For further guidance on AE and SAE assessments, see, for example US FDA 2007 guidance, incorporated herein by reference in its entirety. Where specific guidance for an AE term is not provided, the following general approach is followed: Grade 1 - Mild; does not interfere with the trial subject’s usual function; Grade 2 - Moderate; interferes to some extent with the trial subject’s usual function; Grade 3 - Severe; interferes significantly with the trial subject’s usual function; and Grade 4 - Potentially life-threatening; life-threatening consequences, urgent intervention required. Results [1159] Analysis of the safety data available from Part A shows that pain at the injection site and erythema/redness were the most common local reactions overall, regardless of dose. Fatigue, headache, and muscle pain were the most common systemic reactions. The pooled rates of local reactions (10 subjects [83.3%]) and systemic reactions (11 subjects [91.7%]) were higher among the 30 μg cohort in comparison with the other cohorts. Regarding the severity of the events reported, most of the local reactions were mild. Example 35: Exemplary Immunopeptidomics studies (A375 cell line) [1160] pitopes upon immunization with exemplary
Figure imgf000462_0001
polyribonucleotides as provided herein. [1161] A375 cells were transfected with four different T-cell RNA constructs: x RNA construct 1 (Het 1): comprising a polyribonucleotide sequence (SEQ ID NO: 619) encoding a polypeptide comprising two RL2 antigenic fragments, a RS1 antigenic fragment and a UL54 antigenic fragment (SEQ ID NO: 595). x RNA construct 4 (Het 4): comprising a polyribonucleotide sequence (SEQ ID NO: 622) encoding a polypeptide comprising a UL9 antigenic fragment, a UL49 antigenic fragment, a UL39 antigenic fragment, and a UL29 antigenic fragment (SEQ ID NO: 600). x RNA construct 6 (Het 6): comprising a polyribonucleotide sequence (SEQ ID NO: 624) encoding a polypeptide comprising a UL52 antigenic fragment, two UL5 antigenic fragments, a UL40 antigenic fragment, and two UL30 antigenic fragments (SEQ ID NO: 601). x RNA construct 8 (Het 8): comprising a polyribonucleotide sequence (SEQ ID NO: 626) encoding a polypeptide comprising a UL48 antigenic fragment, a UL25 antigenic fragment, a UL47 antigenic fragment, a UL46 antigenic fragment, two UL27 antigenic fragments, a UL21 antigenic fragment, a UL19 antigenic fragment, and a UL1 antigenic fragment (SEQ ID NO: 602). [1162] A375 cell line carries the following alleles A*02:01 (MAPTAC) and B*07:02 (MAPTAC)), as well as the following alleles that are endogenous to A375 cells: A*01:01, B*44:03, B*57:01, C*06:02, and/or C*16:01. After transfection with the T-cell RNA constructs the A375 cells were lysed. Antigenic fragments were pulled down using monomorphic anti-HLA monoclonal antibodies (W6/32). [1163] Table 33 shows T-cell RNA construct HLA-I epitopes in response to RNA construct 1 (Het 1) detected by mass spectrometry. FIG.24 provides a graphic overview of all HLA-I epitopes (A) and specific A*02:01 epitopes (B) in response to T-cell RNA construct 1 (Het 1). Table 33: HLA-I epitopes. String Antigenic Sequence SEQ ID Length Allele RECON % (Het#) Fragment NO: Rank
Figure imgf000463_0001
1 RS1 YLACEVLPAV 777 10 A*02:01 0.180 1 RS1 YPDAPPLRL 778 9 B*07:02 0.047
Figure imgf000464_0002
detected by mass spectrometry. FIG.25 provides a graphic overview of T-cell RNA construct HLA-I epitopes (A) specific A*02:01 epitopes (B) in response to RNA construct 4 (Het 4). Table 34: HLA-I epitopes. String Antigenic Sequence SEQ ID Length Allele RECON % (Het#) Fragment NO: Rank
Figure imgf000464_0003
[1165] Table 35 shows T-cell RNA construct HLA-I epitopes in response to RNA construct 6 (Het 6) d d b FIG 26 id hi i f T ll RNA HLA I i A %
Figure imgf000464_0001
6 UL30_1 LPAPVVLEF 801 9 B*07:02 0.004 6 UL30_2 GLLPCLHVA 802 9 A*02:01 0.305
Figure imgf000465_0004
detected by mass spectrometry. FIG.27 provides a graphic overview of T-cell RNA construct HLA-I epitopes (top) specific A*02:01 epitopes (bottom) in response to RNA construct 8 (Het 8). Table 36: HLA-I epitopes. String Antigenic Sequence SEQ ID Length Allele RECON % (Het#) Fragment NO: Rank
Figure imgf000465_0005
. String (Het#) Antigenic Fragment Epitope count
Figure imgf000465_0001
Figure imgf000465_0002
Figure imgf000465_0003
1 RS1 5 1 RL2.2 4
Figure imgf000466_0002
, q p p , B*07:02, A*01:01, C*06:02, and/or C*16:01). The 30 detected HLA-I epitopes cover 15 of the 23 HSV-2 antigenic fragments included in the tested RNA constructs. The four RNA constructs (Het 1, 4, 6 and 8) showed a number of highly immunogenic fragments. In particular, fragments RL2 and UL54 in Het 1, fragments UL9, UL39 and UL29 in Het 4, fragments UL5, UL40 and UL30 in Het 6 and fragments UL49, UL46 and UL21 in Het 8 showed high immunogenic potential (300 spots/Mil). Example 36: Exemplary Immunogenicity study in mice [1168] The pre s antigen specific T cell response and polyfunctional CD8 T cell response
Figure imgf000466_0001
upon immunization with exemplary polyribonucleotides as provided herein. [1169] Immunogenicity was assessed using transgenic mice carrying a transgene comprising of the human A*02:01 gene and mouse H2-Kb gene which encodes a chimeric class I molecule. [1170] Mice were grouped into 11 groups and immunized with RNA construct 1(Het 1), RNA constructs 4 (Het 4), RNA constructs (Het 6), or RNA constructs 8 (Het 8), or combinations thereof, as shown below. Two control groups were included; PBS, and a group immunized with three strings encoding an HSV-2 glycoprotein glycoprotein C (gC) antigenic fragment, an HSV-2 glycoprotein D (gD) antigenic fragment, and an HSV-2 glycoprotein E (gE) antigenic fragment. Table 38: Experimental setup Groups String (Het#) Dose (μg) per Immunization schedule Takedown and readout injection 1 1 1
Figure imgf000467_0001
uthanized on Day 33 and Day 35 and spleens were analyzed using ELISpot and intracellular cytokine staining (ICS). [1172] ELISpot: Antigen specific T cells were elicited to most of the antigen fragments when the RNA constructs were given individually (Groups 1-4), in combination of four (Group 5; RNA constructs 1, 4, 6, and 8 combined) (FIG.28), combination of two (Group 6; RNA construct 1 and RNA construct 8) (FIG.29), and a combination of three (Group 7; RNA construct 1, RNA construct 4, and RNA construct 6) (FIG.30). The antigenic fragments were classified into three groups based on the strength of their response; (a) low score (<100 spots per 1x106 cells), (b) moderate score (100-300 spots per 1x106 cells), or (c) high score (>300 spots per 1x106 cells) (FIG. 32). The UL49, UL52, UL27 and UL48 antigenic fragments showed less than 100 spots per 1x106 cells (low score). The RL2a (RL2.1), RL2b (RL2.2), RS1, UL30b (UL30.2), UL19, and UL25 antigenic fragments showed between 100- 400 spots per 1x106 cells (moderate score). The UL54, UL29, UL39, UL9, UL30a (UL30.1), UL40, UL5a (UL5.1), UL5b (UL5.2), UL21, and UL46 antigenic fragments showed more than 400 spots per 1x106 cells (high score). Plots showing T cell response to the antigenic fragments are shown in FIG.33 (Group 5: RNA construct 1, RNA construct 4, RNA construct 6, and RNA construct 8), FIG.34 (Group 7: RNA construct 1, RNA construct 4, and RNA construct 6), and FIG.35 (Group 6: RNA construct 1 and RNA construct 8). [1173] Intracellular cytokine staining (ICS): Spleens were processed to single cell suspension and stimulated with individual peptide pools overnight at 0.3 uM. Concanavalin A was used as a positive control, while DMSO was used as a negative control. After overnight stimulation, anti-CD107a was added for 1 hour followed by a 4-hrs incubation with BFA+ monesin (1:1000). Surface and intracellular staining was performed after BFA + monesin incubation. Mice receiving all four RNA constructs (Group 10; RNA construct 1, RNA construct 4, RNA construct 6, and RNA construct 8) showed UL54, UL29, UL40, and UL47-specific polyfunctional CD8 T cell response (FIG.36 and FIG.37). Example 37: Exemplary RNA construct efficacy in Mice [1174] The present Example shows HSV T-cell efficacy in mice immunized with exemplary T-cell RNA as provided herein. [1175] Female A02 Mice were immunized twice about 3 weeks apart followed by a medroxyprogesterone injection (2 mg/mouse) and a challenge infection with a lethal dose of vaginal HSV-2 (1x106 PFU HSV-2 (500 LD50)). Animals were scored for overall loss of body weight, lethality, survival, genital disease, and vaginal swabs were performed on days 2 and 4 post-infection (FIG.38). [1176] RNA constructs 1, 4, 6, and 8 formulated in lipid nanoparticles (LNP) were administered at doses and combinations as shown in Table 39. RNA construct information is as described in Example 35. Group 8 was administered with trivalent RNA encoding an HSV-2 gC (gC2) antigenic fragment, an HSV-2 gD (gD2) antigenic fragment and an HSV-2 gE (gE2) antigenic fragment formulated in LNPs. Table 39: Experimental setup Groups RNA constructs Dose (μg) per injection 1 1 Immediate early (IE) 1
Figure imgf000468_0001
g . , . , ulative survival (FIGS.45-46) and viral titers (FIG.47). [1178] Weight loss was graphed as % body weight on the observation day (days post challenge) compared to weight taken just prior to challenge. Overall loss of body weight in all groups was observed (FIG.39). [1179] Neither the RNA construct or RNA construct combinations administered to group 1 to 7 protected against lethality. The RNA encoding HSV-2 gC, gD, gE antigenic fragments administered to group 8 elicited full protection against lethal HSV-2 genital infection (FIG.40). A positive trend was observed in the combination groups (Group 5, 6, and 7), in particular the 4-way combination (Group 5) (FIGS.41-42). Comparable data are produced when two different scoring criteria are used. Using both criteria, the 4-RNA construct combination (administered to Group 5) is the most efficacious in prolonging survival of the mice (FIGS.43-44). Improvement in cumulative survival observed in individual RNA constructs: IE (Group 1), E-1 (Group 2) and Late (Group 4) (FIG.45A). Improvement in cumulative survival observed in all combo groups but particularly 4-way combo (Group 5) (FIG. 45B). Cumulative survival for all groups are shown in FIG.46A. Mice immunized with 4-RNA construct (Group 5) have the highest cumulative survival among the tested groups. Mice immunized with Early 2 RNA construct (Group 3) shows the lowest efficacy in this readout and overlaps with saline group (FIG.46B). [1180] Both plaque assay and qPCR were used to measure vaginal viral titers. Significant reduction of vaginal viral titers with IE (Group 1), E-2 (Group 3), 4-way combination (Group 5) at day 2 was observed (FIGS. 47A and 47C). 4-way combination (Group 5) and IE & L combination (Group 6), but none of the individual RNA constructs, showed significantly reduced viral titers at day 4 (FIGS.47B and 47D). Reduction of titers could be due to restriction of primary replication elicited by mucosal immunity. Example 38: Exemplary T-cell RNA construct Protein Expression [1181] The present Example shows that cells transfected with exemplary HSV-2 T-cell RNA construct express the corresponding proteins. [1182] HET293T cells were transfected with ten different T-cell RNA constructs (FIG.48): x RNA construct 15: comprising a polyribonucleotide sequence (SEQ ID NO: 653) encoding a polypeptide comprising a RL2 antigenic fragment, a UL54 antigenic fragment, a UL47 antigenic fragment, a UL46 antigenic fragment, and a UL21 antigenic fragment (SEQ ID NO: 633). x RNA construct 16: comprising a polyribonucleotide sequence (SEQ ID NO: 655) encoding a polypeptide comprising a UL21 antigenic fragment, a UL49 antigenic fragment, a UL54 antigenic fragment, and a RL2.1 antigenic fragment (SEQ ID NO: 634). x RNA construct 17: comprising a polyribonucleotide sequence (SEQ ID NO: 657) encoding a polypeptide comprising a UL29 antigenic fragment, a UL39 antigenic fragment, a UL9 antigenic fragment, a UL5.1 antigenic fragment, a UL5.2 antigenic fragment, a UL40 antigenic fragment, and a UL30.1 antigenic fragment (SEQ ID NO: 635). x RNA construct 18: comprising a polyribonucleotide sequence (SEQ ID NO: 659) encoding a polypeptide comprising a UL30.1 antigenic fragment, a UL40 antigenic fragment, a UL5.2 antigenic fragment, a UL5.1 antigenic fragment, a UL9 antigenic fragment, a UL39 antigenic fragment, and a UL29 antigenic fragment (SEQ ID NO: 636). x RNA construct 19: comprising a polyribonucleotide sequence (SEQ ID NO: 661) encoding a polypeptide comprising a RL2.1 antigenic fragment, a UL54 antigenic fragment, a UL9 antigenic fragment, a UL39 antigenic fragment, and a UL5.1 antigenic fragment (SEQ ID NO: 637). x RNA construct 20: comprising a polyribonucleotide sequence (SEQ ID NO: 663) encoding a polypeptide comprising a UL5.1 antigenic fragment, a UL39 antigenic fragment, a UL9 antigenic fragment, a UL54 antigenic fragment, and a RL2.1 antigenic fragment (SEQ ID NO: 638). x RNA construct 21: comprising a polyribonucleotide sequence (SEQ ID NO: 665) encoding a polypeptide comprising a UL47 antigenic fragment, a UL46 antigenic fragment, a UL21 antigenic fragment, a UL5.2 antigenic fragment, a UL40 antigenic fragment, a UL30.1 antigenic fragment, and a UL29 antigenic fragment (SEQ ID NO: 639). x RNA construct 22: comprising a polyribonucleotide sequence (SEQ ID NO: 667) encoding a polypeptide comprising a UL29 antigenic fragment, a UL30.1 antigenic fragment, a UL40 antigenic fragment, a UL5.2 antigenic fragment, a UL21 antigenic fragment, a UL46 antigenic fragment, and a UL47 antigenic fragment (SEQ ID NO: 640). x RNA construct 23: comprising a polyribonucleotide sequence (SEQ ID NO: 669) encoding a polypeptide comprising a RL2.1 antigenic fragment, a UL54 antigenic fragment, a UL5.2 antigenic fragment, a UL40 antigenic fragment, a UL47 antigenic fragment, and a UL46 antigenic fragment (SEQ ID NO: 641). x RNA construct 24: comprising a polyribonucleotide sequence (SEQ ID NO: 671) encoding a polypeptide comprising a UL46 antigenic fragment, a UL47 antigenic fragment, a UL40 antigenic fragment, a UL5.2 antigenic fragment, a UL54 antigenic fragment, and a RL2.1 antigenic fragment (SEQ ID NO: 642). [1183] All transfections were performed with or without a proteasome inhibitor. Cells were hereafter lysed. Proteins were extracted, digested and TMT0 labeled. Protein expression was quantified by mass spectrometry using a MITD tryptic peptide. [1184] Protein expression of each T-cell RNA construct with proteasome inhibitor is shown in FIG.49 and without proteasome inhibitor is shown in FIG.50. Proteins from HSV-2 T-cell RNA constructs were detected for all RNA constructs (RNA constructs 15-24) with and without proteasome inhibitor. Protein expression was compared between RNA construct 15 and RNA construct 16, RNA construct 17 and RNA construct 18, RNA construct 19 and RNA construct 20, RNA construct 21 and RNA construct 22, and RNA construct 23 and RNA construct 24. Protein expression was higher for RNA construct 15 compared to RNA construct 16, higher for RNA construct 17 compared to RNA construct 18, higher for RNA construct 20 compared to RNA construct 19, RNA construct for RNA construct 22 compared to RNA construct 21, and higher for RNA construct 23 compared to RNA construct 24 for both testing conditions (with or without proteasome inhibitor). RNA construct 15 and RNA construct 17, comparable to RNA construct 20 and RNA construct 22, showed less sensitivity to proteasome inhibitor conditions, suggesting that RNA construct 15 and RNA construct 17 are more stable compared to RNA construct 20 and RNA construct 22 (FIG.49 and FIG.50). Example 39: Exemplary Immunogenicity study in mice (A02 and Balb/C)
Figure imgf000470_0001
olyfunctional CD8 T cell response upon immunization with exemplary polyribonucleotides as provided herein. [1186] Immunogenicity was assessed using transgenic mice carrying a transgene comprising of the human A*02:01 gene and mouse H2-Kb gene which encodes a chimeric class I molecule or balb/c mice. [1187] Mice were grouped into 8 groups and immunized with RNA construct 15 (Het 15), RNA construct 17 (Het 17), RNA construct 20 (Het 20), RNA construct 23 (Het 23), RNA construct 23 (Het 23) or combinations thereof, as shown below in Table 40. Two saline control groups were included. Table 40: Experimental setup Groups RNA constructs Mouse strain Immunization Dose (μg) per injection 1 H 15 + H 17 D0 D21 4 l 4 2 h
