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WO2020056411A1 - Virus herpès simplex 1 (vhs-1) de recombinaison - Google Patents

Virus herpès simplex 1 (vhs-1) de recombinaison Download PDF

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Publication number
WO2020056411A1
WO2020056411A1 PCT/US2019/051303 US2019051303W WO2020056411A1 WO 2020056411 A1 WO2020056411 A1 WO 2020056411A1 US 2019051303 W US2019051303 W US 2019051303W WO 2020056411 A1 WO2020056411 A1 WO 2020056411A1
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Prior art keywords
hsv
glycoprotein
virion
recombinant
virus
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Rory T. WINTER
Andreas STUERMER
Gabriel Alexander LICINA
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Excell Biotech LLC
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Excell Biotech LLC
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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
    • 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
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/005—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from viruses
    • 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
    • C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
    • C12N15/09—Recombinant DNA-technology
    • C12N15/63—Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
    • C12N15/79—Vectors or expression systems specially adapted for eukaryotic hosts
    • C12N15/85—Vectors or expression systems specially adapted for eukaryotic hosts for animal cells
    • C12N15/86—Viral vectors
    • 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
    • C12N7/00—Viruses; Bacteriophages; Compositions thereof; Preparation or purification thereof
    • 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
    • C12N2710/16622—New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes
    • 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
    • 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
    • C12N2710/16641—Use of virus, viral particle or viral elements as a vector
    • C12N2710/16643—Use of virus, viral particle or viral elements as a vector viral genome or elements thereof as genetic vector

Definitions

  • Herpes simplex virus types 1 and 2 are a significant health problem impacting all countries and communities around the world and contributing greatly to the HIV epidemic. Vaccines are needed for these infections, as currently there is no effective vaccine for HSV-l and/or HSV-2.
  • HSV-l is the primary cause of infectious blindness and oral HSV-l cold sores and is the leading cause for new genital HSV infections worldwide.
  • HSV-2 is the primary cause of genital ulcers.
  • HSV-l is now more commonly identified in association with genital tract disease in developed countries.
  • Oral and Genital herpes is a recurrent, lifelong disease that can stigmatize and psychologically impacts those affected.
  • HSV-2 gD subunit gD-2 gD subunit
  • the present invention addresses this need for new and improved HSV-l and HSV-2 vaccines.
  • HSV-l has a relatively large, double-stranded, linear DNA genome (see SEQ ID NO:3) within an icosahedral capsid.
  • the capsid is enveloped in a lipid bilayer called the envelope.
  • the envelope is joined to the capsid by means of a tegument.
  • the complete HSV-l particle is known as the HSV-l virion.
  • the HSV genome (see SEQ ID NO:3 for example) consists of two unique segments, named unique long (UL) and unique short (US), as well as terminal inverted repeats found to the two ends of them named repeat long (RL) and repeat short (RS).
  • the HSV genome encodes five unique short (US) glycoproteins - glycoprotein G (US4 orf), glycoprotein J (US5 orf), glycoprotein D (US6 orf), glycoprotein I (US7 orf), and glycoprotein E (US8 orf).
  • Certain embodiments are directed to a herpes simplex virus-l (HSV-l) having a genomic deletion of an HSV-l glycoprotein.
  • the genomic deletion is of an HSV-l glycoprotein D-encoding gene (Us6).
  • the HSV-l virion comprises a heterologous glycoprotein, for example a HSV-2 glycoprotein.
  • the heterologous glycoprotein in HSV-2 glycoprotein D is directed to a herpes simplex virus-l (HSV-l) having a genomic deletion of an HSV-l glycoprotein.
  • the genomic deletion is of an HSV-l glycoprotein D-encoding gene (Us6).
  • the HSV-l virion comprises a heterologous glycoprotein, for example a HSV-2 glycoprotein.
  • the heterologous glycoprotein in HSV-2 glycoprotein D is particularly preferred embodiments the heterologous glycoprotein in HSV-2 glycoprotein D.
  • Certain embodiments are directed to an isolated cell comprising a recombinant HSV-l or HSV-2 genome as described herein and/or an expression cassette encoding one or more recombinant HSV genome and/or one or more HSV protein as described herein.
  • the cell is not present in a human being, is a cell in culture, or is an isolated cell.
  • Certain embodiments are directed to vaccine compositions comprising recombinant HSV-l genome, and/or a HSV-l virus or HSV virion as described herein.
  • Certain embodiments are directed to compositions comprising a recombinant HSV-l virus as described herein, or a virion as described herein, wherein the genome of the virus or virion comprises at least a deletion of a second gene, wherein the second gene is necessary for HSV-l viral replication or virulence.
  • compositions comprising a recombinant HSV-l virus as described herein, or the virion as described herein, and a pharmaceutically acceptable carrier.
  • Certain embodiments are directed to methods of eliciting an immune response in a subject comprising administering to the subject an amount of a recombinant HSV-l genome as described herein; a virion thereof as described herein, a vaccine as described herein; a composition as described herein; or a pharmaceutical composition as described herein, in an amount effective to elicit an immune response in a subject.
  • Other embodiments are directed to methods of treating an HSV-l, HSV-2, or HSV-l and HSV-2 co-infection in a subject or treating a disease caused by an HSV-l, HSV-2 or co- infection in a subject comprising administering to the subject an amount of the recombinant HSV-l virus as described herein; a virion thereof as described herein, a vaccine as described herein; a composition as described herein; or a pharmaceutical composition as described herein, in an amount effective to treat an HSV-l, HSV-2 or co-infection or treat a disease caused by an HSV-l, HSV-2 or co-infection in a subject.
  • Certain embodiments are directed to methods of vaccinating a subject for HSV-l, HSV-2 or co-infection comprising administering to the subject an amount of the recombinant HSV-l virus genome as described herein; a virion thereof as described herein, a vaccine as described herein; a composition as described herein; or a pharmaceutical composition as described herein, in an amount effective to vaccinate a subject for HSV-l, HSV-2, or co- infection.
  • Certain embodiments are directed to methods of immunizing a subject against HSV-l, HSV-2, or co-infection comprising administering to the subject an amount of the recombinant HSV-l virus genome as described herein; a virion thereof as described herein, a vaccine as described herein; a composition as described herein; or a pharmaceutical composition as described herein, in an amount effective to immunize a subject against HSV-l, HSV-2, or co- infection.
  • the amount of recombinant HSV-l is an amount of pfu of recombinant HSV-l effective to achieve the stated aim.
  • a vaccine, composition, and/or a pharmaceutical composition can have 5x 1o 1 PFU, 5x l0 2 PFU, 5x l0 3 PFU, 5x l0 4 PFU, 5x l0 6 PFU, 5 x 10 7 PFU, 5 x 10 8 PFU, or 5 x 10 9 PFU HS V- 1 virion.
  • Certain embodiments are directed to methods of producing a virion of a recombinant herpes simplex virus-l (HSV-l), the genome having a deletion of an HSV-l glycoprotein D- encoding gene and comprising a heterologous HSV-l or HSV-2 glycoprotein D on the virion lipid bilayer.
  • HSV-l herpes simplex virus-l
  • the method comprises infecting a cell expressing a heterologous nucleic acid encoding a heterologous glycoprotein, e.g., a HSV-l or HSV-2 glycoprotein D, with a recombinant herpes simplex virus-l (HSV-l) having a genomic deletion of an HSV-l glycoprotein D-encoding gene under conditions permitting replication of the recombinant herpes simplex virus-l (HSV-l) and recovering a HSV-l virion produced by the cells.
  • a heterologous nucleic acid encoding a heterologous glycoprotein
  • a heterologous glycoprotein e.g., a HSV-l or HSV-2 glycoprotein D
  • HSV-l herpes simplex virus-l
  • HSV-l herpes simplex virus-l
  • the virion being produced using a cell comprising an expression cassette encoding the heterologous HSV-2 glycoprotein D, the HSV-2 glycoprotein D coding region having an intron as a safety switch (i.e., an intron safety switch).