Figure imgf000470_0002
8 Saline n/a .
Figure imgf000471_0001
[1189] ImmunoSpot: Two saturation criteria were applied: loose (approx. 200 spots per 1x106 cells) and stringent (<100 spots per 1x106 cells). Antigen specific T cell responses were elicited all antigen fragments when RNA constructs 15 and 17 were administered (Group 1; RNA constructs 15 and 17 combined) (FIG.51), when RNA constructs 20 and 22 were administered (Group 2; RNA constructs 20 and 23 combined) (FIG.52) and when RNA construct 23 was administered (Group 3: RNA construct 23 alone) (FIG.53). The different criteria do not affect the specific T-cell response. Immunogenicity in Balb/c mice are shown in FIGS.54-56. UL5.2 lost immunogenicity in Balb/c mice. [1190] Average spot count (minus DMSO)/million cells are shown in Table 41. Group 1 and Group 3 were compared to RNA constructs 1, 4, 6, and 8 (see construct information in Example 35). Table 41: Experimental setup RNA construct Group 1 Group 3 (Het#) (Het 15 and Het 17) (Het 23)
Figure imgf000471_0002
[ 9 ] C vs. C 8 response: Ony 00,000 ce s were pated per we s n C /C 8 depeton we s. CD4 vs. CD8 response results for Group 1 (Het 15 and Het 17) are shown in FIGS.57 and 58 and for Group 3 (Het 23) in FIGS.59 and 60. Example 40: Exemplary RNA construct efficacy in Mice [1192] The present Example shows HSV T-cell efficacy in mice immunized with an exemplary T-cell RNA construct as provided herein. [1193] Female A02 Mice were immunized twice about 3 weeks apart followed by a medroxyprogesterone injection (2 mg/mouse) at day 46 and a challenge infection with a lethal dose of vaginal HSV-2 (5x103 PFU HSV-2 (~10xLD50) at day 51. Animals were scored for overall loss of body weight, lethality, survival, and genital disease. Vaginal swabs were performed 6-hours after the challenge HSV-2 infection as well as on day 2, day 4 and day 7 after the challenge HSV-2 infection (FIG.61). Dorsal root ganglia will be evaluated for HSV-2 DNA copy number by qPCR at the end of the experiment. [1194] RNA constructs 15, 17, and 23 formulated in lipid nanoparticles (LNP) were administered at doses and combinations as shown in Table 42. RNA construct information is as described in Example 38. Group 3 was administered trivalent RNA encoding an HSV-2 gC (gC2) antigenic fragment, an HSV-2 gD (gD2) antigenic fragment and an HSV-2 gE (gE2) antigenic fragment formulated in LNPs. Table 42: Experimental setup Groups RNA constructs (#Het) Dose (μg) per injection 1 PBS n/a
Figure imgf000472_0001
62), survival (FIG.63 and FIG.66), vaginal disease (FIG.64), vaginal replication (FIG.65) and viral titers (FIGS.67-68). [1196] Weight loss was graphed as % body weight on the observation days (days post challenge) compared to weight taken just prior to challenge. Overall loss of body weight in all groups was observed, except in Group 4 (BNT163) (FIG.62). [1197] RNA construct 23 showed 50% improvement in survival rate compared to only 10% in the PBS control group. RNA constructs 15 and 17 in combination showed a positive trend in prolonged survival. RNA encoding HSV-2 gC, gD, gE antigenic fragments administered to group 4 elicited full protection against lethal HSV-2 genital infection (FIG.63). Vaginal disease was not significantly reduced in group 2 (Het 23) and group 3 (Het 15 and Het 17) compared to the PBS group (FIG.64). Viral replication kinetics are shown in FIG.65. Day 2 and Day 4 post- challenge showed the highest replication of virus, compared to 6 hours and 7 days post-challenge. Most virus was cleared by day 7 (FIG.65). A positive trend in cumulative survival was observed in group 2 (Het 23) and group 3 (Het 15 and Het 17) (Group 5, 6, and 7) (FIG.66). [1198] Day 4 titers were reduced significantly four group 2 (Het 23) and group 3 (Het 15 and Het 17) compared to PBS control leading to faster clearance of virus. BNT163 was significantly different from PBS control on Day 2 and Day 4 (FIG.67). Low level viral titers were detected in a few mice at 6-hour eclipse phase, by day 7 most of the virus was cleared from vaginal cavity (FIG.68). [1199] In summary, RNA construct 23 and a combination of RNA construct 15 and 17 have significant effect on viral replication on day 4. A nonsignificant increase in survival was observed with RNA construct 23. Example 41: Exemplary RNA construct efficacy in Mice [1200] The present Example further demonstrates HSV T-cell efficacy in mice immunized with an exemplary T-cell RNA construct or combinations thereof as provided herein. [1201] Female Balb/c mice were immunized twice about 3 weeks apart followed by a medroxyprogesterone injection (2 mg/mouse) at day 46 and a challenge infection with a lethal dose of vaginal HSV-2 (275xLD50 (5x103pfu)) at day 51. Animals were scored for overall loss of body weight, lethality, survival, and genital disease. Vaginal swabs were performed 6-hours after the challenge HSV-2 infection as well as on day 2, day 4 and day 7 after the challenge HSV-2 infection (FIG.61). The swabs were stored at -80C until virus culture was performed. Plaque assay was performed to determine the intravaginal replication titers. [1202] RNA constructs BNT163, Het 15, Het 17, and Het 23 formulated in lipid nanoparticles (LNP) were administered at doses and combinations as shown in Table 43. RNA construct information is as described in Example 33 and Example 38 Table 43: Experimental setup Groups RNA constructs (dose per injection ) 1 Untranslatable control “S-TCS” (4Pg)
Figure imgf000473_0001
[1203] Female Balb/c mice (5 per group) were monitored for weight loss (FIG.69), survival (FIG.70), vaginal disease (FIG.71), vaginal replication (FIG.72). [1204] Weight loss was graphed as % body weight on the observation days (days post challenge) compared to weight taken just prior to challenge. Overall loss of body weight was observed in the mice that received the untranslatable control dose. Mice that received suboptimal does of BNT163 or a combination of either Het 23 or Het 15+17 with a suboptimal dose (10 ng) of BNT163 experienced improved weight loss outcome (FIG.69). [1205] A suboptimal dose (10 ng) of BNT163 did not protect mice from vaginal disease and mice that received only a suboptimal dose (10 ng) of BNT163 had multiple disease days (FIG.71) Combination of either Het 23 or Het 15+17 with a suboptimal (10ng) or optimal dose (100 ng) of BNT163 conferred full protection against disease. [1206] Viral replication kinetics are shown in FIG.72. Day 2 and Day 4 post-challenge showed the highest replication of virus, compared to 6 hours and 7 days post-challenge. Most virus was cleared by day 7 (FIG. 72D). While a suboptimal dose (10 ng) of BNT1643 led to HSV-2 replication in 4/5 mice with titers similar to the negative control (“S-TCS”) at Day 4, addition of Het 23 or Het 15+17 with BNT163 (10 ng) significantly reduced mean viral titers. Furthermore, addition of Het 23 led to complete clearance in 3/5 mice at Day 2 (FIG.72B). [1207] A positive trend in cumulative survival was observed in mice that received a combination of BNT163 (10ng) and Het 23 or Het 15 + 17.) (FIG.70). [1208] Together, these results suggest that combination of suboptimal dose of BNT163 with either Het 23 or Het 15+ Het 17 fully protects against vaginal disease, increases number of viral-cleared mice, and overall increases survivability. Example 42: In Vitro Testing of Further Polyribonucleotides Encoding Exemplary HSV-2 Antigens [1209] The present Example further demonstrates that constructs provided herein are able to express and
Figure imgf000474_0001
secrete HSV-2 immunogenic fragments from cell. In particular, the present Example shows that high transfection rates, expression levels, and secretion levels were observed in HEK293T cells transfected with exemplary nucleoside- modified mRNA (modRNAs) encoding exemplary gC, gD, or gE immunogenic fragment variants. [1210] In the present Example, the following modRNAs were transfected into HEK293T cells: - For the exemplary gC2 antigen immunogenic fragment: construct 1600; construct 3233; construct 1873; construct 1876; construct 2140; construct 2537; construct 2538; construct 2541; construct 2547; construct 2138; construct 2141; construct 2539; construct 2540; construct 2546; construct 2548; construct 2784; construct 2785; construct 2787; construct 2542; construct 2786; construct 3215; construct 3216; and construct 3217. - For the exemplary gD2 antigen immunogenic fragment: construct 1601; construct 1874; construct 1877; construct 1659; and construct 3234. - For the exemplary gE2 antigen immunogenic fragment: construct 1602; construct 1599; construct 3235; construct 1911; construct 1913; construct 2553; construct 2143; construct 2552; construct 2554; construct 2788; construct 2790; construct 2791; construct 2792; construct 4069; and construct 4070. Sequences for the above constructs are included in Table 15. [1211] FIG.73 shows expression levels in HEK293T cells transfected with RNA encoding HSV-2 gC (gC2), gD (gD2) and gE (gE2) antigens. Cells were transfected with either 0.2 μg/mL modRNA encoding gC2 or gD2 antigen constructs, or 0.4μg/mL modRNA encoding gE2 antigen constructs, using a commercial transfection REAGENT (FIG. 73A, FIG.73B AND FIG.73C), or with LNP formulated RNA encoding for a combination of all three antigens in a mass ratio of 1:1:1 (FIG.73D, FIG.73E and FIG.73F; concentrations depicted). Expression of gC2, gD2 and gE2 protein was detected by flow cytometry using primary monoclonal mouse antibodies detecting the respective antigen and a secondary fluorescent tagged anti-mouse antibody. Representative data from one experiment showing median fluorescence intensities (MFI) of the total HEK293T population for gC2 antigen constructs (FIG.73A AND FIG. 73D), for gD2 antigen constructs (FIG.73B and 73E), and for gE2 antigen constructs (FIG.73C and 73F). Data shown are mean+SD of HEK293T transfections performed in triplicates. [1212] FIG.74 shows expression levels in HEK293T cells transfected with RNA encoding HSV-2 gC (gC2) antigens. Cells were transfected with 0.2 μg/mL modRNA encoding gC2 antigen constructs using a commercial transfection reagent. Expression of gC2 proteins was detected by flow cytometry using primary monoclonal mouse antibodies detecting the respective antigen and a secondary fluorescent tagged anti-mouse antibody. Representative data from one experiment showing median fluorescence intensities (MFI) of the total HEK293T population for gC2 antigen constructs. Data shown are mean+SD of HEK293T transfections performed in triplicates. [1213] FIG.75 shows expression levels in HEK293T cells transfected with RNA encoding HSV-2 gD (gD2) antigens. Cells were transfected with 0.2 μg/mL modRNA encoding gD2 antigen constructs using a commercial transfection reagent. Expression of gD2 proteins was detected by flow cytometry using primary monoclonal mouse antibodies detecting the respective antigen and a secondary fluorescent tagged anti-mouse antibody. Representative data from one experiment showing median fluorescence intensities (MFI) of the total HEK293T population for gD2 antigen constructs. Data shown are mean+SD of HEK293T transfections performed in triplicates. [1214] FIG.76 shows expression levels in HEK293T cells transfected with RNA encoding HSV-2 gE (gE2) antigens. Cells were transfected with 0.4 μg/mL modRNA encoding gE2 antigen constructs using a commercial transfection reagent. Expression of gE2 proteins was detected by flow cytometry using a primary monoclonal mouse antibody detecting the respective antigen and a secondary fluorescent tagged anti-mouse antibody. Representative data from one experiment showing median fluorescence intensities (MFI) of the total HEK293T population for gE2 antigen constructs. Data shown are mean+SD of HEK293T transfections performed in triplicates. [1215] FIG.77 shows secretion levels of HEK293T cells transfected with RNA encoding HSV-2 gC (gC2), gD (gD2) and gE (gE2) antigens. Cells were transfected with 0.2 μg/mL modRNA encoding gC2 or gD2 antigen constructs, and 0.4μg/mL modRNA encoding gE2 antigen constructs, using a commercial transfection reagent. Secretion of gC2, gD2 and gE2 antigens was detected in the cell culture supernatants by ELISA using antibodies detecting the respective antigen. Data showing ΔOD (450-620nm) values for gC2 antigen constructs (A), for gD2 antigen constructs (B) , and for gE2 antigen constructs (C). Data shown are mean+SD of HEK293T transfected cell culture supernatants performed in triplicates. Example 43: Methods For Testing Therapeutic Efficacy of Compositions Comprising Exemplary HSV-2 Glycoproteins
Figure imgf000475_0001
[1216] Guinea pigs were infected intravaginally with HSV-2 at 2x105 PFU. The animals were scored daily for genital disease for 28 days, and ranked for disease severity on day 29 based on the number of days with genital lesions. The animals were then assigned to immunization groups using a random table to ensure equal distribution by disease severity in each group. The animals were immunized on days 35 and 65. The animals were scored for genital lesions daily Monday to Friday from day 36-116 (11 weeks). [1217] Scoring For Genital Lesions: A lesion is defined as a red area of approximately 5-10 mm with a white “pustule” in the middle. The scoring scheme is: score 0 for no lesions, 1 for one or more lesions. In contrast, during the acute (primary) infection, many lesions are present, and the scoring scheme is: 0 for no lesions, 1 for one or two lesions, 2 for more than 2 lesions; 3 if the lesions are confluent; and 4 if the lesion is necrotic. Production of RNA, formulation in LNPs [1218] Methods of constructing RNA constructs and formulation in lipid nanoparticles (LNPs) are generally known in the art (see e.g. WO 2019/035066, incorporated herein by reference in its entirety). In some embodiments, to generate nucleoside-modified RNA, m1Ψ-5′-triphosphate (TriLink) was used instead of UTP. RNA was capped using the m7G capping kit with 2′-O-methyltransferase (ScriptCap, CellScript). The RNA contains a 101 nucleotide poly(A) tail. Nucleoside-modified RNAs were purified by Fast Protein Liquid Chromatography (FPLC) (Akta Purifier GE Healthcare) and stored at −20°C. FPLC nucleoside-modified RNAs and polyC RNA (Sigma) were encapsulated in LNPs using a self-assembly process in which an aqueous solution of RNA at acidic pH 4.0 was rapidly mixed with a solution of lipids dissolved in ethanol. LNPs used in this study were similar in composition to those described previously which contain an ionizable cationic lipid/phosphatidylcholine/cholesterol/PEG-lipid (50:10:38.5:1.5 mol/mol) and were encapsulated at an RNA to total lipid ratio of ~0.05 (wt/wt). The LNPs had a diameter of ~80 nm as measured by dynamic light scattering using a Zetasizer Nano ZS (Malvern Instruments Ltd, Malvern, UK) instrument, and were stored at -80°C at a concentration of RNA of ~1 μg/μl. Example 44: Therapeutic Efficacy of Composition s Comprising Exemplary HSV-2 Glycoproteins [1219] Hartley strain guinea pigs were infected intravaginally with 2x105 PFU HSV-2, randomized into 5 equal groups of 12 animals each based on the severity
Figure imgf000476_0001
ons up to the time of randomization (on day 28 post-infection). Animals were then immunized (n=12/group) on days 35 and 65 post infection (30 μg RNA/immunization) with the following nucleoside-modified RNAs encapsulated in lipid nanoparticle 315: [1220] (i) gB2 alone (gB2 group): Modified RNA encoding an N-terminal IL-2 secretory signal peptide and an exemplary immunogenic fragment of HSV-2 glycoprotein B (gB2) truncated prior to the transmembrane domain was encapsulated in lipid nanoparticles (LNP) 315 (provided by Acuitas, Vancouver, Canada). The LNP- encapsulated modified RNA was administered at 30 μg gB2 at each immunization; or [1221] (ii) PBS was used as a control. [1222] Starting one day after the first immunization, the animals were scored daily for genital lesions Monday to Friday from day 36 post-infection until day 116 post-infection (59 days). The results are summarized below. The number of days animals in each group had recurrent genital lesions starting from 1 day after the first immunization (day 36) or starting from 1 day after the second immunization (day 66) were reported. One day after the second immunization was considered as the primary endpoint for the study. [1223] Composition efficacy as determined by reduction in recurrent genital lesions in guinea pigs is shown in Table 44. The cumulative recurrent genital lesion days per group are shown in FIG.78. FIG.79 shows the results for each guinea pig in the study plotting the number of recurrent genital lesions from 1 day after the second immunization until the end of the study on day 116. Table 44: Exemplary HSV Immunogenic Fragment RNA Compositions as Immunotherapy for Recurrent Genital Lesions in Guinea Pigs 1 day after 1st dose 1 day after 2nd dose Immunogens L i n r 708 C m iti n ffi L i n r 444 C m iti n ffi cy
Figure imgf000476_0002
[1224] Conclusions: Administration of LNP-encapsulated RNA encoding exemplary immunogenic fragments of HSV-2 gB reduced the number of recurrent genital lesions compared to animals that were administered PBS control. Administration of LNP-encapsulated RNA encoding exemplary immunogenic fragments of HSV-2 gB reduced recurrent genital lesions by 81% (composition efficacy = 81%) compared to the control group (PBS). [1225] These results suggest that LNP-encapsulated RNA encoding exemplary immunogenic fragments of HSV-2 gB either alone or in combination with other HSV glycoproteins, or immunogenic fragments thereof, or other HSV immunogens may be effective for treating genital herpes in already infected individuals. Example 45: Evaluation of T-cell responses to gB2 immunization [1226] Objective: To evaluate the response of T cells in mice immunized with gB2 mRNA.. [1227] Methods: Mice were immunized twice with 10 μg gB2 mRNA-LNP. Splenocytes from these mice were stimulated with a gB2 overlapping peptide pool. CD4+ T-cells and CD8+ T cells producing cytokines were analyzed by flow cytometry. [1228] Results: gB2 immunization induces single and polyfunctional cytokine CD4+ (FIG.80) and CD8+ (FIG. 81) T-cell activation. Example 46: In vitro Testing of Polyribonucleotides Encoding Exemplary HSV-2 gB Antigens [1229] The resent Exam le demonstrates that constructs rovided herein are able to express a gB
Figure imgf000477_0001
g . p , p p g p ved in HEK293T cells transfected with exemplary polyribonucleotide constructs encoding gB immunogenic fragment variants. [1230] In the present Example, the following modRNAs were transfected into HEK293T cells: construct 4059; construct 4060; construct 4061; construct 4063; a construct 4064; construct 4065. Sequences for these constructs are included in Table 15. [1231] FIG.82 shows expression levels in HEK293T cells transfected with RNA encoding HSV-2 gB (gB2) antigens. Cells were transfected with 0.2 μg/mL modRNA encoding gB2 antigen constructs using a commercial transfection reagent. Expression of gB2 proteins was detected by flow cytometry using primary monoclonal mouse antibodies detecting the respective antigen and a secondary fluorescent tagged anti-mouse antibody. Representative data from one experiment showing median fluorescence intensities (MFI) of the total HEK293T population for gB2 antigen constructs. Data shown are mean+SD of HEK293T transfections performed in triplicates. As shown, significant expression was detected from all six tested RNA constructs. Example 47: Methods for Testing Therapeutic Efficacy of Compositions Comprising Exemplary Immunogenic Fragments of HSV-2 Glycoproteins E and I [1232] Guinea pigs were infected intravaginally with HSV-2 at 2x105 PFU. The animals were scored daily for genital disease for 28 days and ranked for disease severity on day 29 based on the number of days with genital lesions. The animals were then assigned to immunization groups using a random table to ensure equal distribution by disease severity in each group. The animals were immunized on days 35 and 65. The animals were scored for genital lesions daily Monday to Friday from day 36-116 (11 weeks). Scoring For Genital Lesions: A lesion is defined as a red area of approximately 5-10 mm with a white “pustule” in the middle. The scoring criteria is: score 0 for no lesions, 1 for one or more lesions. In contrast, during the acute (primary) infection, many lesions are present, and the scoring scheme is: 0 for no lesions, 1 for one or two lesions, 2 for more than 2 lesions; 3 if the lesions are confluent; and 4 if the lesion is necrotic. Production of RNA, formulation in LNPs [1233] Methods of constructing exemplary RNA constructs and formulation in lipid nanoparticles (LNPs) are generally known in the art (see e.g. WO 2019/035066, incorporated herein by reference in its entirety). In some embodiments, to generate nucleoside-modified RNA, m1Ψ-5′-triphosphate (TriLink) was used instead of UTP. RNA was capped using the m7G capping kit with 2′-O-methyltransferase (ScriptCap, CellScript). The RNA contains a 101- nucleotide polyadenylate (poly(A)) tail, which in this case, is an uninterrupted sequence of adenylate residues located at the 3'-end of the RNA polynucleotides. Nucleoside-modified RNAs were purified by Fast Protein Liquid Chromatography (FPLC) (Akta Purifier GE Healthcare) and stored at −20°C. FPLC nucleoside-modified RNAs and polyC RNA (Sigma) were encapsulated in LNPs using a self-assembly process in which an aqueous solution of RNA at acidic pH 4.0 was rapidly mixed with a solution of lipids dissolved in ethanol. LNPs used in this study were similar in composition to those described previously which contain an ionizable cationic lipid/phosphatidylcholine/cholesterol/PEG-lipid (50:10:38.5:1.5 mol/mol) and were encapsulated at an RNA to total lipid ratio of ~0.05 (wt/wt). The LNPs had a diameter of ~80 nm as measured by dynamic light scattering using a Zetasizer Nano ZS (Malvern Instruments Ltd, Malvern, UK) instrument, and were stored at -80°C at a concentration of RNA of ~1 μg/μl. Ex m l 48 Th r ti Effi f C m iti n C m ri in Ex m l r Imm n ni Fr m nts of