  • the safety switch intron being positioned with in the coding region of a heterologous HSV glycoprotein D. The presence of the intron reduces the possibility of recombination and reversion during virion production or other stages of the viral lifecycle.
  • HSV-l herpes simplex virus-l
  • the HSV-2 glycoprotein D being produced from an expression cassette where the HSV-2 glycoprotein coding region comprises an intron, i.e., an intron safety switch.
  • the intron safety switch is to prevent viral recombination by minimizing or reducing recombination between wild type sequences and recombinant sequences.
  • Certain embodiments are directed to a recombinant nucleic acid, or nucleic acid vector encoding the same, having the same sequence as a genome of a wild-type HSV-l (e.g., SEQ ID NO:3) having a deletion of the coding region for HSV-l glycoprotein D.
  • a wild-type HSV-l e.g., SEQ ID NO:3
  • HSV-l herpes simplex virus- 1
  • HSV-l herpes simplex virus- 1
  • the recombinant HSV-l being useful for treating or preventing an HSV-l, HSV-2, or co-infection in a subject.
  • Certain embodiments are directed to a recombinant HSV-l virion having a genomic deletion of glycoprotein D gene for use in treating or preventing an HSV-l, HSV-2 or co- infection in a subject.
  • the virion does contain HSV-l glycoprotein D.
  • the virion contains and presents a heterologous glycoprotein, e.g., HSV-2 glycoprotein D.
  • the recombinant herpes simplex virus-l has a genomic deletion of an HSV-l glycoprotein D-encoding gene.
  • Certain embodiments are directed to an isolated cell comprising a HSV-l recombinant genome described herein and an expression cassette encoding and/or expressing a heterologous glycoprotein D.
  • the cell expressing the component needed to form a recombinant virus and produce a recombinant HSV-l virion as described herein.
  • the cell is not present in a human being.
  • the cell is in cell culture under the appropriate conditions for virion production.
  • compositions comprising a recombinant virus or virion as described herein.
  • compositions comprising a virus or virion as described herein, wherein the genome of the virus or virion comprises at least a deletion of a second gene, wherein the second gene is necessary for HSV-l viral replication.
  • Certain embodiments are directed to methods of eliciting an immune response in a subject comprising administering to the subject an amount of a virus or virion as described herein, a vaccine as described herein, a composition as described herein, or a pharmaceutical composition as described herein, in an amount effective to elicit an immune response in a subject.
  • Certain embodiments are directed to methods of treating an HSV-l infection in a subject or treating a disease caused by an HSV-l infection in a subject comprising administering to the subject an amount of a virus or virion as described herein, a vaccine as described herein, a composition as described herein, or a pharmaceutical composition as described herein, in an amount effective to treat an HSV-l infection or treat a disease caused by an HSV-l infection in a subject.
  • Certain embodiments are directed to methods of immunizing a subject against HSV-l infection comprising administering to the subject an amount of a virus or virion as described herein, a vaccine as described herein, a composition as described herein, or a pharmaceutical composition as described herein, in an amount effective to immunize a subject against HSV-l.
  • Certain embodiments are directed to methods of producing a recombinant herpes simplex virus-l (HSV-l) virion, having a genomic deletion of an HSV-l glycoprotein D- encoding gene and comprising a heterologous HSV-2 glycoprotein D in the virion lipid bilayer.
  • HSV-l herpes simplex virus-l
  • the method comprising infecting a cell comprising a heterologous nucleic acid encoding a HSV- 2 glycoprotein D (e.g., an HSV-2 glycoprotein D expression vector) with a recombinant herpes simplex virus-l (HSV-l) genome having a deletion of an HSV-l glycoprotein D-encoding gene under conditions permitting replication of the recombinant herpes simplex virus-l (HSV-l) and recovering a recombinant HSV-l virion comprising a heterologous HSV-2 glycoprotein D on the lipid bilayer of the virion produced by the cell.
  • a HSV- 2 glycoprotein D e.g., an HSV-2 glycoprotein D expression vector
  • Certain embodiments are directed to methods of producing a recombinant herpes simplex virus-l (HSV-l) virion having a genomic deletion of an HSV-l glycoprotein D- encoding gene and comprising a heterologous non-HSV-l surface glycoprotein on the lipid bilayer.
  • HSV-l herpes simplex virus-l
  • the method comprising introducing, infecting, or transfecting a cell comprising a heterologous nucleic acid encoding the non-HSV-l surface glycoprotein with a recombinant herpes simplex virus-l (HSV-l) having a deletion of an HSV-l glycoprotein D-encoding gene under conditions permitting replication of the recombinant herpes simplex virus-l (HSV-l) and recovering a recombinant HSV-l virion comprising a heterologous non-HSV-l surface glycoprotein on a lipid bilayer thereof produced by the cell.
  • a recombinant herpes simplex virus-l HSV-l having a deletion of an HSV-l glycoprotein D-encoding gene
  • Certain embodiments are directed to a recombinant nucleic acid having nucleic acid sequence HSV-l (for example see SEQ ID NO:3) except that the sequence does not comprise a sequence encoding an HSV-l glycoprotein D, i.e., genomic deletion of HSV-l glycoprotein D.
  • Certain embodiments are directed to a recombinant herpes simplex virus-l (HSV-l) having a genomic deletion of an HSV-l glycoprotein D gene and further comprising a heterogenous or heterologous antigen of a pathogen.
  • HSV-l herpes simplex virus-l
  • Also provided is a method of inducing antibody dependent cell mediated cytotoxicity (ADCC) against an antigenic target in a subject comprising administering to the subject a recombinant herpes simplex virus-l (HSV-l) presenting an antigen, as described herein in an amount effective to induce antibody dependent cell mediated cytotoxicity (ADCC) against an antigenic target.
  • a recombinant herpes simplex virus-l HSV-l
  • chimeric or“recombinant” as used herein with reference indicates that the nucleic acid, protein, vector, or cell has been modified by the introduction of a heterologous nucleic acid or protein or the alteration of a native nucleic acid or protein.
  • recombinant vectors include nucleic acid sequences that are not found within the native (non-chimeric or non-recombinant) form of the vector.
  • a chimeric adenoviral expression vector refers to an adenoviral expression vector comprising a nucleic acid sequence encoding a heterologous polypeptide.
  • An “expression vector” is a nucleic acid construct, generated recombinantly or synthetically, with a series of specified nucleic acid elements that permit transcription of a particular nucleic acid in a host cell.
  • the expression vector can be a plasmid, virus, or nucleic acid fragment.
  • the expression vector includes a nucleic acid to be transcribed operably linked to a promoter.
  • promoter refers to an array of nucleic acid control sequences that direct transcription of a nucleic acid.
  • a promoter includes necessary nucleic acid sequences near the start site of transcription, such as, in the case of a polymerase II type promoter, a TATA element.
  • a promoter can optionally includes distal enhancer or repressor elements. Promoters include constitutive and inducible promoters.
  • a “constitutive” promoter is a promoter that is active under most environmental and developmental conditions.
  • An“inducible” promoter is a promoter that is active under environmental or developmental regulation.
  • operably linked refers to a functional linkage between a nucleic acid expression control sequence (such as a promoter, or array of transcription factor binding sites) and a second nucleic acid sequence, wherein the expression control sequence directs transcription of the nucleic acid corresponding to the second sequence.
  • a nucleic acid expression control sequence such as a promoter, or array of transcription factor binding sites
  • An“antigen” refers to a protein or part of a polypeptide chain that can be recognized by T cell receptors and/or antibodies. Typically, antigens are derived from bacterial, viral, or fungal proteins.
  • the term“epitope” refers to the portion of the antigen that is recognized by the T cell receptor or antibody. Typically, the term antigen is interpreted to be broader than the term epitope. For example, a T cell receptor or antibody might be specific for a given antigen (e.g., protein X), and recognize or bind to only a few amino acids of protein X, the epitope.
  • A“T cell epitope” is recognized or bound by a T cell receptor.
  • An “immunogenically effective dose or amount” of the presently disclosed compositions is an amount that elicits or modulates an immune response specific for the desired polypeptide, e.g., HSV-2 antigen.
  • Immune responses include humoral immune responses and cell-mediated immune responses.
  • An immunogenic composition can be used therapeutically or prophylactically to treat or prevent HSV infection and outbreak at any stage.
  • “Humoral immune responses” are mediated by cell free components of the blood, i.e., plasma or serum; transfer of the serum or plasma from one individual to another transfers immunity.
  • Cell mediated immune responses are mediated by antigen specific lymphocytes; transfer of the antigen specific lymphocytes from one individual to another transfers immunity.
  • A“therapeutic dose” or“therapeutically effective amount” or“effective amount” of a viral vector or a composition comprising a viral vector is an amount of the vector or composition comprising the vector which prevents, alleviates, abates, or reduces the severity of symptoms of HSV.
  • nucleic acid and“polynucleotide” are used interchangeably herein to refer to deoxyribonucleotide or ribonucleotide polymers in either single- or double-stranded form.
  • A“nucleotide” typically refers to the monomer.
  • the terms encompasses nucleic acids containing known nucleotide analogs or modified backbone residues or linkages, which are synthetic, naturally occurring, and non-naturally occurring, which have similar binding properties as the reference nucleic acid, and which are metabolized in a manner similar to the reference nucleotides.
  • Examples of such analogs include, without limitation, phosphorothioates, phosphoramidates, methyl phosphonates, chiral-methyl phosphonates, 2-O-methyl ribonucleotides, peptide-nucleic acids (PNAs).
  • polypeptide “peptide” and“protein” are used interchangeably herein to refer to a polymer of amino acid residues.
  • the terms apply to amino acid polymers in which one or more amino acid residue is an artificial chemical mimetic of a corresponding naturally occurring amino acid, as well as to naturally occurring amino acid polymers and non-naturally occurring amino acid polymer.
  • amino acid refers to naturally occurring and synthetic amino acids, as well as amino acid analogs and amino acid mimetics that function in a manner similar to the naturally occurring amino acids.
  • Naturally occurring amino acids are those encoded by the genetic code, as well as those amino acids that are later modified, e.g., hydroxyproline, g-carboxy glutamate, and O-phosphoserine.
  • Amino acid analogs refers to compounds that have the same basic chemical structure as a naturally occurring amino acid, i.e., a g carbon that is bound to a hydrogen, a carboxyl group, an amino group, and an R group, e.g., homoserine, norleucine, methionine sulfoxide, methionine methyl sulfonium. Such analogs have modified R groups (e.g., norleucine) or modified peptide backbones, but retain the same basic chemical structure as a naturally occurring amino acid.
  • Amino acid mimetics refers to chemical compounds that have a structure that is different from the general chemical structure of an amino acid, but that functions in a manner similar to a naturally occurring amino acid.
  • Constantly modified variants apply to both amino acid and nucleic acid sequences. With respect to particular nucleic acid sequences, conservatively modified variants refers to those nucleic acids which encode identical or essentially identical amino acid sequences, or where the nucleic acid does not encode an amino acid sequence, to essentially identical sequences. Because of the degeneracy of the genetic code, a large number of functionally identical nucleic acids encode any given protein. For instance, the codons GCA, GCC, GCG and GCU all encode the amino acid alanine. Thus, at every position where an alanine is specified by a codon, the codon can be altered to any of the corresponding codons described without altering the encoded polypeptide.
  • nucleic acid variations are“silent variations,” which are one species of conservatively modified variations. Every nucleic acid sequence herein which encodes a polypeptide also describes every possible silent variation of the nucleic acid.
  • each codon in a nucleic acid except AUG, which is ordinarily the only codon for methionine, and TGG, which is ordinarily the only codon for tryptophan
  • TGG which is ordinarily the only codon for tryptophan
  • amino acid sequences one of skill will recognize that individual substitutions, deletions or additions to a nucleic acid, peptide, polypeptide, or protein sequence which alters, adds or deletes a single amino acid or a small percentage of amino acids in the encoded sequence is a“conservatively modified variant” where the alteration results in the substitution of an amino acid with a chemically similar amino acid. Conservative substitution tables providing functionally similar amino acids are well known in the art. Such conservatively modified variants are in addition to and do not exclude polymorphic variants, interspecies homologs, and alleles of the invention.
  • the following eight groups each contain amino acids that are conservative substitutions for one another: (1) Alanine (A), Glycine (G); (2) Aspartic acid (D), Glutamic acid (E); (3) Asparagine (N), Glutamine (Q); (4) Arginine I, Lysine (K); (5) Isoleucine (I), Leucine (L), Methionine (M). Valine (V); (6) Phenylalanine (F), Tyrosine (Y), Tryptophan (W); (7) Serine (S), Threonine (T); and (8) Cysteine (C), Methionine (M)
  • nucleic acids e.g., a dsRNA sequence
  • polypeptide sequences refer to two or more sequences or subsequences that are the same or have a specified percentage of amino acid residues or nucleotides that are the same (i.e., 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or more identity over a specified region), when compared and aligned for maximum correspondence over a comparison window, or designated region as measured using one of the following sequence comparison algorithms or by manual alignment and visual inspection.
  • sequences are then said to be“substantially identical.”
  • This definition also refers to the compliment of a test sequence.
  • the identity exists over a region that is at least about 10 to about 100, about 20 to about 75, about 30 to about 50 amino acids or nucleotides in length.
  • a suitable algorithm for determining percent sequence identity and sequence similarity are the BLAST and BLAST 2.0 algorithms, which are described in Altschul et al., Nuc. Acids Res. 25:3389-3402 (1977) and Altschul et al., J. Mol. Biol. 215:403-410 (1990), respectively.
  • Software for performing BLAST analyses is publicly available through the National Center for Biotechnology Information (at the website available at ncbi.nlm.nih.gov).
  • the terms“comprises,”“comprising,”“includes,”“including,”“has,” “having,” “contains”, “containing,” “characterized by” or any other variation thereof, are intended to encompass a non-exclusive inclusion, subject to any limitation explicitly indicated otherwise, of the recited components.
  • a chemical composition and/or method that “comprises” a list of elements is not necessarily limited to only those elements (or components or features or steps), but may include other elements (or components or features or steps) not expressly listed or inherent to the chemical composition and/or method.
  • transitional phrases“consists of’ and“consisting of’ exclude any element, step, or component not specified.
  • “consists of’ or“consisting of’ used in a claim would limit the claim to the components, materials or steps specifically recited in the claim except for impurities ordinarily associated therewith (i.e., impurities within a given component).
  • impurities ordinarily associated therewith i.e., impurities within a given component.
  • the phrase“consists of’ or“consisting of’ limits only the elements (or components or steps) set forth in that clause; other elements (or components) are not excluded from the claim as a whole.
  • transitional phrases“consists essentially of’ and“consisting essentially of’ are used to define a chemical composition and/or method that includes materials, steps, features, components, or elements, in addition to those literally disclosed, provided that these additional materials, steps, features, components, or elements do not materially affect the basic and novel characteristic(s) of the claimed invention.
  • the term“consisting essentially of’ occupies a middle ground between“comprising” and“consisting of’.
  • invention is not intended to refer to any particular embodiment or otherwise limit the scope of the disclosure. Although one or more of these embodiments may be preferred, the embodiments disclosed should not be interpreted, or otherwise used, as limiting the scope of the disclosure, including the claims. In addition, one skilled in the art will understand that the following description has broad application, and the discussion of any embodiment is meant only to be exemplary of that embodiment, and not intended to intimate that the scope of the disclosure, including the claims, is limited to that embodiment.