Figure imgf000478_0001
ps of 12 animals each based on the severity of genital lesions up to the time of randomization (on day 28 post-infection). Animals were then immunized (n=12/group) on days 35 and 65 post infection with: (i) RNA encoding exemplary immunogenic fragments of HSV-2 glycoproteins E and I (gE2/gI2) both truncated prior to the transmembrane domain; or (ii) PBS. [1235] Exemplary immunogenic fragments of HSV-2 glycoproteins E and I (gE2/gI2) encoded by RNAs were encapsulated in a lipid nanoparticle (LNP). The human IL-2 signal peptide was used in conjunction with the exemplary gE2 immunogenic fragment, and the MHC-II/HLA-DR signal peptide was used in conjunction with the exemplary gI2 immunogenic fragment. RNA encoding the exemplary immunogenic fragments of gE2 and gI2 were encapsulated in a lipid nanoparticle. [1236] Starting one day after the first immunization, animals were scored daily for genital lesions Monday to Friday from day 36 post-infection until day 116 post-infection (59 days). The results are summarized below. The number of days animals in each group had recurrent genital lesions starting from 1 day after the first immunization (day 36) or starting from 1 day after the second immunization (day 66) were reported. One day after the second immunization was considered as the primary endpoint for the study. [1237] Composition efficacy as determined by reduction in recurrent genital lesions in guinea pigs is shown in Table 45. The cumulative recurrent genital lesion days per group are shown in FIG.83A. FIG.83B shows the results for each guinea pig in the study plotting the number of recurrent genital lesions from 1 day after the second immunization until the end of the study on day 116. Table 45: Cumulative recurrent genital lesion days per group 1 day after 1st dose 1 day after 2nd dose Immunogen Lesions over 708 Composition efficacy Lesions over 444 Composition efficacy =12/
Figure imgf000479_0004
p g g p y u g g gE2 and gI2 reduced the number of recurrent genital lesions compared to animals that were administered PBS control. Administration of the LNP comprising RNA encoding exemplary immunogenic fragments of gE2 and gI2 reduced recurrent genital lesions by 81% (Composition efficacy = 81%) compared to the control group (PBS). Example 49: T-cell responses to gE2 immunization with exemplary immunogenic fragments of HSV-2 glycoproteins E and I g gE2/gI2
Figure imgf000479_0001
N - N or 0μg oy(C) N - N as a contro. en days a ter t e second mmunzaton, speens were arvested from a subset of animals and evaluated for the presence of antigen specific CD4+ and CD8+ T-cell responses to gE2 and gI2 by stimulation of splenocytes with a gE2 or gI2 overlapping peptide pool. CD4+ and CD8+ T-cells producing cytokines were analyzed by flow cytometry. [1240] Results: Mice immunized with gE2/gI2 bivalent RNA Composition generated strong CD4+ (FIG.84A) and CD8+ (FIG.84B) T-cell responses to gE2 peptides and minimal CD4+ T-cell responses to gI2 peptides (FIG.84C). No CD8+ T-cell responses to gI2 peptides were observed (FIG.84D). Example 50: Antibody responses after immunization with exemplary immunogenic fragments of HSV-2 glycoproteins E and I
Figure imgf000479_0002
[1241] Methods: Female BALB/c mice were immunized intramuscularly (IM) twice 1-month apart with 10μg gE2/gI2 RNA-LNP or 10μg Poly(C) RNA-LNP as a control. Sera were collected to evaluate antibody responses (endpoint titers) to gE2/gI2, gE2, and gI2 four weeks after the second immunization. [1242] Results: The composition formulations produced high IgG ELISA endpoint titers to gE2/gI2 (FIGS. 85 AND 86A) and moderate gI2 endpoint titers compared to controls (FIG.86B). Example 51: Prophylactic efficacy of immunization with exemplary immunogenic fragments of HSV-2 glycoproteins E and I
Figure imgf000479_0003
[1243] Female BALB/c mice were immunized intramuscularly (IM) twice 1-month apart with 10μg gE2/gI2 RNA-LNP or 10μg Poly(C) RNA-LNP as a control. The mice were then challenged intravaginally with HSV-2 strain MS (5×103 PFU (275 LD50)) and monitored for signs of clinical disease and subclinical infection including survival, body weight, and genital disease and vaginal virus titers days 2 and 4 post-infection and HSV-2 DNA copy number in DRG (FIG. 87). [1244] Methods: [1245] Scoring For Genital Lesions: A lesion is defined as a red area of approximately 5-10 mm with a white “pustule” in the middle. The scoring scheme is: score 0 for no lesions, 1 for one or more lesions. In contrast, during the acute (primary) infection, many lesions are present, and the scoring scheme is: 0 for no lesions, 1 for one or two lesions, 2 for more than 2 lesions; 3 if the lesions are confluent; and 4 if the lesion is necrotic. [1246] 28 days following the second immunization, mice were challenged intravaginally with HSV-2 and monitored for 28 days for signs of clinical disease including survival, body weight, and genital disease. Animals were evaluated daily for health and signs of disease. Animals were euthanized upon reaching humane endpoints. [1247] Mice were also monitored for signs of subclinical infection including vaginal virus titers days 2 and 4 post- infection and HSV-2 DNA copy number in DRG as determined by qPCR 28 days post-infection or at the time of humane euthanasia. Vaginal virus titers in mice were evaluated days 2 and 4 post-infection by gently swabbing the vaginal cavity and determining the amount of HSV-2 virus present by standard plaque assay on Vero cells. [1248] Sera collected four weeks after the second immunization were evaluated for the presence of antibodies to gE2 or gI2 that could bind to purified gE2/gI2 protein and prevent it from binding the Fc domain of human IgG1, thereby blocking its immune evasive function. Briefly, serial two-fold dilutions of mouse sera were incubated with gE2/gI2 and then added to wells of an ELISA plate that had been coated with human IgG1. The sera dilution that resulted in a 50% reduction in gE2/gI2 binding compared to control was plotted. [1249] Results: By nearly all measures, mice receiving gE2/gI2 RNA-LNP had significantly less clinical disease (FIG. 88A), weight loss (FIG.88B), or genital disease (FIG.88C) compared with control animals. [1250] Mice receiving gE2/gI2 RNA-LNP also had significantly less virus present within their vaginal cavities on days 2 and 4 compared with control animals (FIGS.89A and 89B). In addition, mice receiving gE2/gI2 RNA-LNP had significantly less HSV-2 DNA copy numbers in DRG (FIG.89C). [1251] Administration of exemplary HSV-2 gE/gI RNA-LNP as described herein generates high IgG titers to gE2/gI2 (FIG.86A). HSV-2 gE/gI RNA-LNP also significantly reduced clinical disease and subclinical infection compared to control animals. Example 52: Prophylactic composition trial in guinea pigs [1252] Objective: To determine the efficacy of BNT163 alone and in combination with gB2 or gI2 as a prophylactic composition (e.g., immunogenic composition, e.g., vaccine) for HSV-2 [1253] Methods: Guinea pigs were immunized intramuscularly (IM) twice 1-month apart with PBS, BNT163 (30μg), gE2/gI2 (15μg each), gB2 (30μg), BNT163 (22.5μg) + gB2 or gI2 (7.5μg). Experimental groups comprised 10 animals, and PBS groups comprised 9 animals. The guinea pigs were then challenged intravaginally with HSV-2 strain MS at 5x105 PFU (20 LD50) and monitored daily through day 46 days. Monitoring included determining disease severity, severity of genital lesions, urinary retention, and survival rates. [1254] BNT163 is an RNA composition comprising polyribonucleotides encoding truncated HSV-2 glycoproteins C, D, E (gC2/gD2/gE2) encapsulated in a lipid nanoparticle. The constructs are truncated prior to the transmembrane domain. and have the following signal sequences: gC2 and gE2 have an IL-2-like signal sequence, and gD2 has a gD2 signal sequence. Results: [1255] Animals that received BNT163, gE2/gI2, gB2 or a combination of BNT163 and gB2 or gI2 had an increased probability of survival, compared with animals that received PBS (FIG.91A). Further, the severity of the disease was decreased in animals receiving either BNT163, gE2/gI2, or a combination of BNT163 and gI2. Only one animal from the BNT163 group died, and 2 animals from the gB2 group died. [1256] All treatments groups showed decreased disease severity compared with control animals (FIG.91B). Animals receiving only BNT163 had decreased severity compared to animals receiving either gB2, gE2/gI2, a combination of BNT163 and gB2, or a combination of BNT163 and gI2. Animals receiving a combination of BNT163 and gB2 or gI2 had decreased severity compared to animals receiving gB2 or gE2/gI2 (FIG.91B). These results indicate that adding immunogens to BNT163 alters composition (e.g., immunogenic composition, e.g., vaccine) efficacy if BNT163 concentration is lowered. [1257] Animals receiving only BNT163 had decreased severity of genital lesions compared to control animals, animals receiving either gE2/gI2, or a combination of BNT163 and gB2 or gI2. Animals receiving a combination of BNT163 and gB2 or gI2 had decreased severity of genital lesions compared to animals receiving gE2/gI2 (FIG. 91C). One animal that received only BNT163 died despite having no genital lesions (indicated by black symbol) (FIG.100C). These results indicated that animals receiving BNT163 died despite having no genital lesions. [1258] All treatment groups showed decreased days with urinary retention compared to control animals (FIG. 91D). The BNT163 animal that died had 9 days of urinary retention. 4 out of 10 BNT163 animals had no lesions and no urinary retention. [1259] In conclusion, animals receiving BNT163 at 30μg outperformed other groups for protection against genital lesions than groups that used BNT163 at 22.5μg combined with gB2 or gI2. Example 53: Therapeutic Composition trial in guinea pigs [1260] Objective: To determine the efficacy of BNT163 alone and in combination with gB2 or gI2 as a therapeutic composition (e.g., immunogenic composition, e.g., vaccine) for HSV-2. [1261] Methods: Guinea pigs were then challenged intravaginally with HSV-2 strain MS strain 1991 (4x105 PFU (<1 LD50). 35 and 65 days after infection, animals were immunized intramuscularly (IM) with PBS, BNT163 (30μg), gE2/gI2 (15μg each), BNT163 (22.5μg) + gB2 or gI2 (7.5μg). Treatment groups comprised between 9-12 animals. Animals were monitored until day 116 for disease severity. [1262] Results: [1263] Tables 46-50 show the days with recurrent lesions in each group. Table 46 shows results from a first experiment. Table 47 shows results from a second experiment. Table 48 shows results from a combination of the first and the second experiments. Table 46: Experiment 1 Groups Mean lesion days after 1st immunization Mean lesion days after 2nd immunization to end to end n
Figure imgf000482_0001
Table 47: Experiment 2 Groups Mean lesion days after 1st Mean lesion days after 2nd n
Figure imgf000482_0002
PBS (n=12) 67/708 (9.5%) 5.6 ----- 43/444 (9.7%) 3.6 -----
Figure imgf000483_0001
TABLE 48: Combined data from Experiment 1 and Experiment 2 Groups Mean lesion days after 1st Mean lesion days after 2nd immunization to end immunization to end n
Figure imgf000483_0002
t difference in recurrence of lesions after the second dose compared with animals that received PBS (FIG.92A). Table 49 shows the number and percentage of animals that were recurrence free after the second dose in Experiment 2. [1265] TABLE 49: Experiment 2 data PBS gB2 gE2/gI2 BNT163 , animals receiving gB2, but not gE2/gI2, and or BNT163, had
Figure imgf000484_0001
e compared with animals that received PBS (FIG.92B). Table 50 shows the number and percentage of animals that were recurrence free after the second dose. [1267] TABLE 50: Experiment Data PBS gB2 gE2/gI2 BNT163 6/22 11/22 9/22 11/22 MBODIMENTS
Figure imgf000484_0002
[1268] Embodiment 1. A combination comprising a plurality of polyribonucleotides, wherein the plurality of polyribonucleotides comprises a first set of polyribonucleotides that encode one or more glycoprotein (GP) polypeptides, wherein the one or more GP polypeptides each comprise an HSV glycoprotein or an antigenic portion thereof. [1269] Embodiment 2. The combination of embodiment 1, wherein the first set of polyribonucleotides comprises: (i) a polyribonucleotide encoding an HSV glycoprotein C (gC) or an antigenic portion thereof, (ii) a polyribonucleotide encoding an HSV glycoprotein D (gD) or an antigenic portion thereof, (iii) a polyribonucleotide encoding an HSV glycoprotein E (gE) or an antigenic portion thereof, or (iv) a combination thereof. [1270] Embodiment 3. The combination of embodiment 1 or 2, wherein the first set of polyribonucleotides comprises: (i) a polyribonucleotide that encoding an antigenic portion of HSV gC, (ii) a polyribonucleotide that encoding an antigenic portion of HSV gD, (iii) a polyribonucleotide that encoding an antigenic portion of HSV gE, or (iv) a combination thereof. [1271] Embodiment 4. The combination of any one of embodiments 1-3, wherein the first set of polyribonucleotides comprises: (i) a polyribonucleotide that encoding an antigenic portion of HSV gC, (ii) a polyribonucleotide that encoding an antigenic portion of HSV gD, and (iii) a polyribonucleotide that encoding an antigenic portion of HSV gE. [1272] Embodiment 5. The combination of any one of embodiments 2-4, wherein the antigenic portion of HSV gC comprises or consists of an amino acid sequence according to any one of SEQ ID NO: 1 or 260. [1273] Embodiment 6. The combination of any one of embodiments 2-5, wherein the polyribonucleotide encoding the antigenic portion of HSV gC comprises a ribonucleic acid sequence according to SEQ ID NO: 16-19, 174, 274-281, 290-291, or 336. [1274] Embodiment 7. The combination of any one of embodiments 2-6, wherein the antigenic portion of HSV gD comprises or consists of an amino acid sequence according to SEQ ID NO: 2. [1275] Embodiment 8. The combination of any one of embodiments 2-7, wherein the polyribonucleotide encoding the antigenic portion of HSV gD comprises a ribonucleic acid sequence according to SEQ ID NO: 20-23, 143, 286 or 340. [1276] Embodiment 9. The combination of any one of embodiments 2-8, wherein the antigenic portion of HSV gE comprises or consists of an amino acid sequence according to SEQ ID NO: 3. [1277] Embodiment 10. The combination of any one of embodiments 2-9, wherein the polyribonucleotide encoding the antigenic portion of HSV gE comprises a ribonucleic acid sequence according to SEQ ID NO: 24-27, 149, 282-285, or 341. [1278] Embodiment 11. The combination of any one of embodiments 1-10, wherein the first set of polyribonucleotides further comprises at least one polyribonucleotide encoding a GP polypeptide that comprises an HSV glycoprotein B (gB), variant thereof, or one or more antigenic portions thereof. [1279] Embodiment 12. The combination of embodiment 11, wherein the HSV gB comprises or consists of an amino acid sequence with at least 80%, such as at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to an amino acid sequence according to SEQ ID NO: 379-384. [1280] Embodiment 13. The combination of embodiment 11 or 12, wherein the HSV gB comprises one or more mutations that stabilize the HSV gB relative to a comparable HSV gB that does not comprise the one or more mutations. [1281] Embodiment 14. The combination of embodiment 13, wherein the one or more mutations are one or more amino acid substitutions. [1282] Embodiment 15. The combination of embodiment 14, wherein the one or more amino acid substitutions comprise 120C, 181C, 238C, 251C, 259C, 290C, 291C, 391C, 526C, 571C, 610C, 630C, 636C, 676C, 677C, 680C, 714C, 718C, 725C, 758C, and combinations thereof, wherein the numbering is with reference to SEQ ID NO: 379. [1283] Embodiment 16. The combination of any one of embodiments 13-15, wherein the one or more mutations comprise: (i) 120C and 677C; (ii) 181C and 725C; (iii) 238C and 610C; (iv) 251C and 718C; (v) 259C and 758C; (vi) 290C and 680C; (vii) 291C and 636C; (viii) 391C and 526C; (ix) 571C and 676C; (x) 571C and 680C; and/or (xi) 630C and 714C; wherein the numbering is with reference to SEQ ID NO: 379. [1284] Embodiment 17. The combination of any one of embodiments 1-16, wherein the first set of polyribonucleotides further comprises: (i) a polyribonucleotide encoding an HSV glycoprotein G (gG) or an antigenic portion thereof, (ii) a polyribonucleotide encoding an HSV glycoprotein H (gH) or an antigenic portion thereof, (iii) a polyribonucleotide encoding an HSV glycoprotein I (gI) or an antigenic portion thereof, (iv) a polyribonucleotide encoding an HSV glycoprotein L (gL) or an antigenic portion thereof, (v) a combination thereof. [1285] Embodiment 18. The combination of any one of embodiments 1-17, wherein the combination further comprises a second set of polyribonucleotides that encode one or more T-cell string polypeptides, wherein the one or more T-cell string polypeptides each comprise one or more HSV T-cell antigens or antigenic portions thereof. [1286] Embodiment 19. The combination of embodiment 18, wherein the second set of polyribonucleotides encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof, wherein the one or more HSV T-cell antigens comprises: (i) one or more HSV RS1 polypeptides or antigenic portions thereof, (ii) one or more HSV RL2 polypeptides or antigenic portions thereof, (iii) one or more HSV UL1 polypeptides or antigenic portions thereof, (iv) one or more HSV UL5 polypeptides or antigenic portions thereof, (v) one or more HSV UL9 polypeptides or antigenic portions thereof, (vi) one or more HSV UL19 polypeptides or antigenic portions thereof, (vii) one or more HSV UL21 polypeptides or antigenic portions thereof, (viii) one or more HSV UL25 polypeptides or antigenic portions thereof, (ix) one or more HSV UL27 polypeptides or antigenic portions thereof, (x) one or more HSV UL29 polypeptides or antigenic portions thereof, (xi) one or more HSV UL30 polypeptides or antigenic portions thereof, (xii) one or more HSV UL39 polypeptides or antigenic portions thereof, (xiii) one or more HSV UL40 polypeptides or antigenic portions thereof, (xiv) one or more HSV UL46 polypeptides or antigenic portions thereof, (xv) one or more HSV UL47 polypeptides or antigenic portions thereof, (xvi) one or more HSV UL48 polypeptides or antigenic portions thereof, (xvii) one or more HSV UL49 polypeptides or antigenic portions thereof, (xviii) one or more HSV UL52 polypeptides or antigenic portions thereof, (xix) one or more HSV UL54 polypeptides or antigenic portions thereof, (xx) one or more HSV US10 polypeptides or antigenic portions thereof, (xxi) one or more HSV US12 polypeptides or antigenic portions thereof, (xxii) one or more HSV UL26 polypeptides or antigenic portions thereof, (xxiii) one or more HSV UL50 polypeptides or antigenic portions thereof, or (xxiv) a combination thereof. [1287] Embodiment 20. The combination of embodiment 18 or 19, wherein the second set of polyribonucleotides comprises a polyribonucleotide that encodes a polypeptide that comprises one or more HSV RL2 polypeptides or antigenic portions thereof, one or more HSV RS1 polypeptides or antigenic portions thereof, and one or