  • HSV-l herpes simplex virus type 1
  • HSV-2 herpes simplex virus type 2
  • HSV-l is a highly contagious infection, which is common and endemic throughout the world. Most HSV-l infections are acquired during childhood, and infection is lifelong. The vast majority of HSV-l infections are oral herpes (infections in or around the mouth, sometimes called orolabial, oral-labial or oral-facial herpes), but a large proportion of HSV-l infections are genital herpes (infections in the genital or anal area).
  • HSV-2 The decreasing prevalence of HSV-2 in the US (and other European countries) is linked to an increase in genital HSV-l as evidenced by results in the recent disappointing glycoprotein D (gD) subunit vaccine trial in which the majority of cases of genital herpes disease were caused by HSV-l.
  • gD glycoprotein D
  • HSV-l is associated with fewer recurrences and less genital tract viral shedding compared to HSV-2, both serotypes are transmitted perinataly and cause neonatal disease; neonatal disease is associated with high morbidity and mortality even with acyclovir treatment.
  • Subunit formulations consisting of viral envelope glycoproteins combined with adjuvants have predominated the HSV vaccine field for nearly 20 years and the majority of clinical trials have focused on this strategy. Although subunit preparations are safe and elicit neutralizing antibodies, these formulations provided little efficacy against HSV-l or HSV-2 infection or disease in clinical trials. Surprisingly, an HSV-2 gD subunit vaccine provided protection against genital HSV-l, but not HSV-2. Subsequent studies found that serum HSV-2 gD antibody levels correlated with protection against HSV-l, suggesting that the antibody titers required for HSV-2 protection may be higher than those needed to protect against HSV-l.
  • CD4 + and CD8 + T cells were identified surrounding neurons, and while there was heterogenity in the viral proteins targeted, the tegument protein, virion protein 16 (VP 16), was recognized by multiple trigeminal ganglia T cells in the context of diverse HLA-A and -B alleles; these findings suggest that tegument proteins may be important immunogens.
  • cytotoxic T cells directed at tegument proteins were also identified in studies of humans latently infected with HSV-2.
  • CD8 + T cells persist in genital skin and mucosa at the dermal-epidermal junction following HSV reactivation suggesting that they play a role in immune control.
  • HSV-l AgD an engineered an HSV-l virus genetically deleted for native HSV- 1 gD
  • HSV-2 gD gene encodes an envelope glycoprotein essential for viral entry and cell-to-cell spread.
  • Glycoprotein D also binds to tumor necrosis factor receptor superfamily member 14 (TNFRSF14), an immune-regulatory switch also known as herpesvirus entry mediator (HVEM).
  • HVEM herpesvirus entry mediator
  • gD competes with the natural ligands for this receptor and modulates the cytokine response to the virus.
  • the gD gene was replaced with a DNA fragment encoding the green fluorescent protein (gfp) and transformed complementing Vero cells expressing HSV-2 gD VD60 cells (e.g. gD-2 under gD-2 promoter) with this construct were screened for homologous recombinant virus that formed green plaques.
  • the mutant virus replicates in the complementing Vero cell line to high titers (designated HSV-l when passaged on complementing cells), but is noninfectious in non complementing cells (designated HSV-l AgD _/_ when isolated from non-complementing cells).
  • This virus was purified and characterized in vitro. Intravaginal or subcutaneous inoculation of immunocompetent or immunocompromised humanized mice and or guinea pigs to come within the 12 month time frame to file the final patent application.
  • HSV Herpes Simplex Virus
  • a recombinant herpes simplex virus-l (HSV-l) is provided that has a genomic deletion of the endogenous HSV-l glycoprotein D gene. The resulting virion does not encode endogenous glycoprotein D.
  • the HSV-l glycoprotein D comprises the amino acid sequence set forth in SEQ ID NO: l.
  • the recombinant HSV-l further comprises a herpes simplex virus-2 (HSV-2) glycoprotein D on a lipid bilayer thereof.
  • the HSV- 2 glycoprotein D for example, can have the amino acid sequence set forth in SEQ ID NO:2:
  • the HSV-l glycoprotein D-encoding gene is HSV-l Us 6 gene.
  • the HSV-l genomie in which the HSV-l glycoprotein D-encoding gene is deleted can include, but is not limited to an HSV-l genome (prior to the deletion) as set forth in one of the following Genbank listed sequences: HSV-l(G) (KU310668), HSV-2(4674) (KU310667), B3x l.l (KU310657), B3x l.2 (KU310658), B3x l.3 (KU310659), B3x l.4 (KU310660), B3x l.5 (KU310661), B3x2. l (KU310662), B3x2.2 (KU310663), B3x2.3 (KU310664), B3x2.4 (KU310665), B3x2.5 (KU310666).
  • the HSV-l genome can be at least 90, 91, 92, 93, 94, 95, 96, 97, 98, 99 or 100% identical to SEQ ID NO:3, excluding the US6 gene, e.g., nucleotides 138423 to 139607 of SEQ ID NO:3.
  • HSV-l includes various coding regions including (gene, codons, protein name, protein accession number) RL1 513-1259, neurovirulence protein ICP34.5 (AEQ77029.1); RL2 2087-2092 ubiquitin E3 ligase ICP0 (AEQ77030.1); UL1 9338-10012 envelope glycoprotein L (AEQ77031.1); UL2 9885- 10889 uracil-DNA glycosylase (AEQ77032.1); UL3 10991-11665 nuclear protein UL3 (AEQ77033.1); UL5 complement 11754-12423 helicase-primase helicase subunit (AEQ77035.1); UL4 complement 11754-11759 nuclear protein UL4 (AEQ77034.1); UL6 15131-17161 capsid portal protein (AEQ77036.1); UL7 17136-18026 tegument protein UL7 (AEQ77029.1); RL2 2087-2092 ubiquitin
  • a virion of an isolated, recombinant HSV-l can have a genome with a deletion of an HSV-l glycoprotein D-encoding gene.
  • the virion further comprises a heterologous HSV-l or HSV-2 glycoprotein D on the lipid bilayer.
  • the deleted HSV-l glycoprotein D-encoding gene is an HSV-l ETs6 gene.
  • the virion further comprises an HSV-2 glycoprotein D on the lipid bilayer.
  • the virus can further comprises an HSV-l and/or HSV-2 glycoprotein D in the lipid bilayer.
  • the HSV-l glycoprotein D-encoding gene is an HSV-l Eu gene.
  • the virus further comprises an HSV-2 glycoprotein D in the lipid bilayer.
  • a cell can comprise a recombinant HSV-l genome which does not comprise an HSV- 1 Eu gene, i.e., has a genomic deletion of HSV-l glycorpotein D (HSV-lAgD).
  • certain embodiments are directed to a complementing cell which provides expressed HSV 1 or 2 glycoprotein not encoded for by the recombinant HSV-l AgD genome.
  • the complementing cell comprises a heterologous nucleic acid encoding and/or expressing a HSV-l or HSV-2 glycoprotein D.
  • the cell expresses HSV-2 glycoprotein D.
  • the HSV-2 glycoprotein D is encoded and/or expressed by a heterologous nucleic acid.
  • the heterologous nucleic acid comprises a HSV-l or HSV-2 glycoprotein D gene, or is a nucleic acid having a sequence identical or at least 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, or 100% identical to a HSV-l or HSV-2 glycoprotein D gene.
  • a vaccine composition comprising HSV-lAgD, or a HSV-lAgD virion as described herein.
  • the vaccine comprises an immunological adjuvant.
  • the vaccine does not comprise an immunological adjuvant.
  • compositions or pharmaceutical compositions described herein comprising a recombinant HSV-l (HSV-lAgD).
  • the HSV-l (HSV-lAgD) is live or infective.
  • composition comprising the recombinant HSV-l virus as described herein, or the virion as described herein, wherein the genome of the virus or virion comprises at least a deletion of a second gene, wherein the second gene is necessary for HSV-l viral replication or virulence.
  • a pharmaceutical composition comprising the recombinant HSV-l virus as described herein, or the virion as described herein, and a pharmaceutically acceptable carrier.
  • the composition or pharmaceutical composition or vaccine is formulated so that it is suitable for subcutaneous administration to a human subject. In an embodiment, the composition or pharmaceutical composition or vaccine is formulated so that it is suitable for intravaginal administration to a human subject. In an embodiment, the composition or pharmaceutical composition or vaccine is formulated so that it is suitable for intra-muscular, intra-nasal, or mucosal administration to a human subject.
  • Also provided is a method of eliciting an immune response in a subject comprising administering to the subject an amount of the recombinant HSV-l virus as described herein; a virion thereof as described herein, a vaccine as described herein; a composition as described herein; or a pharmaceutical composition as described herein, in an amount effective to elicit an immune response in a subject.
  • Also provided is a method of treating an HSV-l infection in a subject or treating a disease caused by an HSV-l, HSV-2 or co-infection in a subject comprising administering to the subject an amount of the recombinant HSV-l virus as described herein; a virion thereof as described herein, the vaccine as described herein; a composition as described herein; or a pharmaceutical composition as described herein, in an amount effective to treat an HSV-l, HSV- 2 or co-infection or treat a disease caused by an HSV-l, HSV-2 or co-infection in a subject.
  • the methods comprise treating an HSV-l or HSV-2 pathology caused by an HSV-l, HSV-2 or co-infection.
  • the disease caused by an HSV-l, HSV-2 or co-infection is a genital ulcer.
  • the disease caused by an HSV-l, HSV-2 or co-infection is herpes, oral herpes, herpes whitlow, genital herpes, ocular herpes, herpes gladiatorum, HSV keratitis, HSV retinitis, HSV encephalitis or HSV meningitis.
  • treating, or vaccinating for, an HSV-l, HSV-2 or co-infection i.e. infection with both HSV-l and HSV-2
  • co-infection i.e. infection with both HSV-l and HSV-2
  • separate, individual, embodiments of treating an HSV-l infection, treating an HSV-2 infection, treating a co-infection, vaccinating against an HSV-l infection, vaccinating against an HSV-2 infection, and vaccinating against a co-infection are each provided.
  • Also provided is a method of vaccinating a subject for HSV-l, HSV-2 or co-infection comprising administering to the subject an amount of the recombinant HSV-l virus as described herein; a virion thereof as described herein, the vaccine as described herein; a composition as described herein; or a pharmaceutical composition as described herein, in an amount effective to vaccinate a subject for HSV-l, HSV-2 or co-infection.
  • Also provided is a method of immunizing a subject against HSV-l, HSV-2 or co- infection comprising administering to the subject an amount of the recombinant HSV-l virus as described herein; a virion thereof as described herein, the vaccine as described herein; a composition as described herein; or a pharmaceutical composition as described herein, in an amount effective to immunize a subject against HSV-l, HSV-2 or co-infection.
  • the subject is administered a subcutaneous or intravaginal priming dose and is administered a second dose subcutaneously or intravaginally.
  • the subject is administered as many subcutaneous or intravaginal priming doses to elicit anti-HSV antibodies and T cells.
  • a method of producing a virion of a recombinant herpes simplex virus-l comprising infecting a cell comprising a heterologous nucleic acid encoding a HSV-l or HSV-2 glycoprotein D with a recombinant herpes simplex virus-l (HSV-l) having a deletion of an HSV-l glycoprotein D-encoding gene in the genome thereof under conditions permitting replication of the recombinant herpes simplex virus-l (HSV-l) and recovering a HSV-l virion produced by the cell.
  • HSV-l herpes simplex virus-l
  • the cell expresses HSV-l or HSV-2 glycoprotein D on a membrane thereof.
  • a recombinant nucleic acid having the same sequence as a genome of a wild-type HSV-l except that the recombinant nucleic acid does not comprise a sequence encoding an HSV-l glycoprotein D.
  • the recombinant nucleic acid is a DNA.
  • the recombinant nucleic acid is an RNA.
  • HSV-l herpes simplex virus-l
  • AgD HSV-l glycoprotein D-encoding gene
  • the isolated, recombinant HSV-l further comprises a herpes simplex virus-l (HSV-l) or herpes simplex virus-2 (HSV-2) glycoprotein D on a lipid bilayer thereof.
  • the HSV-l glycoprotein D-encoding gene is an HSV-l Us 6 gene.
  • virion of an isolated, recombinant HSV-l having a deletion of an HSV-l glycoprotein D-encoding gene in the genome thereof for treating or preventing an HSV-l, HSV-2 or co-infection in a subject.
  • the virion further comprises an HSV-l or HSV-2 glycoprotein D on a lipid bilayer thereof.
  • the HSV-l glycoprotein D- encoding gene is an HSV-l Us 6 gene.
  • the HSV-l, HSV-2, or co- infection causes a genital ulcer.
  • an isolated, recombinant herpes simplex virus-l (HSV-2) is provided having a deletion of an HSV-l glycoprotein D-encoding gene in the genome thereof.
  • the isolated, recombinant HSV-l further comprises a surface glycoprotein on a lipid bilayer thereof which is a herpes simplex virus-2 (HSV-2) glycoprotein D.
  • the isolated, recombinant HSV-l further comprises a non-HSV-l viral surface glycoprotein on a lipid bilayer thereof.
  • the isolated, recombinant HSV-l further comprises a bacterial surface glycoprotein on a lipid bilayer thereof.
  • the isolated, recombinant HSV-l further comprises a parasitic surface glycoprotein on a lipid bilayer thereof, wherein the parasite is a parasite of a mammal.
  • the HSV-l glycoprotein D-encoding gene is an HSV-l US6 gene.
  • the surface glycoprotein is encoded by a transgene that has been inserted into the genome of the recombinant HSV-l.
  • the surface glycoprotein is present on a lipid bilayer thereof by way of infecting a cell with a recombinant HSV-l having a deletion of an HSV-l glycoprotein D-encoding gene, wherein the cell is or has been transfected to express the surface glycoprotein on a cell membrane thereof, and wherein the recombinant HSV-l comprising the surface glycoprotein present on a lipid bilayer is produced from the cell.
  • a virion of an isolated, recombinant HSV-l having a deletion of an HSV-l glycoprotein D-encoding gene in the genome thereof.
  • the virion of the isolated, recombinant HSV-l further comprises a surface glycoprotein on a lipid bilayer thereof which is a herpes simplex virus-2 (HSV-l) glycoprotein D.
  • the virion of the isolated, recombinant HSV-l further comprises a non-HSV-l viral surface glycoprotein on a lipid bilayer thereof.
  • the virion of the isolated, recombinant HSV-l further comprises a bacterial surface glycoprotein on a lipid bilayer thereof.
  • the virion of the isolated, recombinant HSV-l further comprises a parasitic surface glycoprotein on a lipid bilayer thereof, wherein the parasite is a parasite of a mammal.
  • the HSV-l glycoprotein D-encoding gene is an HSV-l Us 6 gene.
  • the surface glycoprotein is encoded by a transgene that has been inserted into the genome of the recombinant HSV-l of the virion.
  • the surface glycoprotein is present on a lipid bilayer thereof by way of infecting a cell with a recombinant HSV-l having a deletion of an HSV-l glycoprotein D-encoding gene, wherein the cell is or has been transfected to express the surface glycoprotein on a cell membrane thereof, and wherein the recombinant HSV-l comprising the surface glycoprotein present on a lipid bilayer is produced from the cell.