more HSV UL54 polypeptides or antigenic portions thereof. [1288] Embodiment 21. The combination of any one of embodiments 18-20, wherein the second set of polyribonucleotides comprises a polyribonucleotide that encodes a polypeptide that comprises, in N-terminus to C- terminus order, an RL2 polypeptide or antigenic fragment thereof, a linker, an RL2 polypeptide or antigenic fragment thereof, a linker, an RS1 polypeptide or antigenic fragment thereof, a linker, a UL54 polypeptide or antigenic fragment thereof, and a linker. [1289] Embodiment 22. The combination of any one of embodiments 18-20, wherein the second set of polyribonucleotides comprises a polyribonucleotide that encodes a polypeptide that comprises, in N-terminus to C- terminus order, an UL54 polypeptide or antigenic fragment thereof, a linker, an RS1 polypeptide or antigenic fragment thereof, a linker, an RL2 polypeptide or antigenic fragment thereof, a linker, a RL2 polypeptide or antigenic fragment thereof, and a linker. [1290] Embodiment 23. The combination of embodiment 18 or 19, wherein the second set of polyribonucleotides comprises a polyribonucleotide that encodes a polypeptide that comprises one or more HSV UL29 polypeptides or antigenic portions thereof, one or more HSV UL39 polypeptides or antigenic portions thereof, one or more HSV UL49 polypeptides or antigenic portions thereof, and one or more HSV UL9 polypeptides or antigenic portions thereof. [1291] Embodiment 24. The combination of any one of embodiments 18, 19, and 23, wherein the second set of polyribonucleotides comprises a polyribonucleotide that encodes a polypeptide that comprises, in N-terminus to C-terminus order, an UL29 polypeptide or antigenic fragment thereof, a linker, an UL39 polypeptide or antigenic fragment thereof, a linker, an UL49 polypeptide or antigenic fragment thereof, a linker, a UL9 polypeptide or antigenic fragment thereof, and a linker. [1292] Embodiment 25. The combination of any one of embodiments 18, 19, and 23, wherein the second set of polyribonucleotides comprises a polyribonucleotide that encodes a polypeptide that comprises, in N-terminus to C-terminus order, an UL9 polypeptide or antigenic fragment thereof, a linker, an UL49 polypeptide or antigenic fragment thereof, a linker, an UL39 polypeptide or antigenic fragment thereof, a linker, a UL29 polypeptide or antigenic fragment thereof, and a linker. [1293] Embodiment 26. The combination of embodiment 18 or 19, wherein the second set of polyribonucleotides comprises a polyribonucleotide that encodes a polypeptide that comprises one or more HSV UL30 polypeptides or antigenic portions thereof, one or more HSV UL40 polypeptides or antigenic portions thereof, one or more HSV UL5 polypeptides or antigenic portions thereof, and one or more HSV UL52 polypeptides or antigenic portions thereof. [1294] Embodiment 27. The combination of any one of embodiments 18, 19, and 26, wherein the second set of polyribonucleotides comprises a polyribonucleotide that encodes a polypeptide that comprises, in N-terminus to C-terminus order, an UL30 polypeptide or antigenic fragment thereof, a linker, an UL30 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, a UL5 polypeptide or antigenic fragment thereof, a linker, a UL5 polypeptide or antigenic fragment thereof, a linker, a UL52 polypeptide or antigenic fragment thereof, and a linker. [1295] Embodiment 28. The combination of any one of embodiments 18, 19, and 26, wherein the second set of polyribonucleotides comprises a polyribonucleotide that encodes a polypeptide that comprises, in N-terminus to C-terminus order, an UL52 polypeptide or antigenic fragment thereof, a linker, an UL5 polypeptide or antigenic fragment thereof, a linker, an UL5 polypeptide or antigenic fragment thereof, a linker, a UL40 polypeptide or antigenic fragment thereof, a linker, a UL30 polypeptide or antigenic fragment thereof, a linker, a UL30 polypeptide or antigenic fragment thereof, and a linker. [1296] Embodiment 29. The combination of embodiment 18 or 19, wherein the second set of polyribonucleotides comprises a polyribonucleotide that encodes a polypeptide that comprises one or more HSV UL1 polypeptides or antigenic portions thereof, one or more HSV UL19 polypeptides or antigenic portions thereof, one or more HSV UL21 polypeptides or antigenic portions thereof, one or more HSV UL27 polypeptides or antigenic portions thereof, one or more HSV UL46 polypeptides or antigenic portions thereof, one or more HSV UL47 polypeptides or antigenic portions thereof, one or more UL48 polypeptides or antigenic portions thereof, and one or more HSV UL25 polypeptides or antigenic portions thereof. [1297] Embodiment 30. The combination of any one of embodiments 18, 19, and 29, wherein the second set of polyribonucleotides comprises a polyribonucleotide that encodes a polypeptide that comprises, in N-terminus to C-terminus order, an HSV UL1 polypeptide or antigenic fragment thereof, a linker, an HSV UL19 polypeptide or antigenic fragment thereof, a linker, an HSV UL21 polypeptide or antigenic fragment thereof, a linker, an HSV UL27 polypeptide or antigenic fragment thereof, a linker, an HSV UL27 polypeptide or antigenic fragment thereof, a linker, an HSV UL46 polypeptide or antigenic fragment thereof, a linker, an HSV UL47 polypeptide or antigenic fragment thereof, a linker, an HSV UL25 polypeptide or antigenic fragment thereof, a linker, an HSV UL48 polypeptide or antigenic fragment thereof, and a linker. [1298] Embodiment 31. The combination of any one of embodiments 18, 19, and 30, wherein the second set of polyribonucleotides comprises a polyribonucleotide that encodes a polypeptide that comprises, in N-terminus to C-terminus order, an HSV UL48 polypeptide or antigenic fragment thereof, a linker, an HSV UL25 polypeptide or antigenic fragment thereof, a linker, an HSV UL47 polypeptide or antigenic fragment thereof, a linker, an HSV UL46 polypeptide or antigenic fragment thereof, a linker, an HSV UL27 polypeptide or antigenic fragment thereof, a linker, an HSV UL27 polypeptide or antigenic fragment thereof, a linker, an HSV UL21 polypeptide or antigenic fragment thereof, a linker, an HSV UL19 polypeptide or antigenic fragment thereof, a linker, an HSV UL1 polypeptide or antigenic fragment thereof, and a linker. [1299] Embodiment 32. The combination of embodiment 18 or 19, wherein the second set of polyribonucleotides comprises a polyribonucleotide that encodes a polypeptide that comprises one or more HSV US1 polypeptides or antigenic fragments thereof, one or more HSV US8 polypeptides or antigenic fragments thereof, one or more HSV US12 polypeptides or antigenic fragments thereof, one or more HSV UL50 polypeptides or antigenic fragments thereof, one or more HSV UL26 polypeptides or antigenic fragments thereof, and one or more HSV US10 polypeptides or antigenic fragments thereof. [1300] Embodiment 33. The combination of any one of embodiments 18, 19, and 32, wherein the second set of polyribonucleotides comprises a polyribonucleotide that encodes a polypeptide that comprises, in N-terminus to C-terminus order, an US1 polypeptide or antigenic fragment thereof, a linker, an US1 polypeptide or antigenic fragment thereof, a linker, an US8 polypeptide or antigenic fragment thereof, a linker, a US8 polypeptide or antigenic fragment thereof, a linker, a US8 polypeptide or antigenic fragment thereof, a linker, a US12 polypeptide or antigenic fragment thereof, a linker, a UL50 polypeptide or antigenic fragment thereof, a linker, a UL26 polypeptide or antigenic fragment thereof, a linker, a UL26 polypeptide or antigenic fragment thereof, a linker, a US10 polypeptide or antigenic fragment thereof, and a linker. [1301] Embodiment 34. The combination of any one of embodiments 18, 19, and 32, wherein the second set of polyribonucleotides comprises a polyribonucleotide that encodes a polypeptide that comprises, in N-terminus to C-terminus order, an UL26 polypeptide or antigenic fragment thereof, a linker, an UL26 polypeptide or antigenic fragment thereof, a linker, an US10 polypeptide or antigenic fragment thereof, a linker, a UL50 polypeptide or antigenic fragment thereof, a linker, a US12 polypeptide or antigenic fragment thereof, a linker, a US8 polypeptide or antigenic fragment thereof, a linker, a US8 polypeptide or antigenic fragment thereof, a linker, a US8 polypeptide or antigenic fragment thereof, a linker, a US1 polypeptide or antigenic fragment thereof, a linker, a US1 polypeptide or antigenic fragment thereof, and a linker. [1302] Embodiment 35. The combination of embodiment 18 or 19, wherein the polypeptide comprises one or more HSV RL2 polypeptides or antigenic fragments thereof, one or more HSV UL54 polypeptides or antigenic fragments thereof, one or more HSV UL47 polypeptides or antigenic fragments thereof, one or more HSV UL46 polypeptides or antigenic fragments thereof, and one or more HSV UL21 polypeptides or antigenic fragments thereof. [1303] Embodiment 36. The combination of any one of embodiments 18, 19, and 35, wherein the polypeptide comprises, in N-terminus to C-terminus order, an RL2 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, a UL46 polypeptide or antigenic fragment thereof, a linker, a UL21 polypeptide or antigenic fragment thereof, and a linker. [1304] Embodiment 37. The combination of any one of embodiments 18, 19, and 35, wherein the polypeptide comprises, in N-terminus to C-terminus order, an UL21 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, an RL2 polypeptide or antigenic fragment thereof, and a linker. [1305] Embodiment 38. The combination of embodiment 18 or 19, wherein the polypeptide comprises one or more HSV UL29 polypeptides or antigenic fragments thereof, one or more HSV UL39 polypeptides or antigenic fragments thereof, one or more HSV UL9 polypeptides or antigenic fragments thereof, one or more HSV UL5 polypeptides or antigenic fragments thereof, one or more HSV UL40 polypeptides or antigenic fragments thereof, and one or more HSV UL30 polypeptides or antigenic fragments thereof. [1306] Embodiment 39. The combination of any one of embodiments 18, 19, and 38, wherein the polypeptide comprises, in N-terminus to C-terminus order, an UL29 polypeptide or antigenic fragment thereof, a linker, an UL39 polypeptide or antigenic fragment thereof, a linker, an UL9 polypeptide or antigenic fragment thereof, a linker, an UL5.1 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL30.1 polypeptide or antigenic fragment thereof, and a linker. [1307] Embodiment 40. The combination of any one of embodiments 18, 19, and 38, wherein the polypeptide comprises, in N-terminus to C-terminus order, an UL30.1 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL5.1 polypeptide or antigenic fragment thereof, a linker, an UL9 polypeptide or antigenic fragment thereof, a linker, an UL39 polypeptide or antigenic fragment thereof, a linker, an UL29 polypeptide or antigenic fragment thereof, and a linker. [1308] Embodiment 41. The combination of embodiment 18 or 19, wherein the polypeptide comprises one or more HSV RL2 polypeptides or antigenic fragments thereof, one or more HSV UL54 polypeptides or antigenic fragments thereof, one or more HSV UL9 polypeptides or antigenic fragments thereof, one or more HSV UL39 polypeptides or antigenic fragments thereof, and one or more HSV UL5 polypeptides or antigenic fragments thereof. [1309] Embodiment 42. The combination of any one of embodiments 18, 19, and 41, wherein the polypeptide comprises, in N-terminus to C-terminus order, an RL2.1 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, an UL9 polypeptide or antigenic fragment thereof, a linker, a UL39 polypeptide or antigenic fragment thereof, a linker, a UL5.1 polypeptide or antigenic fragment thereof, and a linker. [1310] Embodiment 43. The combination of any one of embodiments 18, 19, and 41, wherein the polypeptide comprises, in N-terminus to C-terminus order, an UL5.1 polypeptide or antigenic fragment thereof, a linker, an UL39 polypeptide or antigenic fragment thereof, a linker, an UL9 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, an RL2.1 polypeptide or antigenic fragment thereof, and a linker. [1311] Embodiment 44. The combination of embodiment 18 or 19, wherein the polypeptide comprises one or more HSV UL47 polypeptides or antigenic fragments thereof, one or more HSV UL46 polypeptides or antigenic fragments thereof, one or more HSV UL21 polypeptides or antigenic fragments thereof, one or more HSV UL5.2 polypeptides or antigenic fragments thereof, one or more HSV UL40 polypeptides or antigenic fragments thereof, one or more HSV UL30.1 polypeptides or antigenic fragments thereof, and one or more HSV UL29 polypeptides or antigenic fragments thereof. [1312] Embodiment 45. The combination of any one of embodiments 18, 19, and 44, wherein the polypeptide comprises, in N-terminus to C-terminus order, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL21 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL30.1 polypeptide or antigenic fragment thereof, a linker, an UL29 polypeptide or antigenic fragment thereof, and a linker. [1313] Embodiment 46. The combination of any one of embodiments 18, 19, and 44, wherein the polypeptide comprises, in N-terminus to C-terminus order, an UL29 polypeptide or antigenic fragment thereof, a linker, an UL30.1 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL21 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, and a linker. [1314] Embodiment 47. The combination of embodiment 18 or 19, wherein the polypeptide comprises one or more HSV RL2.1 polypeptides or antigenic fragments thereof, one or more HSV UL54 polypeptides or antigenic fragments thereof, one or more HSV UL5.2 polypeptides or antigenic fragments thereof, one or more HSV UL40 polypeptides or antigenic fragments thereof, one or more HSV UL47 polypeptides or antigenic fragments thereof, and one or more HSV UL46 polypeptides or antigenic fragments thereof. [1315] Embodiment 48. The combination of any one of embodiments 18, 19, and 47, wherein the polypeptide comprises, in N-terminus to C-terminus order, a RL2.1 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, and a linker. [1316] Embodiment 49. The combination of any one of embodiments 18, 19, and 47, wherein the polypeptide comprises, in N-terminus to C-terminus order, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, a RL2.1 polypeptide or antigenic fragment thereof, and a linker. [1317] Embodiment 50. The combination of embodiment 18 or 19, wherein the polypeptide comprises one or more HSV RL2.1 polypeptides or antigenic fragments thereof, one or more HSV RS1 polypeptides or antigenic fragments thereof, and one or more HSV UL54 polypeptides or antigenic fragments thereof. [1318] Embodiment 51. The combination of any one of embodiments 18, 19, and 50, wherein the polypeptide comprises, in N-terminus to C-terminus order, a RL2.1 polypeptide or antigenic fragment thereof, a linker, a RS1 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, and a linker. [1319] Embodiment 52. The combination of embodiment 18 or 19, wherein the polypeptide comprises one or more HSV UL29 polypeptides or antigenic fragments thereof, one or more HSV UL39 polypeptides or antigenic fragments thereof, and one or more HSV UL9 polypeptides or antigenic fragments thereof. [1320] Embodiment 53. The combination of any one of embodiments 18, 19, and 52, wherein the polypeptide comprises, in N-terminus to C-terminus order, an UL29 polypeptide or antigenic fragment thereof, a linker, an UL39 polypeptide or antigenic fragment thereof, a linker, an UL9 polypeptide or antigenic fragment thereof, and a linker. [1321] Embodiment 54. The combination of embodiment 18 or 19, wherein the polypeptide comprises one or more HSV UL5.1 polypeptides or antigenic fragments thereof, one or more HSV UL5.2 polypeptides or antigenic fragments thereof, one or more HSV UL40 polypeptides or antigenic fragments thereof, and one or more HSV UL30.1 polypeptides or antigenic fragments thereof. [1322] Embodiment 55. The combination of any one of embodiments 18, 19, and 54, wherein the polypeptide comprises, in N-terminus to C-terminus order, an UL5.1 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL30.1 polypeptide or antigenic fragment thereof, and a linker. [1323] Embodiment 56. The combination of embodiment 18 or 19, wherein the polypeptide comprises one or more HSV UL47 polypeptides or antigenic fragments thereof, one or more HSV UL46 polypeptides or antigenic fragments thereof, and one or more HSV UL21 polypeptides or antigenic fragments thereof. [1324] Embodiment 57. The combination of any one of embodiments 18, 19, and 56, wherein the polypeptide comprises, in N-terminus to C-terminus order, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL21 polypeptide or antigenic fragment thereof, and a linker. [1325] Embodiment 58. The combination of embodiment 18 or 19, wherein the polypeptide comprises one or more HSV UL29 polypeptides or antigenic fragments thereof, one or more HSV UL39 polypeptides or antigenic fragments thereof, one or more HSV UL9 polypeptides or antigenic fragments thereof, one or more HSV UL5.1 polypeptides or antigenic fragments thereof, one or more HSV UL5.2 polypeptides or antigenic fragments thereof, one or more HSV UL40 polypeptides or antigenic fragments thereof, and one or more HSV UL30.1 polypeptides or antigenic fragments thereof. [1326] Embodiment 59. The combination of any one of embodiments 18, 19, and 58, wherein the polypeptide comprises, in N-terminus to C-terminus order, an UL29 polypeptide or antigenic fragment thereof, a linker, an UL39 polypeptide or antigenic fragment thereof, a linker, an UL9 polypeptide or antigenic fragment thereof, a linker, an UL5.1 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL30.1 polypeptide or antigenic fragment thereof, and a linker. [1327] Embodiment 60. The combination of any one of embodiments 18, 19, and 58, wherein the polypeptide comprises, in N-terminus to C-terminus order, an UL30.1 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL5.1 polypeptide or antigenic fragment thereof, a linker, an UL9 polypeptide or antigenic fragment thereof, a linker, an UL39 polypeptide or antigenic fragment thereof, a linker, an UL29 polypeptide or antigenic fragment thereof, and a linker. [1328] Embodiment 61. The combination of embodiment 18 or 19, wherein the polypeptide comprises one or more HSV RL2.1 polypeptides or antigenic fragments thereof, one or more HSV RS1 polypeptides or antigenic fragments thereof, one or more HSV UL54 polypeptides or antigenic fragments thereof, one or more HSV UL47 polypeptides or antigenic fragments thereof, one or more HSV UL46 polypeptides or antigenic fragments thereof, and one or more HSV UL21 polypeptides or antigenic fragments thereof. [1329] Embodiment 62. The combination of any one of embodiments 18, 19, and 61, wherein the polypeptide comprises, in N-terminus to C-terminus order, a RL2.1 polypeptide or antigenic fragment thereof, a linker, a RS1 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL21 polypeptide or antigenic fragment thereof, and a linker. [1330] Embodiment 63. The combination of any one of embodiments 18, 19, and 61, wherein the polypeptide comprises, in N-terminus to C-terminus order, an UL21 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, a RS1 polypeptide or antigenic fragment thereof, a linker, a RL2.1 polypeptide or antigenic fragment thereof, and a linker. [1331] Embodiment 64. The combination of embodiment 18 or 19, wherein the polypeptide comprises one or more HSV UL5.1 polypeptides or antigenic fragments thereof, one or more HSV UL5.2 polypeptides or antigenic fragments thereof, one or more HSV UL40 polypeptides or antigenic fragments thereof, one or more HSV UL30.1 polypeptides or antigenic fragments thereof, one or more HSV UL47 polypeptides or antigenic fragments thereof, one or more HSV UL46 polypeptides or antigenic fragments thereof, and one or more HSV UL21 polypeptides or antigenic fragments thereof. [1332] Embodiment 65. The combination of any one of embodiments 18, 19, and 64, wherein the polypeptide comprises, in N-terminus to C-terminus order, an UL5.1 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL30.1 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL21 polypeptide or antigenic fragment thereof, and a linker. [1333] Embodiment 66. The combination of any one of embodiments 18, 19, and 64, wherein the polypeptide comprises, in N-terminus to C-terminus order, an UL21 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL30.1 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL5.1 polypeptide or