  • the virion has been recovered from such.
  • an isolated cell comprising therein a virus as described herein or a virion as described herein, wherein the cell is not present in a human being.
  • the cell comprises a heterologous nucleic acid encoding a HSV-2 glycoprotein D.
  • the cell expresses HSV-2 glycoprotein D on a membrane thereof.
  • the HSV-2 glycoprotein D is encoded by the heterologous nucleic acid, which heterologous nucleic acid is a HSV-2 glycoprotein D gene, or is a nucleic acid having a sequence identical to a HSV-2 glycoprotein D gene.
  • a vaccine composition comprising a virus as described herein, or a virion as described herein.
  • the vaccine composition comprises an immunological adjuvant.
  • composition comprising a virus as described herein, or a virion as described herein, wherein the genome of the virus or virion comprises at least a deletion of a second gene, wherein the second gene is necessary for HSV-l viral replication.
  • the composition comprises serum from, or is derived from serum from, a mammal into which the virus or virion has been previously introduced so as to elicit an immune response.
  • composition comprising a virus as described herein, or a virion as described herein, and a pharmaceutically acceptable carrier.
  • Also provided is a method of eliciting an immune response in a subject comprising administering to the subject an amount of a virus as described herein; a virion as described herein, a vaccine as described herein; a composition as described herein; or a pharmaceutical composition as described herein, in an amount effective to elicit an immune response in a subject.
  • Also provided is a method of treating an HSV-l infection in a subject or treating a disease caused by an HSV-l infection in a subject comprising administering to the subject an amount of a virus as described herein; a virion as described herein, a vaccine as described herein; a composition as described herein; or a pharmaceutical composition as described herein, in an amount effective to treat an HSV-linfection or treat a disease caused by an HSV-l infection in a subject.
  • Also provided is a method of vaccinating a subject for HSV-l infection comprising administering to the subject an amount of a virus as described herein; a virion as described herein, a vaccine as described herein; a composition as described herein; or a pharmaceutical composition as described herein, in an amount effective to vaccinate a subject for HSV-l.
  • Also provided is a method of immunizing a subject against HSV-l infection comprising administering to the subject an amount of a virus as described herein; a virion as described herein, a vaccine as described herein; a composition as described herein; or a pharmaceutical composition as described herein, in an amount effective to immunize a subject against HSV-l.
  • HSV-l and HSV-2 diseases are known in the art, and are also described herein. Both treatment and prevention of HSV-l and HSV-2 diseases are each separately encompassed. Also treatment or prevention of a HSV-l and HSV-2 co-infection are covered. Prevention is understood to mean amelioration of the extent of development of the relevant disease or infection in a subject treated with the virus, virion, vaccine or compositions described herein, as compared to an untreated subject.
  • a method of producing a virion of a recombinant herpes simplex virus-l comprising infecting a cell comprising a heterologous nucleic acid encoding a HSV-2 glycoprotein D with a recombinant herpes simplex virus-l (HSV-2) having a deletion of an HSV-l glycoprotein D- encoding gene in the genome thereof under conditions permitting replication of the recombinant herpes simplex virus-l (HSV-l) and recovering a recombinant HSV-l virion comprising an HSV-2 glycoprotein D on a lipid bilayer thereof produced by the cell.
  • HSV-l herpes simplex virus-l
  • a method of producing a virion of a recombinant herpes simplex virus-l having a deletion of an HSV-l glycoprotein D-encoding gene in the genome thereof and comprising a non-HSV-l surface glycoprotein on a lipid bilayer thereof, comprising infecting a cell comprising a heterologous nucleic acid encoding the non-HSV-l surface glycoprotein with a recombinant herpes simplex virus-l (HSV-l) having a deletion of an HSV-l glycoprotein D-encoding gene in the genome thereof under conditions permitting replication of the recombinant herpes simplex virus-l (HSV-l) and recovering a recombinant HSV-l virion comprising a non-HSV-l surface glycoprotein on a lipid bilayer thereof produced by the cell.
  • HSV-l herpes simplex virus-l
  • a recombinant nucleic acid having the same sequence as a genome of a HSV-l except that the sequence does not comprise a sequence encoding an HSV-l glycoprotein D.
  • HSV-l herpes simplex virus-l having a deletion of an HSV-l glycoprotein D-encoding gene in the genome thereof for treating or preventing an HSV-l infection in a subject.
  • HSV-l herpes simplex virus-l having a deletion of an HSV-l glycoprotein D-encoding gene in the genome thereof for treating or preventing an HSV-2 infection in a subject.
  • a virion of an isolated, recombinant HSV-l having a deletion of an HSV-l glycoprotein D-encoding gene in the genome thereof for treating or preventing an HSV-l infection in a subject.
  • Also provided is a method of treating an HSV-2 infection, or HSV-l and HSV-2 co- infection, in a subject, or treating a disease caused by an HSV-l infection or HSV-l and HSV-l co-infection in a subject comprising administering to the subject an amount of virus as described herein; a virion as described herein, a vaccine as described herein; a composition as described herein; or a pharmaceutical composition as described herein, in an amount effective to treat an HSV-l infection or treat a disease caused by an HSV-l infection in a subject or an amount effective to treat an HSV-l and HSV-2 co-infection or treat a disease caused by an HSV-l and HSV-2 co-infection in a subject.
  • the subject is a mammalian subject.
  • the mammalian subject is a human subject.
  • HSV-l herpes simplex virus-l
  • the heterogenous antigen is a protein, peptide, polypeptide or glycoprotein.
  • the heterogenous antigen heterogenous antigen with respect to HSV-l but is an antigen found on or in the relevant “pathogen.”
  • Pathogens, viral and bacterial are described herein.
  • the pathogen is a bacterial pathogen of a mammal or a viral pathogen of a mammal.
  • the antigen or the transgene encoding the pathogen is not actually taken or physically removed from the pathogen, but nevertheless has the same sequence as the pathogen antigen or encoding nucleic acid sequence.
  • the isolated, recombinant HSV-l comprises a heterogenous antigen of a pathogen on a lipid bilayer thereof.
  • the pathogen is bacterial or viral.
  • the pathogen is a parasite of a mammal.
  • the HSV-l glycoprotein D-encoding gene is an HSV-l Us 6 gene.
  • the isolated, recombinant HSV-l, the heterogenous antigen is encoded by a transgene that has been inserted into the genome of the recombinant HSV-l.
  • a method of inducing antibody dependent cell mediated cytotoxicity (ADCC) against an antigenic target in a subject comprising administering to the subject an isolated, recombinant herpes simplex virus-l (HSV-l) having a deletion of an HSV-l glycoprotein D-encoding gene in the genome thereof and further comprising a heterogenous antigen on a lipid bilayer thereof in an amount effective to induce antibody dependent cell mediated cytotoxicity (ADCC) against an antigenic target.
  • HSV-l herpes simplex virus-l
  • ADCC antibody dependent cell mediated cytotoxicity
  • Recombinant HSV-l AgD expressing the appropriate transgenes will selectively induce antibodies and cellular immune responses that protect against skin or mucosal infections by pathogens.
  • the heterogenous/heterologous antigen is a surface antigen. In an embodiment, the heterogenous/heterologous antigen is a surface antigen of an antigenic target. In an embodiment, the heterogenous/heterologous antigen is a parasite antigen. In an embodiment, the heterogenous/heterologous antigen is a bacterial antigen or a viral antigen.
  • the subject is a human. In an embodiment of the methods described herein, the subject has not yet been infected with HSV-
  • HSV-2 or co-infection.
  • the subject has been infected with HSV-l, HSV-2 or co-infection.
  • a co-infection means a co-infection with HSV-l and HSV-2.
  • compositions for HSV vaccination of uninfected and infected individuals can be formulated for oral or mucosal delivery as described below.