antigenic fragment thereof, and a linker. [1334] Embodiment 67. The combination of any one of the preceding embodiments, wherein the one or more GP polypeptides comprise a secretory signal. [1335] Embodiment 68. The combination of any one of embodiments 11-67, wherein at least one of the one or more GP polypeptides comprise (i) an HSV gB, variant thereof, or one or more antigenic portions thereof and (ii) a secretory signal. [1336] Embodiment 69. The combination of any one of embodiments 18-67, wherein one or more T-cell string polypeptides comprise a secretory signal. [1337] Embodiment 70. The combination of any of embodiments 67-69, wherein the secretory signal is located at the N-terminus of the polypeptide. [1338] Embodiment 71. The combination of any of embodiments 67-70, wherein the secretory signal is a viral secretory signal. [1339] Embodiment 72. The combination of embodiment 71, wherein the viral secretory signal is an HSV secretory signal. [1340] Embodiment 73. The combination of embodiment 72, wherein the HSV secretory signal is an HSV- 1 secretory signal. [1341] Embodiment 74. The combination of embodiment 73, wherein the HSV-1 secretory signal is an HSV-1 gB secretory signal. [1342] Embodiment 75. The combination of embodiment 74, wherein the HSV-1 gB secretory signal comprises or consists of an amino acid sequence of SEQ ID NO: 214, 359 or 385. [1343] Embodiment 76. The combination of embodiment 73, wherein the HSV-1 secretory signal is an HSV-1 gC secretory signal. [1344] Embodiment 77. The combination of embodiment 76, wherein the HSV-1 gC secretory signal comprises or consists of an amino acid sequence of SEQ ID NO: 357. [1345] Embodiment 78. The combination of embodiment 73, wherein the HSV-1 secretory signal is an HSV-1 gD secretory signal. [1346] Embodiment 79. The combination of embodiment 78, wherein the HSV-1 gD secretory signal comprises or consists of an amino acid sequence of SEQ ID NO: 213, 356, or 407. [1347] Embodiment 80. The combination of embodiment 73, wherein the HSV-1 secretory signal is an HSV-1 gE secretory signal. [1348] Embodiment 81. The combination of embodiment 80, wherein the HSV-1 gE secretory signal comprises or consists of an amino acid sequence of SEQ ID NO: 358. [1349] Embodiment 82. The combination of embodiment 72, wherein the HSV secretory signal is an HSV- 2 secretory signal. [1350] Embodiment 83. The combination of embodiment 82, wherein the HSV-2 secretory signal is an HSV-2 gB secretory signal. [1351] Embodiment 84. The combination of embodiment 83, wherein the HSV-2 gB secretory signal comprises or consists of an amino acid sequence of SEQ ID NO: 360. [1352] Embodiment 85. The combination of embodiment 82, wherein the HSV-2 secretory signal is an HSV-2 gC secretory signal. [1353] Embodiment 86. The combination of embodiment 85, wherein the HSV-2 gC secretory signal comprises or consists of an amino acid sequence of SEQ ID NO: 32. [1354] Embodiment 87. The combination of embodiment 82, wherein the HSV-2 secretory signal is an HSV-2 gD secretory signal. [1355] Embodiment 88. The combination of embodiment 87, wherein the HSV-2 gD secretory signal comprises or consists of an amino acid sequence of SEQ ID NO: 29-31, or 354-355. [1356] Embodiment 89. The combination of embodiment 82, wherein the HSV-2 secretory signal is an HSV-2 gE secretory signal. [1357] Embodiment 90. The combination of embodiment 89, wherein the HSV-2 gE secretory signal comprises or consists of an amino acid sequence of SEQ ID NO: 33, 216, or 292. [1358] Embodiment 91. The combination of embodiment 82, wherein the HSV-2 secretory signal is an HSV-2 gI secretory signal. [1359] Embodiment 92. The combination of embodiment 91, wherein the HSV-2 gI secretory signal comprises or consists of an amino acid sequence of SEQ ID NO: 215, 361, 362, or 409. [1360] Embodiment 93. The combination of any one of embodiments 67-69, wherein the secretory signal is an Ebola spike glycoprotein secretory signal. [1361] Embodiment 94. The combination of embodiment 93, wherein the Ebola spike glycoprotein secretory signal comprises or consists of an amino acid sequence of SEQ ID NO: 217 or 386. [1362] Embodiment 95. The combination of any one of embodiments 67-69, wherein the secretory signal is an IL2 secretory signal. [1363] Embodiment 96. The combination of embodiment 95, wherein the IL2 secretory signal comprises or consists of an amino acid sequence of SEQ ID NO: 28. [1364] Embodiment 97. The combination of any one of embodiments 67-69, wherein the secretory signal is a human secretory signal. [1365] Embodiment 98. The combination of embodiment 97, wherein the secretory signal is a human Ig heavy chain secretory signal. [1366] Embodiment 99. The combination of embodiment 98, wherein the human Ig heavy chain secretory signal comprises or consists of an amino acid sequence of SEQ ID NO: 395-401, or 402. [1367] Embodiment 100. The combination of any one of the preceding embodiments, wherein the one or more GP polypeptides comprise a transmembrane region. [1368] Embodiment 101. The combination of any one of embodiments 11-100, wherein at least one of the one or more GP polypeptides that comprise (i) an HSV gB, variant thereof, or one or more antigenic portions thereof and (ii) a transmembrane region. [1369] Embodiment 102. The combination of any one of embodiments 18-101, wherein the one or more T-cell string polypeptides comprise a transmembrane region. [1370] Embodiment 103. The combination of any one of embodiments 100-102, wherein the transmembrane region comprises or consists of a heterologous transmembrane region. [1371] Embodiment 104. The combination of any one of embodiments 100-103, wherein the transmembrane region comprises or consists of a viral transmembrane region. [1372] Embodiment 105. The polyribonucleotide of embodiment 104, wherein the transmembrane region comprises or consists of an HSV transmembrane region. [1373] Embodiment 106. The combination of embodiment 105, wherein the HSV transmembrane region comprises or consists of an HSV-1 or HSV-2 transmembrane region. [1374] Embodiment 107. The combination of embodiment 105 or 106, wherein the transmembrane region comprises or consists of an HSV gD transmembrane region. [1375] Embodiment 108. The combination of embodiment 107, wherein the HSV gD transmembrane region comprises an amino acid sequence according to SEQ ID NO: 468. [1376] Embodiment 109. The combination of any one of embodiments 100-103, wherein the transmembrane region comprises or consists of an hDAF-GPI anchor region. [1377] Embodiment 110. The combination of embodiment 109, wherein the hDAF-GPI anchor region comprises an amino acid sequence according to SEQ ID NO: 469. [1378] Embodiment 111. The combination of embodiment 105 or 106, wherein the HSV transmembrane region comprises or consists of an HSV gB transmembrane region. [1379] Embodiment 112. The combination of embodiment 111, wherein the HSV gB transmembrane region consists of an amino acid sequence according to SEQ ID NO: 470. [1380] Embodiment 113. The combination of embodiment 111, wherein the HSV gB transmembrane region consists of an amino acid sequence according to SEQ ID NO: 471. [1381] Embodiment 114. The combination of any one of embodiments 100-103, wherein the transmembrane region comprises or consists of a VSV-G transmembrane region. [1382] Embodiment 115. The combination of embodiment 114, wherein the VSV-G transmembrane region comprises an amino acid sequence according to SEQ ID NO: 472. [1383] Embodiment 116. The combination of any one of embodiments 1-99, wherein the one or more GP polypeptides do not comprise a transmembrane region. [1384] Embodiment 117. The combination of any one of embodiments 18-99, wherein the one or more T- cell string polypeptides do not comprise a transmembrane region. [1385] Embodiment 118. The combination of any one of the preceding embodiments, wherein one or more GP polypeptides comprise a MHC Class I Trafficking Signal (MITD). [1386] Embodiment 119. The combination of any one of embodiments 11-118, wherein at least one of the one or more GP polypeptides comprises (i) an HSV gB, variant thereof, or one or more antigenic portions thereof and (ii) a MITD. [1387] Embodiment 120. The combination of any one of embodiments 18-119, wherein one or more T- cell string polypeptides comprise a MITD. [1388] Embodiment 121. The combination of any one of embodiments 118-120, wherein the MITD comprises an amino acid sequence according to SEQ ID NO: 473. [1389] Embodiment 122. The combination of any one of the preceding embodiments, wherein the one or more GP polypeptides comprise a multimerization region. [1390] Embodiment 123. The combination of any one of embodiments 11-122, wherein at least one of the one or more GP polypeptides comprises (i) an HSV gB, variant thereof, or one or more antigenic portions thereof and (ii) a multimerization region. [1391] Embodiment 124. The combination of any one of embodiments 18-123, wherein the one or more T-cell string polypeptides comprise a multimerization region. [1392] Embodiment 125. The combination of any one of embodiments 122-124, wherein the multimerization region comprising or consisting of the amino acid sequence of SEQ ID NO: 474. [1393] Embodiment 126. The combination of any one of the preceding embodiments, wherein the one or more GP polypeptides comprise one or more linkers. [1394] Embodiment 127. The combination of any one of embodiments 11-126, wherein at least one of the one or more GP polypeptides comprises (i) an HSV gB, variant thereof, or one or more antigenic portions thereof and (ii) one or more linkers. [1395] Embodiment 128. The combination of any one of embodiments 18-127, wherein the one or more T-cell string polypeptides comprise one or more linkers. [1396] Embodiment 129. The combination of any one of embodiments 126-128, wherein the one or more linkers comprise one or more glycine-serine linkers. [1397] Embodiment 130. The combination of any one of embodiments 126-129, wherein one or more linkers comprise an amino acid sequence according to SEQ ID NO: 477. [1398] Embodiment 131. The combination of any one of the preceding embodiments, wherein at least one of the plurality of polyribonucleotides is a codon-optimized polyribonucleotides. [1399] Embodiment 132. The combination of any one of embodiments 1-131, wherein the first set of polyribonucleotides comprises at least one polyribonucleotide that is a codon-optimized polyribonucleotide. [1400] Embodiment 133. The combination of any one of embodiments 1-132, wherein the polyribonucleotides in the first set of polyribonucleotides are codon-optimized polyribonucleotides. [1401] Embodiment 134. The combination of any one of embodiments 1-133, wherein the second set of polyribonucleotides comprises at least one polyribonucleotide that is a codon-optimized polyribonucleotide. [1402] Embodiment 135. The combination of any one of embodiments 1-134, wherein at least one of the one or more GP polypeptides comprises an amino acid sequence according to SEQ ID NO: 65-68, 131, or 159-163. [1403] Embodiment 136. The combination of any one of embodiments 2-135, wherein the polyribonucleotide encoding an antigenic portion of HSV gC comprises a ribonucleic acid sequence according to SEQ ID NO: 104-115, 189-208, 318-321, or 348-351. [1404] Embodiment 137. The combination of any one of embodiments 1-136, wherein the at least one of the one or more GP polypeptides comprise an amino acid sequence according to SEQ ID NO: 69-71, or 72. [1405] Embodiment 138. The combination of any one of embodiments 2-137, wherein the polyribonucleotide encoding an antigenic portion of HSV gD comprises a ribonucleic acid sequence according to SEQ ID NO: 116-120, or 352. [1406] Embodiment 139. The combination of any one of embodiments 1-138, wherein at least one of the one or more GP polypeptides comprise an amino acid sequence according to SEQ ID NO: 73-76, 132, 164, or 327- 328. [1407] Embodiment 140. The combination of any one of embodiments 2-139, wherein the polyribonucleotide encoding an antigenic portion of HSV gD comprises a ribonucleic acid sequence according to SEQ ID NO: 121-130, 209-212, 322-326, or 353. [1408] Embodiment 141. The combination of any one of embodiments 18-140, wherein at least one of the one or more T-cell string polypeptides comprises an HSV-1 gD secretory signal, one or more RL2 polypeptides or antigenic fragments thereof, one or more RS1 polypeptides or antigenic fragments thereof, one or more UL54 polypeptides or antigenic fragments thereof, and a MITD. [1409] Embodiment 142. The combination of any one of embodiments 18-141, wherein at least one of the one or more T-cell string polypeptides comprises, in N-terminus to C-terminus order, an HSV-1 gD secretory signal, an RL2 polypeptide or antigenic fragment thereof, a linker, an RL2 polypeptide or antigenic fragment thereof, a linker, an RS1 polypeptide or antigenic fragment thereof, a linker, a UL54 polypeptide or antigenic fragment thereof, a linker, and a MITD. [1410] Embodiment 143. The combination of embodiment 142, wherein at least one of the one or more T-cell string polypeptides comprises or consists of an amino acid sequence according to SEQ ID NO: 595. [1411] Embodiment 144. The combination of embodiment 143, wherein the second set of polyribonucleotides comprises a polyribonucleotide that comprises or consists of an amino acid sequence according to SEQ ID NO: 619. [1412] Embodiment 145. The combination of any one of embodiments 18-140, wherein at least one of the one or more T-cell string polypeptides comprise, in N-terminus to C-terminus order, an HSV-1 gD secretory signal, an UL54 polypeptide or antigenic fragment thereof, a linker, an RS1 polypeptide or antigenic fragment thereof, a linker, an RL2 polypeptide or antigenic fragment thereof, a linker, a RL2 polypeptide or antigenic fragment thereof, a linker, and a MITD. [1413] Embodiment 146. The combination of embodiment 145, wherein at least one of the one or more T-cell string polypeptides comprises or consists of an amino acid sequence according to SEQ ID NO: 599. [1414] Embodiment 147. The combination of embodiment 146, wherein the second set of polyribonucleotides comprises a polyribonucleotide that comprises or consists of an amino acid sequence according to SEQ ID NO: 620. [1415] Embodiment 148. The combination of any one of embodiments 18-140, wherein at least one of the one or more T-cell string polypeptides comprises an HSV-1 gD secretory signal, an UL29 polypeptide or antigenic fragment thereof, a linker, an UL39 polypeptide or antigenic fragment thereof, a linker, an UL49 polypeptide or antigenic fragment thereof, a linker, a UL9 polypeptide or antigenic fragment thereof, and a linker. [1416] Embodiment 149. The combination of any one of embodiments 18-140, and 148 wherein at least one of the one or more T-cell string polypeptides comprises, in N-terminus to C-terminus order, an HSV-1 gD secretory signal, an UL29 polypeptide or antigenic fragment thereof, a linker, an UL39 polypeptide or antigenic fragment thereof, a linker, an UL49 polypeptide or antigenic fragment thereof, a linker, a UL9 polypeptide or antigenic fragment thereof, a linker, and a MITD. [1417] Embodiment 150. The combination of embodiment 149, wherein at least one of the one or more T-cell string polypeptides comprises or consists of an amino acid sequence according to SEQ ID NO: 596. [1418] Embodiment 151. The combination of embodiment 150, wherein the second set of polyribonucleotides comprises a polyribonucleotide that comprises or consists of an amino acid sequence according to SEQ ID NO: 621. [1419] Embodiment 152. The combination of any one of embodiments 18-140, wherein at least one of the one or more T-cell string polypeptides comprises, in N-terminus to C-terminus order, an HSV-1 gD secretory signal, an UL9 polypeptide or antigenic fragment thereof, a linker, an UL49 polypeptide or antigenic fragment thereof, a linker, an UL39 polypeptide or antigenic fragment thereof, a linker, a UL29 polypeptide or antigenic fragment thereof, a linker, and a MITD. [1420] Embodiment 153. The combination of embodiment 152, wherein at least one of the one or more T-cell string polypeptides comprises or consists of an amino acid sequence according to SEQ ID NO: 600. [1421] Embodiment 154. The combination of embodiment 153, wherein the second set of polyribonucleotides comprises a polyribonucleotide that comprises or consists of an amino acid sequence according to SEQ ID NO: 622. [1422] Embodiment 155. The combination of any one of embodiments 18-140, wherein at least one of the one or more T-cell string polypeptides comprises an HSV-1 gD secretory signal, one or more HSV UL30 polypeptides or antigenic fragments thereof, one or more HSV UL40 polypeptides or antigenic fragments thereof, one or more HSV UL5 polypeptides or antigenic fragments thereof, one or more HSV UL52 polypeptides or antigenic fragments thereof, and a MITD. [1423] Embodiment 156. The combination of any one of embodiments 18-140, and 155, wherein at least one of the one or more T-cell string polypeptides comprises, in N-terminus to C-terminus order, an HSV-1 gD secretory signal, an UL30 polypeptide or antigenic fragment thereof, a linker, an UL30 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, a UL5 polypeptide or antigenic fragment thereof, a linker, a UL5 polypeptide or antigenic fragment thereof, a linker, a UL52 polypeptide or antigenic fragment thereof, a linker, and a MITD. [1424] Embodiment 157. The combination of embodiment 156, wherein at least one of the one or more T-cell string polypeptides comprises or consists of an amino acid sequence according to SEQ ID NO: 597. [1425] Embodiment 158. The combination of embodiment 157, wherein the second set of polyribonucleotides comprises a polyribonucleotide that comprises or consists of an amino acid sequence according to SEQ ID NO: 623. [1426] Embodiment 159. The combination of any one of embodiments 18-140, wherein at least one of the one or more T-cell string polypeptides comprises, in N-terminus to C-terminus order, an HSV-1 gD secretory signal, an UL52 polypeptide or antigenic fragment thereof, a linker, an UL5 polypeptide or antigenic fragment thereof, a linker, an UL5 polypeptide or antigenic fragment thereof, a linker, a UL40 polypeptide or antigenic fragment thereof, a linker, a UL30 polypeptide or antigenic fragment thereof, a linker, a UL30 polypeptide or antigenic fragment thereof, a linker, and a MITD. [1427] Embodiment 160. The combination of embodiment 159, wherein at least one of the one or more T-cell string polypeptides comprises or consists of an amino acid sequence according to SEQ ID NO: 601. [1428] Embodiment 161. The combination of embodiment 160, wherein the second set of polyribonucleotides comprises a polyribonucleotide that comprises or consists of an amino acid sequence according to SEQ ID NO: 624. [1429] Embodiment 162. The combination of any one of embodiments 18-140, wherein at least one of the one or more T-cell string polypeptides comprises an HSV-1 gD secretory signal, one or more HSV UL1 polypeptides or antigenic fragments thereof, one or more HSV UL19 polypeptides or antigenic fragments thereof, one or more HSV UL21 polypeptides or antigenic fragments thereof, one or more HSV UL27 polypeptides or antigenic fragments thereof, one or more HSV UL46 polypeptides or antigenic fragments thereof, one or more HSV UL47 polypeptides or antigenic fragments thereof, one or more UL48 polypeptides or antigenic fragments thereof, one or more HSV UL25 polypeptides or antigenic fragments thereof, and a MITD. [1430] Embodiment 163. The combination of any one of embodiments 18-140, and 162, wherein at least one of the one or more T-cell string polypeptides comprises, in N-terminus to C-terminus order, an HSV-1 gD secretory signal, an HSV UL1 polypeptide or antigenic fragment thereof, a linker, an HSV UL19 polypeptide or antigenic fragment thereof, a linker, an HSV UL21 polypeptide or antigenic fragment thereof, a linker, an HSV UL27 polypeptide or antigenic fragment thereof, a linker, an HSV UL27 polypeptide or antigenic fragment thereof, a linker, an HSV UL46 polypeptide or antigenic fragment thereof, a linker, an HSV UL47 polypeptide or antigenic fragment thereof, a linker, an HSV UL25 polypeptide or antigenic fragment thereof, a linker, an HSV UL48 polypeptide or antigenic fragment thereof, a linker, and a MITD. [1431] Embodiment 164. The combination of embodiment 163, wherein at least one of the one or more T-cell string polypeptides comprises or consists of an amino acid sequence according to SEQ ID NO: 598. [1432] Embodiment 165. The combination of embodiment 164, wherein the second set of polyribonucleotides comprises a polyribonucleotide that comprises or consists of an amino acid sequence according to SEQ ID NO: 625. [1433] Embodiment 166. The combination of any one of embodiments 18-140, wherein at least one of the one or more T-cell string polypeptides comprises, in N-terminus to C-terminus order, an HSV-1 gD secretory signal, an HSV UL48 polypeptide or antigenic fragment thereof, a linker, an HSV UL25 polypeptide or antigenic fragment thereof, a linker, an HSV UL47 polypeptide or antigenic fragment thereof, a linker, an HSV UL46 polypeptide or antigenic fragment thereof, a linker, an HSV UL27 polypeptide or antigenic fragment thereof, a linker, an HSV UL27 polypeptide or antigenic fragment thereof, a linker, an HSV UL21 polypeptide or antigenic fragment thereof, a linker, an HSV UL19 polypeptide or antigenic fragment thereof, a linker, an HSV UL1 polypeptide or antigenic fragment thereof, a linker, and a MITD. [1434] Embodiment 167. The combination of embodiment 166, wherein at least one of the one or more T-cell string polypeptides comprises or consists of an amino acid sequence according to SEQ ID NO: 602. [1435] Embodiment 168. The combination of embodiment 167, wherein the second set of polyribonucleotides comprises a polyribonucleotide that comprises or consists of an amino acid sequence according to SEQ ID NO: 626. [1436] Embodiment 169. The combination of any one of embodiments 18-140, wherein at least one of the one or more T-cell string polypeptides comprises an HSV-2 gD secretory signal, one or more RL2 polypeptides or antigenic fragments thereof, one or more RS1 polypeptides or antigenic fragments thereof, one or more UL54 polypeptides or antigenic fragments thereof, and a MITD. [1437] Embodiment 170. The combination of any one of embodiments 18-140, and 169, wherein at least one of the one or more T-cell string polypeptides comprises, in N-terminus to C-terminus order, an HSV-2 gD secretory signal, an RL2 polypeptide or antigenic fragment thereof, a linker, an RL2 polypeptide or antigenic fragment thereof, a linker, an RS1 polypeptide or antigenic fragment thereof, a linker, a UL54 polypeptide or antigenic fragment thereof, a linker, and a MITD. [1438] Embodiment 171. The combination of embodiment 170, wherein at least one of the one or more T-cell string polypeptides comprises or consists of an amino acid sequence according to SEQ ID NO: 603. [1439] Embodiment 172. The combination of embodiment 171, wherein the second set of polyribonucleotides comprises a polyribonucleotide that comprises or consists of an amino acid sequence according to SEQ ID NO: 627. [1440] Embodiment 173. The combination of any one of embodiments 18-140, wherein at least one of the one or more T-cell string polypeptides comprises, in N-terminus to C-terminus order, HSV-2 gD secretory signal, an UL54 polypeptide or antigenic fragment thereof, a linker, an RS1 polypeptide or antigenic fragment thereof, a linker, an RL2 polypeptide or antigenic fragment thereof, a linker, a RL2 polypeptide or fragment thereof, a linker, and a MITD. [1441] Embodiment 174. The combination of embodiment 173, wherein at least one of the one or more T-cell string polypeptides comprises or consists of an amino acid sequence according to SEQ ID NO: 604. [1442] Embodiment 175. The combination of embodiment 174, wherein the second set of polyribonucleotides comprises a polyribonucleotide that comprises or consists of an amino acid sequence according to SEQ ID NO: 628. [1443] Embodiment 176. The combination of any one of embodiments 18-140, wherein at least one of the one or more T-cell string polypeptides comprises an HSV-1 gD secretory signal, one or more HSV US1 polypeptides or antigenic fragments thereof, one or more HSV US8 polypeptides or antigenic fragments thereof, one or more HSV US12 polypeptides or antigenic fragments thereof, one or more HSV UL50 polypeptides or antigenic fragments thereof, one or more HSV UL26 polypeptides or antigenic fragments thereof, and one or more HSV US10 polypeptides or antigenic fragments thereof, and a MITD. [1444] Embodiment 177. The combination of any one of embodiments 18-140, and 176, wherein at least one of the one or more T-cell string polypeptides comprises, in N-terminus to C-terminus order, an HSV-1 gD secretory signal, an US1 polypeptide or antigenic fragment thereof, a linker, an US1 polypeptide or antigenic fragment thereof, a linker, an US8 polypeptide or antigenic fragment thereof, a linker, a US8 polypeptide or antigenic fragment thereof, a linker, a US8 polypeptide or antigenic fragment thereof, a linker, a US12 polypeptide or antigenic fragment thereof, a linker, a UL50 polypeptide or antigenic fragment thereof, a linker, a UL26 polypeptide or antigenic fragment thereof, a linker, a UL26 polypeptide or antigenic fragment thereof, a linker, a US10 polypeptide or antigenic fragment thereof, a linker, and a MITD. [1445] Embodiment 178. The combination of embodiment 177, wherein at least one of the one or more T-cell string polypeptides comprises or consists of an amino acid sequence according to SEQ ID NO: 605. [1446] Embodiment 179. The combination of embodiment 178, wherein the second set of polyribonucleotides comprises a polyribonucleotide that comprises or consists of an amino acid sequence according to SEQ ID NO: 629. [1447] Embodiment 180. The combination of any one of embodiments 18-140, wherein at least one of the one or more T-cell string polypeptides comprises, in N-terminus to C-terminus order, an HSV-1 gD secretory signal, an UL26 polypeptide or antigenic fragment thereof, a linker, an UL26 polypeptide or antigenic fragment thereof, a linker, an US10 polypeptide or antigenic fragment thereof, a linker, a UL50 polypeptide or antigenic fragment thereof, a linker, a US12 polypeptide or antigenic fragment thereof, a linker, a US8 polypeptide or antigenic fragment thereof, a linker, a US8 polypeptide or antigenic fragment thereof, a linker, a US8 polypeptide or antigenic fragment thereof, a linker, a US1 polypeptide or antigenic fragment thereof, a linker, a US1 polypeptide or antigenic fragment thereof, a linker, and a MITD. [1448] Embodiment 181. The combination of embodiment 180, wherein at least one of the one or more T-cell string polypeptides comprises or consists of an amino acid sequence according to SEQ ID NO: 606. [1449] Embodiment 182. The combination of embodiment 181, wherein the second set of polyribonucleotides comprises a polyribonucleotide that comprises or consists of an amino acid sequence according to SEQ ID NO: 630. [1450] Embodiment 183. The combination of any one of embodiments 18-140, wherein at least one of the one or more T-cell string polypeptides comprises an HSV-1 gD secretory signal, one or more RL2 polypeptides or antigenic fragments thereof, one or more RS1 polypeptides or antigenic fragments thereof, one or more UL54 polypeptides or antigenic fragments thereof, and an HSV-1 gD transmembrane region. [1451] Embodiment 184. The combination of any one of embodiments 18-140, and 183, wherein at least one of the one or more T-cell string polypeptides comprises, in N-terminus to C-terminus order, an HSV-1 gD secretory signal, an RL2 polypeptide or antigenic fragment thereof, a linker, an RL2 polypeptide or antigenic fragment thereof, a linker, an RS1 polypeptide or antigenic fragment thereof, a linker, a UL54 polypeptide or antigenic fragment thereof, a linker, and an HSV-1 gD transmembrane region. [1452] Embodiment 185. The combination of embodiment 184, wherein at least one of the one or more T-cell string polypeptides comprises or consists of an amino acid sequence according to SEQ ID NO: 607. [1453] Embodiment 186. The combination of embodiment 185, wherein the second set of polyribonucleotides comprises a polyribonucleotide that comprises or consists of an amino acid sequence according to SEQ ID NO: 631. [1454] Embodiment 187. The combination of any one of embodiments 18-140, wherein at least one of the one or more T-cell string polypeptides comprises an HSV-1 gD secretory signal, one or more RL2 polypeptides or antigenic fragments thereof, one or more RS1 polypeptides or antigenic fragments thereof, one or more UL54 polypeptides or antigenic fragments thereof, and a VSV-G transmembrane region. [1455] Embodiment 188. The combination of any one of embodiments 18-140, and 187, wherein at least one of the one or more T-cell string polypeptides comprises, in N-terminus to C-terminus order, an HSV-1 gD secretory signal, an RL2 polypeptide or antigenic fragment thereof, a linker, an RL2 polypeptide or antigenic fragment thereof, a linker, an RS1 polypeptide or antigenic fragment thereof, a linker, a UL54 polypeptide or antigenic fragment thereof, a linker, and a VSV-G transmembrane region. [1456] Embodiment 189. The combination of embodiment 188, wherein at least one of the one or more T-cell string polypeptides comprises or consists of an amino acid sequence according to SEQ ID NO: 608. [1457] Embodiment 190. The combination of embodiment 189, wherein the second set of polyribonucleotides comprises a polyribonucleotide that comprises or consists of an amino acid sequence according to SEQ ID NO: 632. [1458] Embodiment 191. The combination of any one of embodiments 1-2 and 18-19, wherein the polypeptide comprises an HSV-1 gD secretory signal, one or more HSV RL2 polypeptides or antigenic fragments thereof, one or more HSV UL54 polypeptides or antigenic fragments thereof, one or more HSV UL47 polypeptides or antigenic fragments thereof, one or more HSV UL46 polypeptides or antigenic fragments thereof, one or more HSV UL21 polypeptides or antigenic fragments thereof, and a MITD. [1459] Embodiment 192. The combination of any one of embodiments 191, wherein the polypeptide comprises, in N-terminus to C-terminus order, an HSV-1 gD secretory signal, an RL2 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, a UL46 polypeptide or antigenic fragment thereof, a linker, a UL21 polypeptide or antigenic fragment thereof, a linker, and a MITD. [1460] Embodiment 193. The combination of embodiment 192, wherein the polypeptide comprises or consists of an amino acid sequence according to SEQ ID NO: 633. [1461] Embodiment 194. The combination of embodiment 193, wherein the polyribonucleotide comprises or consists of an amino acid sequence according to SEQ ID NO: 653. [1462] Embodiment 195. The combination of any one of embodiments 1-2, wherein the polypeptide comprises, in N-terminus to C-terminus order, an HSV-1 gD secretory signal, an UL21 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, an RL2 polypeptide or antigenic fragment thereof, a linker, and a MITD. [1463] Embodiment 196. The combination of embodiment 195, wherein the polypeptide comprises or consists of an amino acid sequence according to SEQ ID NO: 634. [1464] Embodiment 197. The combination of embodiment 196, wherein the polyribonucleotide comprises or consists of an amino acid sequence according to SEQ ID NO: 655. [1465] Embodiment 198. The combination of any one of embodiments 1-2, wherein the polypeptide comprises an HSV-1 gD secretory signal, one or more HSV UL29 polypeptides or antigenic fragments thereof, one or more HSV UL39 polypeptides or antigenic fragments thereof, one or more HSV UL9 polypeptides or antigenic fragments thereof, one or more HSV UL5 polypeptides or antigenic fragments thereof, one or more HSV UL40 polypeptides or antigenic fragments thereof, one or more HSV UL30 polypeptides or antigenic fragments thereof, and a MITD. [1466] Embodiment 199. The combination of any one of embodiments 198, wherein the polypeptide comprises, in N-terminus to C-terminus order, an HSV-1 gD secretory signal, an UL29 polypeptide or antigenic fragment thereof, a linker, an UL39 polypeptide or antigenic fragment thereof, a linker, an UL9 polypeptide or antigenic fragment thereof, a linker, an UL5.1 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL30.1 polypeptide or antigenic fragment thereof, a linker, and a MITD. [1467] Embodiment 200. The combination of embodiment 199, wherein the polypeptide comprises or consists of an amino acid sequence according to SEQ ID NO: 636. [1468] Embodiment 201. The combination of embodiment 200, wherein the polyribonucleotide comprises or consists of an amino acid sequence according to SEQ ID NO: 657. [1469] Embodiment 202. The combination of any one of embodiments 198, wherein the polypeptide comprises, in N-terminus to C-terminus order, an HSV-1 gD secretory signal, an UL30.1 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL5.1 polypeptide or antigenic fragment thereof, a linker, an UL9 polypeptide or antigenic fragment thereof, a linker, an UL39 polypeptide or antigenic fragment thereof, a linker, an UL29 polypeptide or antigenic fragment thereof, a linker, and a MITD. [1470] Embodiment 203. The combination of embodiment 202, wherein the polypeptide comprises or consists of an amino acid sequence according to SEQ ID NO: 275. [1471] Embodiment 204. The combination of embodiment 203, wherein the polyribonucleotide comprises or consists of an amino acid sequence according to SEQ ID NO: 659. [1472] Embodiment 205. The combination of any one of embodiments 1-2, wherein the polypeptide comprises an HSV-1 gD secretory signal, one or more HSV RL2 polypeptides or antigenic fragments thereof, one or more HSV UL54 polypeptides or antigenic fragments thereof, one or more HSV UL9 polypeptides or antigenic fragments thereof, one or more HSV UL39 polypeptides or antigenic fragments thereof, one or more HSV UL5 polypeptides or antigenic fragments thereof, and a MITD. [1473] Embodiment 206. The combination of any one of embodiments 205, wherein the polypeptide comprises, in N-terminus to C-terminus order, an HSV-1 gD secretory signal, an RL2.1 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, an UL9 polypeptide or antigenic fragment thereof, a linker, a UL39 polypeptide or antigenic fragment thereof, a linker, a UL5.1 polypeptide or antigenic fragment thereof, a linker, and a MITD. [1474] Embodiment 207. The combination of embodiment 206, wherein the polypeptide comprises or consists of an amino acid sequence according to SEQ ID NO: 637. [1475] Embodiment 208. The combination of embodiment 207, wherein the polyribonucleotide comprises or consists of an amino acid sequence according to SEQ ID NO: 661. [1476] Embodiment 209. The combination of any one of embodiments 205, wherein the polypeptide comprises, in N-terminus to C-terminus order, an HSV-1 gD secretory signal, an UL5.1 polypeptide or antigenic fragment thereof, a linker, an UL39 polypeptide or antigenic fragment thereof, a linker, an UL9 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, an RL2.1 polypeptide or antigenic fragment thereof, a linker, an UL29 polypeptide or antigenic fragment thereof, a linker, and a MITD. [1477] Embodiment 210. The combination of embodiment 209, wherein the polypeptide comprises or consists of an amino acid sequence according to SEQ ID NO: 638. [1478] Embodiment 211. The combination of embodiment 210, wherein the polyribonucleotide comprises or consists of an amino acid sequence according to SEQ ID NO: 663. [1479] Embodiment 212. The combination of any one of embodiments 1-2, wherein the polypeptide comprises an HSV-1 gD secretory signal, one or more HSV UL47 polypeptides or antigenic fragments thereof, one or more HSV UL46 polypeptides or antigenic fragments thereof, one or more HSV UL21 polypeptides or antigenic fragments thereof, one or more HSV UL5.2 polypeptides or antigenic fragments thereof, one or more HSV UL40 polypeptides or antigenic fragments thereof, one or more HSV UL30.1 polypeptides or antigenic fragments thereof, one or more HSV UL29 polypeptides or antigenic fragments thereof, and a MITD. [1480] Embodiment 213. The combination of any one of embodiments 212, wherein the polypeptide comprises, in N-terminus to C-terminus order, an HSV-1 gD secretory signal, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL21 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL30.1 polypeptide or antigenic fragment thereof, a linker, an UL29 polypeptide or antigenic fragment thereof, a linker, and a MITD. [1481] Embodiment 214. The combination of embodiment 213, wherein the polypeptide comprises or consists of an amino acid sequence according to SEQ ID NO: 639. [1482] Embodiment 215. The combination of embodiment 214, wherein the polyribonucleotide comprises or consists of an amino acid sequence according to SEQ ID NO: 665. [1483] Embodiment 216. The combination of any one of embodiments 212, wherein the polypeptide comprises, in N-terminus to C-terminus order, an HSV-1 gD secretory signal, an UL29 polypeptide or antigenic fragment thereof, a linker, an UL30.1 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL21 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, and a MITD. [1484] Embodiment 217. The combination of embodiment 216, wherein the polypeptide comprises or consists of an amino acid sequence according to SEQ ID NO: 640. [1485] Embodiment 218. The combination of embodiment 217, wherein the polyribonucleotide comprises or consists of an amino acid sequence according to SEQ ID NO: 667. [1486] Embodiment 219. The combination of any one of embodiments 1-2, wherein the polypeptide comprises an HSV-1 gD secretory signal, one or more HSV RL2.1 polypeptides or antigenic fragments thereof, one or more HSV UL54 polypeptides or antigenic fragments thereof, one or more HSV UL5.2 polypeptides or antigenic fragments thereof, one or more HSV UL40 polypeptides or antigenic fragments thereof, one or more HSV UL47 polypeptides or antigenic fragments thereof, one or more HSV UL46 polypeptides or antigenic fragments thereof, and a MITD. [1487] Embodiment 220. The combination of any one of embodiments 219, wherein the polypeptide comprises, in N-terminus to C-terminus order, an HSV-1 gD secretory signal, a RL2.1 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, and a MITD. [1488] Embodiment 221. The combination of embodiment 220, wherein the polypeptide comprises or consists of an amino acid sequence according to SEQ ID NO: 641. [1489] Embodiment 222. The combination of embodiment 221, wherein the polyribonucleotide comprises or consists of an amino acid sequence according to SEQ ID NO: 669. [1490] Embodiment 223. The combination of any one of embodiments 219, wherein the polypeptide comprises, in N-terminus to C-terminus order, an HSV-1 gD secretory signal, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, a RL2.1 polypeptide or antigenic fragment thereof, a linker, and a MITD. [1491] Embodiment 224. The combination of embodiment 223, wherein the polypeptide comprises or consists of an amino acid sequence according to SEQ ID NO: 642. [1492] Embodiment 225. The combination of embodiment 224, wherein the polyribonucleotide comprises or consists of an amino acid sequence according to SEQ ID NO: 671. [1493] Embodiment 226. The combination of any one of embodiments 1-2, wherein the polypeptide comprises an HSV-1 gD secretory signal, one or more HSV RL2.1 polypeptides or antigenic fragments thereof, one or more HSV RS1 polypeptides or antigenic fragments thereof, one or more HSV UL54 polypeptides or antigenic fragments thereof, and a MITD. [1494] Embodiment 227. The combination of any one of embodiments 226, wherein the polypeptide comprises, in N-terminus to C-terminus order, an HSV-1 gD secretory signal, a RL2.1 polypeptide or antigenic fragment thereof, a linker, a RS1 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, and a MITD. [1495] Embodiment 228. The combination of embodiment 227, wherein the polypeptide comprises or consists of an amino acid sequence according to SEQ ID NO: 643. [1496] Embodiment 229. The combination of embodiment 228, wherein the polyribonucleotide comprises or consists of an amino acid sequence according to SEQ ID NO: 673. [1497] Embodiment 230. The combination of any one of embodiments 1-2, wherein the polypeptide comprises an HSV-1 gD secretory signal, one or more HSV UL29 polypeptides or antigenic fragments thereof, one or more HSV UL39 polypeptides or antigenic fragments thereof, and one or more HSV UL9 polypeptides or antigenic fragments thereof, and a MITD. [1498] Embodiment 231. The combination of any one of embodiments 230, wherein the polypeptide comprises, in N-terminus to C-terminus order, an HSV-1 gD secretory signal, an UL29 polypeptide or antigenic fragment thereof, a linker, an UL39 polypeptide or antigenic fragment thereof, a linker, an UL9 polypeptide or antigenic fragment thereof, a linker, and a MITD. [1499] Embodiment 232. The combination of embodiment 231, wherein