  • the pharmaceutical composition can also be formulated for injection (e.g., intravenous, intramuscular, intraperitoneal, subcutaneous, etc.).
  • the composition comprises a viral vector encoding an HSV-2 antigen (e.g., glycoprotein D).
  • HSV-2 antigen e.g., glycoprotein D
  • Pharmaceutical compositions comprising the compositions described herein can contain other compounds, which may be biologically active or inactive.
  • Pharmaceutical compositions can be composed to protect against stomach degradation such that the administered composition (e.g., viral vector) reaches the desired location (e.g., ileum).
  • compositions can be delivered using any delivery system known to those of ordinary skill in the art. Numerous gene delivery techniques are well known in the art.
  • the presently described immunogenic compositions can contain pharmaceutically acceptable salts.
  • Such salts may be prepared from pharmaceutically acceptable non-toxic bases, including organic bases (e.g., salts of primary, secondary and tertiary amines and basic amino acids) and inorganic bases (e.g., sodium, potassium, lithium, ammonium, calcium and magnesium salts).
  • organic bases e.g., salts of primary, secondary and tertiary amines and basic amino acids
  • inorganic bases e.g., sodium, potassium, lithium, ammonium, calcium and magnesium salts.
  • Some particular examples of salts include phosphate buffered saline and saline for injection.
  • Suitable carriers include, for example, water, saline, alcohol, a fat, a wax, a buffer, a solid carrier, such as mannitol, lactose, starch, magnesium stearate, sodium saccharine, talcum, cellulose, glucose, sucrose, and magnesium carbonate, or biodegradable microspheres (e.g., polylactate polyglycolate).
  • compositions may also comprise buffers (e.g., neutral buffered saline or phosphate buffered saline), carbohydrates (e.g., glucose, mannose, sucrose or dextrans), mannitol, proteins, polypeptides or amino acids such as glycine, antioxidants, bacteriostats, chelating agents such as EDTA or glutathione, adjuvants (e.g., aluminum hydroxide), solutes that render the formulation isotonic, hypotonic or weakly hypertonic with the blood of a recipient, suspending agents, thickening agents and/or preservatives.
  • buffers e.g., neutral buffered saline or phosphate buffered saline
  • carbohydrates e.g., glucose, mannose, sucrose or dextrans
  • mannitol proteins
  • proteins polypeptides or amino acids
  • proteins e.glycine
  • antioxidants e.g., g., bacteriostats,
  • compositions further comprise an adjuvant such as a TLR-3 agonist (e.g., dsRNA or a mimetic thereof such as poly I:C or poly A:U).
  • TLR-3 agonist is used to stimulate immune recognition of an antigen of interest.
  • TLR-3 agonists include, for example, short hairpin RNA, virally derived RNA, short segments of RNA that can form double-strands or short hairpin RNA, and short interfering RNA (siRNA).
  • the TLR-3 agonist can be virally derived dsRNA, such as for example, a dsRNA derived from a Sindbis virus or dsRNA viral intermediates (Alexopoulou et al.
  • the TLR-3 agonist is a short hairpin RNA.
  • Short hairpin RNA sequences typically comprise two complementary sequences joined by a linker sequence.
  • the particular linker sequence is not critical. Any linker sequence can be used so long as it does not interfere with the binding of the two complementary sequences to form a dsRNA.
  • Suitable adjuvants include, for example, the lipids and non-lipid compounds, cholera toxin (CT), CT subunit B, CT derivative CTK63, E. coli heat labile enterotoxin (LT), LT derivative LTK63, Al(OH)3, and polyionic organic acids as described in e.g., WO 04/020592, Anderson and Crowle, Infect. Immun. 31(1):413-418 (1981), Roterman et al., J. Physiol. Pharmacol., 44(3):2l3-32 (1993), Arora and Crowle. J. Reticuloendothel. 24(3):27l-86 (1978), and Crowle and May, Infect. Immun.
  • Suitable polyionic organic acids include for example, 6,6'-[3,3'-demithyl[l,l'-biphenyl]-4,4'-diyl]bis(azo)bis[4-amino-5-hydroxy- 1, 3 -naphthalene-di sulfonic acid] (Evans Blue) and 3,3'-[l,l'biphenyl]-4,4'-diylbis(azo)bis[4- amino-l-naphthalenesulfonic acid] (Congo Red).
  • Suitable adjuvants include topical immunomodulators such as, members of the imidazoquinoline family such as, for example, imiquimod and resiquimod (see, e.g., Hengge et al., Lancet Infect. Dis. l(3):l89-98 (2001).
  • Additional suitable adjuvants are commercially available as, for example, additional alum-based adjuvants (e.g., Alhydrogel, Rehydragel, aluminum phosphate, Algammulin); oil based adjuvants (Freund's Incomplete Adjuvant and Complete Adjuvant (Difco Laboratories, Detroit.
  • additional alum-based adjuvants e.g., Alhydrogel, Rehydragel, aluminum phosphate, Algammulin
  • oil based adjuvants Freund's Incomplete Adjuvant and Complete Adjuvant (Difco Laboratories, Detroit.
  • cytokines e.g., GM-CSF or Flat3 -ligand
  • Merck Adjuvant 65 Merck and Company, Inc., Rahway, N.J.
  • AS-2 SmithKline Beecham, Philadelphia, Pa.
  • salts of calcium, iron or zinc an insoluble suspension of acylated tyrosine; acylated sugars; cationically or anionically derivatized polysaccharides; polyphosphazenes; biodegradable microspheres; monophosphoryl lipid A and Quil A.
  • Cytokines such as GM-CSF or interleukin-2, -7, or -12, are also suitable adjuvants.
  • Hemocyanins e.g., keyhole limpet hemocyanin
  • Polysaccharide adjuvants such as, for example, chitin, chitosan, and deacetylated chitin are also suitable as adjuvants.
  • Other suitable adjuvants include muramyl dipeptide (MDP, N acetylmuramyl L alanyl D isoglutamine) bacterial peptidoglycans and their derivatives (e.g., threonyl-MDP, and MTPPE).
  • BCG and BCG cell wall skeleton may also be used as adjuvants in the invention, with or without trehalose dimycolate.
  • Trehalose dimycolate may be used itself (see, e.g., U.S. Pat. No. 4,579,945).
  • Detoxified endotoxins are also useful as adjuvants alone or in combination with other adjuvants (see, e.g., U.S. Pat. Nos. 4,866,034; 4,435,386; 4,505,899; 4,436,727; 4,436,728; 4,505,900; and 4,520,019.
  • the saponins QS21, QS17, QS7 are also useful as adjuvants (see, e.g., U.S. Pat. No. 5,057,540; EP 0362 279; WO 96/33739; and WO 96/11711).
  • Other suitable adjuvants include Montanide ISA 720 (Seppic, France), SAF (Chiron, Calif., United States), ISCOMS (CSL), MF-59 (Chiron), the SBAS series of adjuvants (e.g., SBAS-2, SBAS-4 or SBAS-6 or variants thereof, available from SmithKline Beecham, Rixensart, Belgium), Detox (Corixa, Hamilton. Mont.), and RC-529 (Corixa, Hamilton, Mont.).
  • compositions can be packaged in unit-dose or multi-dose containers, such as sealed ampoules or vials. Such containers can be hermetically sealed to preserve integrity or sterility of the formulation until use.
  • Formulations can be stored as suspensions, solutions, or emulsions in oily or aqueous vehicles, in a lyophilized condition (e.g., for addition of sterile liquid carrier before use), or in an oral delivery formulation, e.g. capsule, tablet, or pill.
  • compositions can be by any non-parenteral route (e.g., vaginally, orally, intranasally, or otherwise mucosally via, e.g., lungs, salivary glands, nasal cavities, small intestine, colon, rectum, tonsils, or Peyer's patches), or any parenteral route (e.g., intramuscular, subcutaneous, intraperitoneal, intravenous, etc.).