the polypeptide comprises or consists of an amino acid sequence according to SEQ ID NO: 644. [1500] Embodiment 233. The combination of embodiment 232, wherein the polyribonucleotide comprises or consists of an amino acid sequence according to SEQ ID NO: 675. [1501] Embodiment 234. The combination of any one any one of embodiments 1-2, wherein the polypeptide comprises an HSV-1 gD secretory signal, one or more HSV UL5.1 polypeptides or antigenic fragments thereof, one or more HSV UL5.2 polypeptides or antigenic fragments thereof, one or more HSV UL40 polypeptides or antigenic fragments thereof, one or more HSV UL30.1 polypeptides or antigenic fragments thereof, and a MITD. [1502] Embodiment 235. The combination of any one of embodiments 234, wherein the polypeptide comprises, in N-terminus to C-terminus order, an HSV-1 gD secretory signal, an UL5.1 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL30.1 polypeptide or antigenic fragment thereof, a linker, and a MITD. [1503] Embodiment 236. The combination of embodiment 235, wherein the polypeptide comprises or consists of an amino acid sequence according to SEQ ID NO: 645. [1504] Embodiment 237. The combination of embodiment 236, wherein the polyribonucleotide comprises or consists of an amino acid sequence according to SEQ ID NO: 677. [1505] Embodiment 238. The polyribonucleotide of any one of embodiments 234, wherein the polypeptide comprises an HSV-1 gD secretory signal, one or more HSV UL47 polypeptides or antigenic fragments thereof, one or more HSV UL46 polypeptides or antigenic fragments thereof, one or more HSV UL21 polypeptides or antigenic fragments thereof, and a MITD. [1506] Embodiment 239. The combination of any one of embodiments 238, wherein the polypeptide comprises, in N-terminus to C-terminus order, an HSV-1 gD secretory signal, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL21 polypeptide or antigenic fragment thereof, a linker, and a MITD. [1507] Embodiment 240. The combination of embodiment 239, wherein the polypeptide comprises or consists of an amino acid sequence according to SEQ ID NO: 646. [1508] Embodiment 241. The combination of embodiment 240, wherein the polyribonucleotide comprises or consists of an amino acid sequence according to SEQ ID NO: 679. [1509] Embodiment 242. The combination of any one of embodiments 1-2, wherein the polypeptide comprises an HSV-1 gD secretory signal, one or more HSV UL29 polypeptides or antigenic fragments thereof, one or more HSV UL39 polypeptides or antigenic fragments thereof, one or more HSV UL9 polypeptides or antigenic fragments thereof, one or more HSV UL5.1 polypeptides or antigenic fragments thereof, one or more HSV UL5.2 polypeptides or antigenic fragments thereof, one or more HSV UL40 polypeptides or antigenic fragments thereof, one or more HSV UL30.1 polypeptides or antigenic fragments thereof, and a MITD. [1510] Embodiment 243. The combination of any one of embodiments 242, wherein the polypeptide comprises, in N-terminus to C-terminus order, an HSV-1 gD secretory signal, an UL29 polypeptide or antigenic fragment thereof, a linker, an UL39 polypeptide or antigenic fragment thereof, a linker, an UL9 polypeptide or antigenic fragment thereof, a linker, an UL5.1 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL30.1 polypeptide or antigenic fragment thereof, a linker, and a MITD. [1511] Embodiment 244. The combination of embodiment 243, wherein the polypeptide comprises or consists of an amino acid sequence according to SEQ ID NO: 647. [1512] Embodiment 245. The combination of embodiment 244, wherein the polyribonucleotide comprises or consists of an amino acid sequence according to SEQ ID NO: 681. [1513] Embodiment 246. The combination of any one of embodiments 242, wherein the polypeptide comprises, in N-terminus to C-terminus order, an HSV-1 gD secretory signal, an UL30.1 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL5.1 polypeptide or antigenic fragment thereof, a linker, an UL9 polypeptide or antigenic fragment thereof, a linker, an UL39 polypeptide or antigenic fragment thereof, a linker, an UL29 polypeptide or antigenic fragment thereof, a linker, and a MITD. [1514] Embodiment 247. The combination of embodiment 246, wherein the polypeptide comprises or consists of an amino acid sequence according to SEQ ID NO: 648. [1515] Embodiment 248. The combination of embodiment 247, wherein the polyribonucleotide comprises or consists of an amino acid sequence according to SEQ ID NO: 683. [1516] Embodiment 249. The combination of any one of embodiments 1-2, wherein the polypeptide comprises an HSV-1 gD secretory signal, comprises one or more HSV RL2.1 polypeptides or antigenic fragments thereof, one or more HSV RS1 polypeptides or antigenic fragments thereof, one or more HSV UL54 polypeptides or antigenic fragments thereof, one or more HSV UL47 polypeptides or antigenic fragments thereof, one or more HSV UL46 polypeptides or antigenic fragments thereof, one or more HSV UL21 polypeptides or antigenic fragments thereof, and a MITD. [1517] Embodiment 250. The combination of any one of embodiments 249, wherein the polypeptide comprises, in N-terminus to C-terminus order, an HSV-1 gD secretory signal, a RL2.1 polypeptide or antigenic fragment thereof, a linker, a RS1 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL21 polypeptide or antigenic fragment thereof, a linker, and a MITD. [1518] Embodiment 251. The combination of embodiment 250, wherein the polypeptide comprises or consists of an amino acid sequence according to SEQ ID NO: 649. [1519] Embodiment 252. The combination of embodiment 251, wherein the polyribonucleotide comprises or consists of an amino acid sequence according to SEQ ID NO: 685. [1520] Embodiment 253. The combination of any one of embodiments 249, wherein the polypeptide comprises, in N-terminus to C-terminus order, an HSV-1 gD secretory signal, an UL21 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL54 polypeptide or antigenic fragment thereof, a linker, a RS1 polypeptide or antigenic fragment thereof, a linker, a RL2.1 polypeptide or antigenic fragment thereof, a linker, and a MITD. [1521] Embodiment 254. The combination of embodiment 253, wherein the polypeptide comprises or consists of an amino acid sequence according to SEQ ID NO: 650. [1522] Embodiment 255. The combination of embodiment 254, wherein the polyribonucleotide comprises or consists of an amino acid sequence according to SEQ ID NO: 687. [1523] Embodiment 256. The combination of any one of embodiments 1-2, wherein the polypeptide comprises an HSV-1 gD secretory signal, comprises one or more HSV UL5.1 polypeptides or antigenic fragments thereof, one or more HSV UL5.2 polypeptides or antigenic fragments thereof, one or more HSV UL40 polypeptides or antigenic fragments thereof, one or more HSV UL30.1 polypeptides or antigenic fragments thereof, one or more HSV UL47 polypeptides or antigenic fragments thereof, one or more HSV UL46 polypeptides or antigenic fragments thereof, one or more HSV UL21 polypeptides or antigenic fragments thereof, and a MITD. [1524] Embodiment 257. The combination of any one of embodiments 256, wherein the polypeptide comprises, in N-terminus to C-terminus order, an UL5.1 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL30.1 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL21 polypeptide or antigenic fragment thereof, a linker, and a MITD. [1525] Embodiment 258. The combination of embodiment 257, wherein the polypeptide comprises or consists of an amino acid sequence according to SEQ ID NO: 651. [1526] Embodiment 259. The combination of embodiment 258, wherein the polyribonucleotide comprises or consists of an amino acid sequence according to SEQ ID NO: 689. [1527] Embodiment 260. The combination of any one of embodiments 256, wherein the polypeptide comprises, in N-terminus to C-terminus order, an HSV-1 gD secretory signal, an UL21 polypeptide or antigenic fragment thereof, a linker, an UL46 polypeptide or antigenic fragment thereof, a linker, an UL47 polypeptide or antigenic fragment thereof, a linker, an UL30.1 polypeptide or antigenic fragment thereof, a linker, an UL40 polypeptide or antigenic fragment thereof, a linker, an UL5.2 polypeptide or antigenic fragment thereof, a linker, an UL5.1 polypeptide or antigenic fragment thereof, a linker, and a MITD. [1528] Embodiment 261. The combination of embodiment 260, wherein the polypeptide comprises or consists of an amino acid sequence according to SEQ ID NO: 692. [1529] Embodiment 262. The combination of embodiment 261, wherein the polyribonucleotide comprises or consists of an amino acid sequence according to SEQ ID NO: 691. [1530] Embodiment 263. The combination of any one of embodiments 1-262, wherein at least one of the plurality of polyribonucleotides is an isolated polyribonucleotide. [1531] Embodiment 264. The combination of any one of embodiments 1-263, wherein the first set of polyribonucleotides comprises at least one polyribonucleotide that is an isolated polyribonucleotide. [1532] Embodiment 265. The combination of any one of embodiments 18-264, wherein the second set of polyribonucleotides comprises at least one polyribonucleotide that is an isolated polyribonucleotide. [1533] Embodiment 266. The combination of any one of embodiments 1-265, wherein at least one of the plurality of polyribonucleotides is an engineered polyribonucleotides. [1534] Embodiment 267. The combination of any one of embodiments 1-266, wherein the first set of polyribonucleotides comprises at least one polyribonucleotide that is an engineered polyribonucleotide. [1535] Embodiment 268. The combination of any one of embodiments 18-267, wherein the second set of polyribonucleotides comprises at least one polyribonucleotide that is an engineered polyribonucleotide. [1536] Embodiment 269. The combination of any one of embodiments 1-268, wherein at least one of the first set of polyribonucleotides is comprised in a first RNA construct, wherein the first RNA construct comprises in 5' to 3' order: (i) a 5' UTR; (ii) one or more HSV polyribonucleotides; (iii) a 3' UTR; and (iv) a polyA tail sequence. [1537] Embodiment 270. The combination of any one of embodiments 18-268, wherein at least one of the second set of polyribonucleotides is comprised in a second RNA construct, wherein the second RNA construct comprises in 5' to 3' order: (i) a 5' UTR; (ii) one or more HSV T-cell polyribonucleotides; (iii) a 3' UTR; and (iv) a polyA tail sequence. [1538] Embodiment 271. The combination of any one of embodiments 269 or 270, wherein [1539] (i) the 5' UTR of the first and/or second RNA construct comprises or consists of a modified human alpha-globin 5'-UTR; and [1540] (ii) the 3' UTR of the first and/or second RNA construct comprises or consists of a first sequence from the amino terminal enhancer of split (AES) messenger RNA and a second sequence from the mitochondrial encoded 12S ribosomal RNA. [1541] Embodiment 272. The combination of any one of embodiments 269-271, wherein the 5' UTR of the first and/or second RNA construct consists of a ribonucleic acid sequence according to SEQ ID NO: 152. [1542] Embodiment 273. The combination of any one of embodiments 269-272, wherein the 3' UTR of the first and/or second RNA construct consists of a ribonucleic acid sequence according to SEQ ID NOs: 158. [1543] Embodiment 274. The combination of any one of embodiments 269-273, wherein the polyA tail sequence is a split polyA tail sequence. [1544] Embodiment 275. The combination of embodiment 274, wherein the split polyA tail sequence consists of a ribonucleic acid sequence according to SEQ ID NO: 155. [1545] Embodiment 276. The combination of any one of embodiments 269-275, wherein the first and/or second RNA construct further comprise a 5' cap. [1546] Embodiment 277. The combination of embodiment 276, wherein the first and/or second RNA construct comprise a cap proximal sequence comprising positions +1, +2, +3, +4, and +5 of the polyribonucleotide. [1547] Embodiment 278. The combination of embodiment 276 or 277, wherein the 5' cap comprises or consists of m7(3’OMeG)(5')ppp(5')(2'OMeA1)pG2, wherein A1 is position +1 of the polyribonucleotide, and G2 is position +2 of the polyribonucleotide. [1548] Embodiment 279. The combination of embodiment 277 or 278, wherein the cap proximal sequence comprises A1 and G2 of the Cap1 structure, and a sequence comprising: A3A4U5 (SEQ ID NO: 150) at positions +3, +4 and +5 respectively of the polyribonucleotide. [1549] Embodiment 280. The combination of any one of embodiments 269-279, wherein the first and/or second RNA construct includes modified uridines in place of all uridines, optionally wherein modified uridines are each N1-methyl-pseudouridine. [1550] Embodiment 281. The combination of any one of embodiments 1-280, wherein: a first composition comprises at least one polyribonucleotide of the first set of polyribonucleotides. [1551] Embodiment 282. The composition of embodiment 281, wherein the first composition comprises (i) a polyribonucleotide that encoding an antigenic portion of HSV gC, (ii) a polyribonucleotide that encoding an antigenic portion of HSV gD, and (iii) a polyribonucleotide that encoding an antigenic portion of HSV gE. [1552] Embodiment 283. The combination of embodiment 281 or 282, wherein: a second composition comprises at least one polyribonucleotide encoding a polypeptide that comprises an HSV gB, variant thereof, or one or more antigenic portions thereof. [1553] Embodiment 284. The combination of any one of embodiments 281-283, wherein: a third composition comprises at least one polyribonucleotide of the second set of polyribonucleotides. [1554] Embodiment 285. The combination of any one of embodiments 281-284, wherein the first and/or second, and/or third composition further comprises lipid nanoparticles, polyplexes (PLX), lipidated polyplexes (LPLX), or liposomes, [1555] Embodiment 286. The combination of embodiment 285, wherein the first and/or second, and/or third composition is fully or partially encapsulated within the lipid nanoparticles, polyplexes (PLX), lipidated polyplexes (LPLX), or liposomes. [1556] Embodiment 287. The combination of any one of embodiments 281-286, wherein first and/or second, and/or third composition further comprises lipid nanoparticles. [1557] Embodiment 288. The combination of any one of embodiments 281-287, wherein first and/or second, and/or third composition is encapsulated within the lipid nanoparticles. [1558] Embodiment 289. The combination of any one of embodiments 281-288, wherein the first, second, and/or third composition comprises at least one pharmaceutically acceptable excipient. [1559] Embodiment 290. The combination of any one of embodiments 281-289, wherein the first, second and/or third composition comprises a cryoprotectant, optionally wherein the cryoprotectant is sucrose. [1560] Embodiment 291. The combination of any one of embodiments 281-290, wherein the first, second and/or third composition comprises an aqueous buffered solution, optionally wherein the aqueous buffered solution comprises one or more of Tris base, Tris HCl, NaCl, KCl, Na2HPO4, and KH2PO4. [1561] Embodiment 292. The combination of any one of embodiments 1-291 for use in the treatment of an HSV infection. [1562] Embodiment 293. The combination of any one of embodiments 1-291 for use in the prevention of an HSV infection. [1563] Embodiment 294. A method comprising administering a combination according to any one of embodiments 1-291 to a subject. [1564] Embodiment 295. A method comprising administering a first and/or second, and/or third composition according to any one of embodiments 281-284 to a subject. [1565] Embodiment 296. The method of embodiment 296 or the combination for use of embodiment 292 or 293, comprising administering two or more doses of the first, second and/or third composition to a subject. [1566] Embodiment 297. The method of embodiment 295, or the combination for use of embodiment 292 or 293, comprising administering three or more doses of the composition to a subject. [1567] Embodiment 298. The method of any one of embodiments 295-297, wherein the first composition and the second composition are administered on the same day. [1568] Embodiment 299. The method of any one of embodiments 295-297, wherein the first composition and the second composition are administered on different days. [1569] Embodiment 300. The method of any one of embodiments 295-299, wherein the first composition and the second composition are administered to the subject at different locations on the subject’s body. [1570] Embodiment 301. The method of embodiment 295, wherein the first composition and the third composition are administered on the same day. [1571] Embodiment 302. The method of embodiment 295, wherein the first composition and the third composition are administered on different days. [1572] Embodiment 303. The method of embodiment 301 or 302, wherein the first composition and the third composition are administered to the subject at different locations on the subject’s body. [1573] Embodiment 304. The method of embodiment 295, wherein the second composition and the third composition are administered on the same day. [1574] Embodiment 305. The method of embodiment 295, wherein the second composition and the third composition are administered on different days. [1575] Embodiment 306. The method of embodiment 304 or 305, wherein the second composition and the third composition are administered to the subject at different locations on the subject’s body. [1576] Embodiment 307. The method of any one of embodiments 294-306, wherein the method is a method of treating an HSV infection. [1577] Embodiment 308. The method of embodiment 294 or 307, wherein the method is a method of preventing an HSV infection. [1578] Embodiment 309. The method of any one of embodiments 294, 307, and 308, wherein the subject has or is at risk of developing an HSV infection. [1579] Embodiment 310. The method of any one of embodiments 294-309, wherein the subject is a human. [1580] Embodiment 311. The method of any one of embodiments 294-310, wherein administration induces an anti-HSV immune response in the subject. [1581] Embodiment 312. The method of embodiment 311, wherein the anti-HSV immune response in the subject comprises an adaptive immune response. [1582] Embodiment 313. The method of embodiment 311 or 312, wherein the anti-HSV immune response in the subject comprises a T-cell response. [1583] Embodiment 314. The method of embodiment 313, wherein the T-cell response is or comprises a CD4+ T cell response. [1584] Embodiment 315. The method of embodiment 313, wherein the T-cell response is or comprises a CD8+ T cell response. [1585] Embodiment 316. The method of embodiment 311 or 312, wherein the anti-HSV immune system response comprises a B-cell response. [1586] Embodiment 317. The method of any one of embodiments 311-316, wherein the anti-HSV immune system response comprises the production of antibodies directed against the one or more GP polypeptides and/or T- cell string polypeptides. [1587] Embodiment 318. Use of the combination of any one of embodiments 1-293, in the treatment of an HSV infection. [1588] Embodiment 319. Use of the combination of any one of embodiments 1-293 in the prevention of an HSV infection. EQUIVALENTS [1589] It is to be understood that while the disclosure has been described in conjunction with the detailed description thereof, the foregoing description is intended to illustrate and not limit the scope of the invention, which is defined by the scope of the appended claims. Other aspects, advantages, and modifications are within the scope of the following claims.

Claims

CLAIMS 1. A combination comprising a plurality of polyribonucleotides, wherein the plurality of polyribonucleotides comprises a first set of polyribonucleotides that encode one or more glycoprotein (GP) polypeptides, wherein the one or more GP polypeptides each comprise an HSV glycoprotein or an antigenic portion thereof.
2. The combination of claim 1, wherein the first set of polyribonucleotides comprises: (i) a polyribonucleotide encoding an HSV glycoprotein C (gC) or an antigenic portion thereof, (ii) a polyribonucleotide encoding an HSV glycoprotein D (gD) or an antigenic portion thereof, (iii) a polyribonucleotide encoding an HSV glycoprotein E (gE) or an antigenic portion thereof, or (iv) a combination thereof.
3. The combination of claim 1 or 2, wherein the first set of polyribonucleotides further comprises at least one polyribonucleotide encoding a GP polypeptide that comprises an HSV glycoprotein B (gB), variant thereof, or one or more antigenic portions thereof.
4. The combination of any one of claims 1-3, wherein the combination further comprises a second set of polyribonucleotides that encode one or more T-cell string polypeptides, wherein the one or more T-cell string polypeptides each comprise one or more HSV T-cell antigens or antigenic portions thereof.
5. The combination of claim 4, wherein the second set of polyribonucleotides encodes at least one T-cell string polypeptide that comprises one or more HSV T-cell antigens or antigenic portions thereof, wherein the one or more HSV T-cell antigens comprises: (i) one or more HSV RS1 polypeptides or antigenic portions thereof, (ii) one or more HSV RL2 polypeptides or antigenic portions thereof, (iii) one or more HSV UL1 polypeptides or antigenic portions thereof, (iv) one or more HSV UL5 polypeptides or antigenic portions thereof, (v) one or more HSV UL9 polypeptides or antigenic portions thereof, (vi) one or more HSV UL19 polypeptides or antigenic portions thereof, (vii) one or more HSV UL21 polypeptides or antigenic portions thereof, (viii) one or more HSV UL25 polypeptides or antigenic portions thereof, (ix) one or more HSV UL27 polypeptides or antigenic portions thereof, (x) one or more HSV UL29 polypeptides or antigenic portions thereof, (xi) one or more HSV UL30 polypeptides or antigenic portions thereof, (xii) one or more HSV UL39 polypeptides or antigenic portions thereof, (xiii) one or more HSV UL40 polypeptides or antigenic portions thereof, (xiv) one or more HSV UL46 polypeptides or antigenic portions thereof, (xv) one or more HSV UL47 polypeptides or antigenic portions thereof, (xvi) one or more HSV UL48 polypeptides or antigenic portions thereof, (xvii) one or more HSV UL49 polypeptides or antigenic portions thereof, (xviii) one or more HSV UL52 polypeptides or antigenic portions thereof, (xix) one or more HSV UL54 polypeptides or antigenic portions thereof, (xx) one or more HSV US10 polypeptides or antigenic portions thereof, (xxi) one or more HSV US12 polypeptides or antigenic portions thereof, (xxii) one or more HSV UL26 polypeptides or antigenic portions thereof, (xxiii) one or more HSV UL50 polypeptides or antigenic portions thereof, or (xxiv) a combination thereof.
6. The combination of claim 4 or 5, wherein the second set of polyribonucleotides comprises a polyribonucleotide that encodes a polypeptide that comprises one or more HSV RL2 polypeptides or antigenic portions thereof, one or more HSV RS1 polypeptides or antigenic portions thereof, and one or more HSV UL54 polypeptides or antigenic portions thereof.
7. The combination of claim 4or 5, wherein the second set of polyribonucleotides comprises a polyribonucleotide that encodes a polypeptide that comprises one or more HSV UL29 polypeptides or antigenic portions thereof, one or more HSV UL39 polypeptides or antigenic portions thereof, one or more HSV UL49 polypeptides or antigenic portions thereof, and one or more HSV UL9 polypeptides or antigenic portions thereof.
8. The combination of claim 4 or 5, wherein the second set of polyribonucleotides comprises a polyribonucleotide that encodes a polypeptide that comprises one or more HSV UL30 polypeptides or antigenic portions thereof, one or more HSV UL40 polypeptides or antigenic portions thereof, one or more HSV UL5 polypeptides or antigenic portions thereof, and one or more HSV UL52 polypeptides or antigenic portions thereof.
9. The combination of claim 4 or 5, wherein the second set of polyribonucleotides comprises a polyribonucleotide that encodes a polypeptide that comprises one or more HSV UL1 polypeptides or antigenic portions thereof, one or more HSV UL19 polypeptides or antigenic portions thereof, one or more HSV UL21 polypeptides or antigenic portions thereof, one or more HSV UL27 polypeptides or antigenic portions thereof, one or more HSV UL46 polypeptides or antigenic portions thereof, one or more HSV UL47 polypeptides or antigenic portions thereof, one or more UL48 polypeptides or antigenic portions thereof, and one or more HSV UL25 polypeptides or antigenic portions thereof.
10. The combination of claim 4 or 5, wherein the second set of polyribonucleotides comprises a polyribonucleotide that encodes a polypeptide that comprises one or more HSV US1 polypeptides or antigenic fragments thereof, one or more HSV US8 polypeptides or antigenic fragments thereof, one or more HSV US12 polypeptides or antigenic fragments thereof, one or more HSV UL50 polypeptides or antigenic fragments thereof, one or more HSV UL26 polypeptides or antigenic fragments thereof, and one or more HSV US10 polypeptides or antigenic fragments thereof.
11. The combination of claim 4 or 5, wherein the polypeptide comprises one or more HSV RL2 polypeptides or antigenic fragments thereof, one or more HSV UL54 polypeptides or antigenic fragments thereof, one or more HSV UL47 polypeptides or antigenic fragments thereof, one or more HSV UL46 polypeptides or antigenic fragments thereof, and one or more HSV UL21 polypeptides or antigenic fragments thereof.
12. The combination of claim 4 or 5, wherein the polypeptide comprises one or more HSV RL2 polypeptides or antigenic fragments thereof, one or more HSV UL54 polypeptides or antigenic fragments thereof, one or more HSV UL9 polypeptides or antigenic fragments thereof, one or more HSV UL39 polypeptides or antigenic fragments thereof, and one or more HSV UL5 polypeptides or antigenic fragments thereof.
13. The combination of claim 4 or 5, wherein the polypeptide comprises one or more HSV UL47 polypeptides or antigenic fragments thereof, one or more HSV UL46 polypeptides or antigenic fragments thereof, one or more HSV UL21 polypeptides or antigenic fragments thereof, one or more HSV UL5.2 polypeptides or antigenic fragments thereof, one or more HSV UL40 polypeptides or antigenic fragments thereof, one or more HSV UL30.1 polypeptides or antigenic fragments thereof, and one or more HSV UL29 polypeptides or antigenic fragments thereof.
14. The combination of claim 4 or 5, wherein the polypeptide comprises one or more HSV RL2.1 polypeptides or antigenic fragments thereof, one or more HSV UL54 polypeptides or antigenic fragments thereof, one or more HSV UL5.2 polypeptides or antigenic fragments thereof, one or more HSV UL40 polypeptides or antigenic fragments thereof, one or more HSV UL47 polypeptides or antigenic fragments thereof, and one or more HSV UL46 polypeptides or antigenic fragments thereof.
15. The combination of claim 4 or 5, wherein the polypeptide comprises one or more HSV RL2.1 polypeptides or antigenic fragments thereof, one or more HSV RS1 polypeptides or antigenic fragments thereof, and one or more HSV UL54 polypeptides or antigenic fragments thereof.
16. The combination of claim 4 or 5, wherein the polypeptide comprises one or more HSV UL29 polypeptides or antigenic fragments thereof, one or more HSV UL39 polypeptides or antigenic fragments thereof, and one or more HSV UL9 polypeptides or antigenic fragments thereof.
17. The combination of claim 4 or 5, wherein the polypeptide comprises one or more HSV UL5.1 polypeptides or antigenic fragments thereof, one or more HSV UL5.2 polypeptides or antigenic fragments thereof, one or more HSV UL40 polypeptides or antigenic fragments thereof, and one or more HSV UL30.1 polypeptides or antigenic fragments thereof.
18. The combination of claim 4 or 5, wherein the polypeptide comprises one or more HSV UL47 polypeptides or antigenic fragments thereof, one or more HSV UL46 polypeptides or antigenic fragments thereof, and one or more HSV UL21 polypeptides or antigenic fragments thereof.
19. The combination of claim 4 or 5, wherein the polypeptide comprises one or more HSV UL29 polypeptides or antigenic fragments thereof, one or more HSV UL39 polypeptides or antigenic fragments thereof, one or more HSV UL9 polypeptides or antigenic fragments thereof, one or more HSV UL5.1 polypeptides or antigenic fragments thereof, one or more HSV UL5.2 polypeptides or antigenic fragments thereof, one or more HSV UL40 polypeptides or antigenic fragments thereof, and one or more HSV UL30.1 polypeptides or antigenic fragments thereof.
20. The combination of claim 4 or 5, wherein the polypeptide comprises one or more HSV RL2.1 polypeptides or antigenic fragments thereof, one or more HSV RS1 polypeptides or antigenic fragments thereof, one or more HSV UL54 polypeptides or antigenic fragments thereof, one or more HSV UL47 polypeptides or antigenic fragments thereof, one or more HSV UL46 polypeptides or antigenic fragments thereof, and one or more HSV UL21 polypeptides or antigenic fragments thereof.
21. The combination of claim 4 or 5, wherein the polypeptide comprises one or more HSV UL5.1 polypeptides or antigenic fragments thereof, one or more HSV UL5.2 polypeptides or antigenic fragments thereof, one or more HSV UL40 polypeptides or antigenic fragments thereof, one or more HSV UL30.1 polypeptides or antigenic fragments thereof, one or more HSV UL47 polypeptides or antigenic fragments thereof, one or more HSV UL46 polypeptides or antigenic fragments thereof, and one or more HSV UL21 polypeptides or antigenic fragments thereof.
22. The combination of any one of claims 4-21, wherein at least one of the one or more T-cell string polypeptides comprises an HSV-1 gD secretory signal, one or more RL2 polypeptides or antigenic fragments thereof, one or more RS1 polypeptides or antigenic fragments thereof, one or more UL54 polypeptides or antigenic fragments thereof, and a MITD.
23. The combination of any one of claims 4-21, wherein at least one of the one or more T-cell string polypeptides comprises an HSV-1 gD secretory signal, an UL29 polypeptide or antigenic fragment thereof, a linker, an UL39 polypeptide or antigenic fragment thereof, a linker, an UL49 polypeptide or antigenic fragment thereof, a linker, a UL9 polypeptide or antigenic fragment thereof, and a linker.
24. The combination of any one of claims 4-21, wherein at least one of the one or more T-cell string polypeptides comprises an HSV-1 gD secretory signal, one or more HSV UL30 polypeptides or antigenic fragments thereof, one or more HSV UL40 polypeptides or antigenic fragments thereof, one or more HSV UL5 polypeptides or antigenic fragments thereof, one or more HSV UL52 polypeptides or antigenic fragments thereof, and a MITD.
25. The combination of any one of claims 4-21, wherein at least one of the one or more T-cell string polypeptides comprises an HSV-1 gD secretory signal, one or more HSV UL1 polypeptides or antigenic fragments thereof, one or more HSV UL19 polypeptides or antigenic fragments thereof, one or more HSV UL21 polypeptides or antigenic fragments thereof, one or more HSV UL27 polypeptides or antigenic fragments thereof, one or more HSV UL46 polypeptides or antigenic fragments thereof, one or more HSV UL47 polypeptides or antigenic fragments thereof, one or more UL48 polypeptides or antigenic fragments thereof, one or more HSV UL25 polypeptides or antigenic fragments thereof, and a MITD.
26. The combination of any one of claims 4-21, wherein at least one of the one or more T-cell string polypeptides comprises an HSV-2 gD secretory signal, one or more RL2 polypeptides or antigenic fragments thereof, one or more RS1 polypeptides or antigenic fragments thereof, one or more UL54 polypeptides or antigenic fragments thereof, and a MITD.
27. The combination of any one of claims 4-21, wherein at least one of the one or more T-cell string polypeptides comprises an HSV-1 gD secretory signal, one or more HSV US1 polypeptides or antigenic fragments thereof, one or more HSV US8 polypeptides or antigenic fragments thereof, one or more HSV US12 polypeptides or antigenic fragments thereof, one or more HSV UL50 polypeptides or antigenic fragments thereof, one or more HSV UL26 polypeptides or antigenic fragments thereof, and one or more HSV US10 polypeptides or antigenic fragments thereof, and a MITD.
28. The combination of any one of claims 4-21, wherein at least one of the one or more T-cell string polypeptides comprises an HSV-1 gD secretory signal, one or more RL2 polypeptides or antigenic fragments thereof, one or more RS1 polypeptides or antigenic fragments thereof, one or more UL54 polypeptides or antigenic fragments thereof, and an HSV-1 gD transmembrane region.
29. The combination of any one of claims 4-21, wherein at least one of the one or more T-cell string polypeptides comprises an HSV-1 gD secretory signal, one or more RL2 polypeptides or antigenic fragments thereof, one or more RS1 polypeptides or antigenic fragments thereof, one or more UL54 polypeptides or antigenic fragments thereof, and a VSV-G transmembrane region.
30. The combination of any one of claims 1-2 and 4-5, wherein the polypeptide comprises an HSV-1 gD secretory signal, one or more HSV RL2 polypeptides or antigenic fragments thereof, one or more HSV UL54 polypeptides or antigenic fragments thereof, one or more HSV UL47 polypeptides or antigenic fragments thereof, one or more HSV UL46 polypeptides or antigenic fragments thereof, one or more HSV UL21 polypeptides or antigenic fragments thereof, and a MITD.
31. The combination of any one of claims 1-2, wherein the polypeptide comprises an HSV-1 gD secretory signal, one or more HSV UL29 polypeptides or antigenic fragments thereof, one or more HSV UL39 polypeptides or antigenic fragments thereof, one or more HSV UL9 polypeptides or antigenic fragments thereof, one or more HSV UL5 polypeptides or antigenic fragments thereof, one or more HSV UL40 polypeptides or antigenic fragments thereof, one or more HSV UL30 polypeptides or antigenic fragments thereof, and a MITD.
32. The combination of any one of claims 1-2, wherein the polypeptide comprises an HSV-1 gD secretory signal, one or more HSV RL2 polypeptides or antigenic fragments thereof, one or more HSV UL54 polypeptides or antigenic fragments thereof, one or more HSV UL9 polypeptides or antigenic fragments thereof, one or more HSV UL39 polypeptides or antigenic fragments thereof, one or more HSV UL5 polypeptides or antigenic fragments thereof, and a MITD.
33. The combination of any one of claims 1-2, wherein the polypeptide comprises an HSV-1 gD secretory signal, one or more HSV UL47 polypeptides or antigenic fragments thereof, one or more HSV UL46 polypeptides or antigenic fragments thereof, one or more HSV UL21 polypeptides or antigenic fragments thereof, one or more HSV UL5.2 polypeptides or antigenic fragments thereof, one or more HSV UL40 polypeptides or antigenic fragments thereof, one or more HSV UL30.1 polypeptides or antigenic fragments thereof, one or more HSV UL29 polypeptides or antigenic fragments thereof, and a MITD.
34. The combination of any one of claims 1-2, wherein the polypeptide comprises an HSV-1 gD secretory signal, one or more HSV RL2.1 polypeptides or antigenic fragments thereof, one or more HSV UL54 polypeptides or antigenic fragments thereof, one or more HSV UL5.2 polypeptides or antigenic fragments thereof, one or more HSV UL40 polypeptides or antigenic fragments thereof, one or more HSV UL47 polypeptides or antigenic fragments thereof, one or more HSV UL46 polypeptides or antigenic fragments thereof, and a MITD.
35. The combination of any one of claims 1-2, wherein the polypeptide comprises an HSV-1 gD secretory signal, one or more HSV RL2.1 polypeptides or antigenic fragments thereof, one or more HSV RS1 polypeptides or antigenic fragments thereof, one or more HSV UL54 polypeptides or antigenic fragments thereof, and a MITD.
36. The combination of any one of claims 1-2, wherein the polypeptide comprises an HSV-1 gD secretory signal, one or more HSV UL29 polypeptides or antigenic fragments thereof, one or more HSV UL39 polypeptides or antigenic fragments thereof, and one or more HSV UL9 polypeptides or antigenic fragments thereof, and a MITD.
37. The combination of any one any one of claims 1-2, wherein the polypeptide comprises an HSV-1 gD secretory signal, one or more HSV UL5.1 polypeptides or antigenic fragments thereof, one or more HSV UL5.2 polypeptides or antigenic fragments thereof, one or more HSV UL40 polypeptides or antigenic fragments thereof, one or more HSV UL30.1 polypeptides or antigenic fragments thereof, and a MITD.
38. The polyribonucleotide of any one of claims 1-2, wherein the polypeptide comprises an HSV-1 gD secretory signal, one or more HSV UL47 polypeptides or antigenic fragments thereof, one or more HSV UL46 polypeptides or antigenic fragments thereof, one or more HSV UL21 polypeptides or antigenic fragments thereof, and a MITD.
39. The combination of any one of claims 1-2, wherein the polypeptide comprises an HSV-1 gD secretory signal, one or more HSV UL29 polypeptides or antigenic fragments thereof, one or more HSV UL39 polypeptides or antigenic fragments thereof, one or more HSV UL9 polypeptides or antigenic fragments thereof, one or more HSV UL5.1 polypeptides or antigenic fragments thereof, one or more HSV UL5.2 polypeptides or antigenic fragments thereof, one or more HSV UL40 polypeptides or antigenic fragments thereof, one or more HSV UL30.1 polypeptides or antigenic fragments thereof, and a MITD.
40. The combination of any one of claims 1-2, wherein the polypeptide comprises an HSV-1 gD secretory signal, comprises one or more HSV RL2.1 polypeptides or antigenic fragments thereof, one or more HSV RS1 polypeptides or antigenic fragments thereof, one or more HSV UL54 polypeptides or antigenic fragments thereof, one or more HSV UL47 polypeptides or antigenic fragments thereof, one or more HSV UL46 polypeptides or antigenic fragments thereof, one or more HSV UL21 polypeptides or antigenic fragments thereof, and a MITD.
41. The combination of any one of claims 1-2, wherein the polypeptide comprises an HSV-1 gD secretory signal, comprises one or more HSV UL5.1 polypeptides or antigenic fragments thereof, one or more HSV UL5.2 polypeptides or antigenic fragments thereof, one or more HSV UL40 polypeptides or antigenic fragments thereof, one or more HSV UL30.1 polypeptides or antigenic fragments thereof, one or more HSV UL47 polypeptides or antigenic fragments thereof, one or more HSV UL46 polypeptides or antigenic fragments thereof, one or more HSV UL21 polypeptides or antigenic fragments thereof, and a MITD.
42. The combination of any one of claims 1-41, wherein at least one of the first set of polyribonucleotides is comprised in a first and/or second RNA construct, wherein the first and/or second RNA construct comprises in 5' to 3' order: (i) a 5' UTR; (ii) one or more HSV polyribonucleotides; (iii) a 3' UTR; and (iv) a polyA tail sequence.
43. The combination of any one of claims 1-42, wherein: a first composition comprises at least one polyribonucleotide of the first set of polyribonucleotides.
44. The composition of claim 43, wherein the first composition comprises (i) a polyribonucleotide that encoding an antigenic portion of HSV gC, (ii) a polyribonucleotide that encoding an antigenic portion of HSV gD, and (iii) a polyribonucleotide that encoding an antigenic portion of HSV gE.
45. The combination of claim 43 or 44, wherein: a second composition comprises at least one polyribonucleotide encoding a polypeptide that comprises an HSV gB, variant thereof, or one or more antigenic portions thereof.
46. The combination of any one of claims 43-45, wherein: a third composition comprises at least one polyribonucleotide of the second set of polyribonucleotides.
47. The combination of any one of claims 43-46, wherein the first and/or second, and/or third composition further comprises lipid nanoparticles, polyplexes (PLX), lipidated polyplexes (LPLX), or liposomes,
48. A method comprising administering a combination according to any one of claims 1-47 to a subject.
49. A method comprising administering a first and/or second, and/or third composition according to any one of claims 43-46 to a subject.
50. Use of the combination of any one of claims 1-47, in the treatment or prevention of an HSV infection.
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