  • the composition can be administered alone or with an adjuvant.
  • the adjuvant is administered at the same time as the composition.
  • the adjuvant is administered after the composition, e.g., 1, 2, 6, 12, 18, 24, 36, 48, 60, or 72 hours after administration of the composition.
  • the presently disclosed compositions can be administered in combination with other immunogenic compositions.
  • compositions can be administered prophylactically, to an individual that does not have detectable HSV, has not displayed symptoms of HSV infection, or one that is at risk of infection.
  • the presently disclosed compositions can also be administered to reduce severity of HSV symptoms in an individual that is already infected, and reduce the likelihood of the individual spreading the virus (e.g., by reducing viral shedding).
  • Frequency of administration of the prophylactic or therapeutic compositions described herein, as well as dosage, will vary from individual to individual, and can be readily established using standard techniques. Between 1 and 10 doses may be administered over a 52 week period. In some embodiments, the presently disclosed composition is administered upon early indication of an outbreak. In some embodiments, administration is once/year. In some embodiments, 3 doses are administered, at intervals of 1 month, or 2-3 doses are administered every 2-3 months. Booster vaccinations can be given periodically thereafter. Alternate protocols may be appropriate for individual patients and particular diseases and disorders.
  • a suitable dose is an amount of the composition that, when administered as described above, is capable of promoting an anti-viral immune response, and is at least 10-50% above the basal (i.e., untreated) level.
  • Such response can be monitored by measuring vaccine-dependent generation or activation of cytolytic CD8 T cells capable of killing virally infected cells, e.g., as determined in vitro.
  • Such vaccines should also be capable of causing an immune response that leads to an improved clinical outcome (e.g., less frequent outbreaks, or complete or partial remission) in vaccinated as compared to non- vaccinated individuals.
  • the dose size may be adjusted based on the particular patient.
  • the presently disclosed compositions can conveniently be formulated in a coated tablet, pill, or capsule.
  • gel, ointment, or suppository can be used.
  • An appropriate dosage and treatment regimen provides the presently disclosed compositions in an amount sufficient to provide therapeutic and/or prophylactic benefit. Such a response can be monitored by establishing an improved clinical outcome (e.g., less frequent outbreaks, prevention of appearance of symptoms, complete or partial, reduced rate of spreading the infection) in treated individuals as compared to non-treated individuals.
  • Immune responses to the presently disclosed compositions can be evaluated using standard proliferation, cytotoxicity or cytokine assays, which may be performed using samples obtained from a patient before and after treatment.
  • An immune response to a given antigen can be detected using any means know in the art including, for example, detecting specific activation of CD4+ or CD8+ T cells or by detecting the presence of antibodies that specifically bind to the polypeptide.
  • Specific activation of CD4+ or CD8+ T cells associated with a mucosal, humoral, or cell-mediated immune response can be detected in a variety of ways.
  • Methods for detecting specific T cell activation include, but are not limited to, detecting the proliferation of T cells, the production of cytokines (e.g., lymphokines), or the generation of cytolytic activity (i.e., generation of cytotoxic T cells specific for the immunogenic polypeptide).
  • cytokines e.g., lymphokines
  • cytolytic activity i.e., generation of cytotoxic T cells specific for the immunogenic polypeptide.
  • specific T cell activation is indicated by proliferation of T cells.
  • cytolytic activity e.g., detectable using 5lCr release assays (see, e.g., Brossart and Bevan, Blood 90(4): 1594-1599 (1997) and Lenz et ak, J. Exp. Med. 192(8): 1135-1142 (2000)).
  • T cell proliferation can be detected by measuring the rate of DNA synthesis in T cells (e.g., isolated CD8 T cells).
  • a typical way to measure the rate of DNA synthesis is, for example, by pulse-labeling cultures of T cells with tritiated thymidine, a nucleoside precursor which is incorporated into newly synthesized DNA. The amount of tritiated thymidine incorporated can be determined using a liquid scintillation spectrophotometer.
  • T cell proliferation examples include measuring increases in interleukin-2 (IL-2) production, Ca2+ flux, or dye uptake, such as 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium.
  • lymphokines e.g., interferon-gamma
  • ICP0 or other HSV-2 antigen the immunogenic polypeptide
  • Antibody immune responses can be detected using immunoassays known in the art (see, e.g., Tucker el al., Mol Therapy, 8, 392-399 (2003); Tucker el ak, Vaccine, 22, 2500-2504 (2004)).
  • Suitable immunoassays include the double monoclonal antibody sandwich immunoassay technique of David et al. (U.S. Pat. No. 4,376,110); monoclonal-polyclonal antibody sandwich assays (Wide et al., in Kirkham and Hunter, eds., Radioimmunoassay Methods, E. and S.
  • HSV-2 antigens such as ICP0 or an HSV-2 capsid protein (e.g, gD or gB).

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Abstract

Certains modes de réalisation concernent un virus herpès simplex 1 (VHS-1) de recombinaison ayant une délétion génomique d'une glycoprotéine du VHS-1. Dans certains aspects, la délétion génomique est un gène codant pour la glycoprotéine D du VHS-1 (Us6). Dans certains aspects, le virion VHS-1 comprend une glycoprotéine hétérologue, par exemple une glycoprotéine du VHS-1 et/ou VHS-2. Dans un aspect particulier, la glycoprotéine hétérologue est la glycoprotéine D de VHS-2.
PCT/US2019/051303 2018-09-14 2019-09-16 Virus herpès simplex 1 (vhs-1) de recombinaison Ceased WO2020056411A1 (fr)

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Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2005005637A2 (fr) * 2003-05-09 2005-01-20 Medigene, Inc. Ciblage de virus sur des recepteurs de cellules de surface
US20110177125A1 (en) * 2006-09-08 2011-07-21 Harvey Friedman Hsv-1 and hsv-2 vaccines and methods of use thereof
US20170202952A1 (en) * 2014-03-03 2017-07-20 Albert Einstein College Of Medicine, Inc Recombinant herpes simplex virus 2 (hsv-2) vaccine vectors

Patent Citations (3)

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Publication number Priority date Publication date Assignee Title
WO2005005637A2 (fr) * 2003-05-09 2005-01-20 Medigene, Inc. Ciblage de virus sur des recepteurs de cellules de surface
US20110177125A1 (en) * 2006-09-08 2011-07-21 Harvey Friedman Hsv-1 and hsv-2 vaccines and methods of use thereof
US20170202952A1 (en) * 2014-03-03 2017-07-20 Albert Einstein College Of Medicine, Inc Recombinant herpes simplex virus 2 (hsv-2) vaccine vectors

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Title
LIGAS, M ET AL.: "A Herpes Simplex Virus Mutant In Which Glycoprotein D Sequences Are Replaced By beta-Galactosidase Sequences Binds To But Is Unable To Penetrate Into Cells", JOURNAL OF VIROLOGY, vol. 62, no. 5, May 1998 (1998-05-01), pages 1486 - 1494, XP002443290 *
WARNER, M ET AL.: "Cell Surface Protein With Herpesvirus Entry Activity (HveB) Confers Susceptibility To Infection By Mutants Of Herpes Simplex Virus Type 1, Herpes Simplex Virus Type 2, And Pseudorabies Virus", VIROLOGY, vol. 246, no. 1, 1998, pages 179 - 189, XP004445786, DOI: 10.1006/viro.1998.9218 *
ZHOU, G ET AL.: "Glycoprotein D or J Delivered In Trans Blocks Apoptosis In SK -N-SH Cells Induced by A Herpes Simplex Virus 1 Mutant Lacking Intact Genes Expressing Both Glycoproteins", JOURNAL OF VIROLOGY, vol. 74, no. 24, December 2000 (2000-12-01), pages 11782 - 11791 *

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