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WO2024226815A2 - Épitopes de lymphocytes b d'antigènes treponema pallidum destinés à être utilisés dans un vaccin contre la syphilis - Google Patents

Épitopes de lymphocytes b d'antigènes treponema pallidum destinés à être utilisés dans un vaccin contre la syphilis Download PDF

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WO2024226815A2
WO2024226815A2 PCT/US2024/026293 US2024026293W WO2024226815A2 WO 2024226815 A2 WO2024226815 A2 WO 2024226815A2 US 2024026293 W US2024026293 W US 2024026293W WO 2024226815 A2 WO2024226815 A2 WO 2024226815A2
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seq
sequence
epitope
set forth
virus
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WO2024226815A3 (fr
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Lorenzo GIACANI
Sheila A. Lukehart
Barbara MOLINI
Anna Wald
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University of Washington
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University of Washington
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    • 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
    • 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
    • A61K39/02—Bacterial antigens
    • A61K39/0225—Spirochetes, e.g. Treponema, Leptospira, Borrelia
    • 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
    • A61K2039/525—Virus
    • A61K2039/5256—Virus expressing foreign proteins
    • 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
    • A61K2039/525—Virus
    • A61K2039/5258—Virus-like particles
    • 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
    • A61K2039/545—Medicinal preparations containing antigens or antibodies characterised by the dose, timing or administration schedule
    • 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
    • A61K2039/55505—Inorganic adjuvants
    • 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
    • A61K2039/55577—Saponins; Quil A; QS21; ISCOMS
    • 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
    • A61K2039/572—Medicinal preparations containing antigens or antibodies characterised by the type of response, e.g. Th1, Th2 cytotoxic 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
    • A61K2039/57—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

Definitions

  • the present disclosure also describes epitopes in a concatenated form or expressed on a delivery scaffold such as a scaffold protein, a virus-like particle, or a nanoparticle for use in vaccines.
  • the vaccines can be used to selectively elicit antibodies capable of neutralizing T. pallidum.
  • BACKGROUND OF THE DISCLOSURE [0005] After years of steady decline during the 1990's, syphilis has become resurgent in many developed countries including the United States (US), Canada, and China, as well as in many European nations. Furthermore, it continues to be endemic in developing countries, where more than 90% of cases occur.
  • WHO World Health Organization
  • CS Congenital syphilis
  • Treponema pallidum subsp. pallidum T. pallidum
  • T. pallidum epitopes include sequences as set forth in SEQ ID NOs: 1-12.
  • an epitope described herein can be expressed on a delivery scaffold such as a scaffold protein, a virus-like particle, or a nanoparticle.
  • an epitope described herein can be expressed as a concatemer including a chimeric concatemer.
  • Various implementations of the present disclosure relate to syphilis epitopes and vaccines that can be used to elicit or increase an immune response to syphilis.
  • the syphilis epitopes described herein can be used in a vaccine to prevent or decrease the severity of a syphilis infection.
  • the syphilis vaccine includes an epitope described in SEQ ID NOs: 1-12.
  • the syphilis vaccine includes an epitope described in SEQ ID NOs: 1-12 expressed on a delivery scaffold.
  • the syphilis vaccine includes an epitope described in SEQ ID NOs: 1-12 expressed as a concatemer.
  • a vaccine as described herein includes one or more of the syphilis epitopes described herein and an adjuvant.
  • FIGs.1A, 1B (1A) 3D model of Nichols strain TprC (TP0117) generated using I-TASSER. Loops are predicted to be surface exposed. Variability among strains is found in loops without white arrow (loops 5, and 9-11), while loops indicated with white arrow are conserved among strains.
  • FIG.2 Multiple sequence alignment of the COOH-terminal region (amino acids (aa) 405-600) of five TprC variants found in T. pallidum subsp. pallidum isolates, where the most antigenic variability is located. External loop (ExL) 5 (not shown) is also diverse.
  • FIG.3 Schematic of T. pallidum tprK gene (TP0897) and mechanism of antigenic variation of the variable (V) regions.
  • the tprK gene (top) represents the expression site for the TprK protein.
  • Seven discrete V regions (termed V1-V7) are located within the tprK open reading frame (ORF) (Centurion-Lara et al., Molecular Microbiology. 2004;52(6):1579-96 (“Centurion-Lara 2”)). Dark grey and white (C) regions are conserved among all tprK genes sequenced to date.
  • ORF tprK open reading frame
  • FIGs. 4A, 4B (4A) Transcription of IFNy by PBMC from control and immunized rabbits incubated with mitogen (ConA) or T. pallidum vaccine candidate.
  • FIG.5 Phagocytosis of T. pallidum in the presence of antisera against recombinant peptides, compared to normal (NRS) and infected (IRS, positive control) rabbit serum. TprK antiserum is directed against the protein conserved NH 2 -terminus.
  • TprC/D antiserum is to the Tpr Subfamily I group of gene products.
  • FlaA is a periplasmic protein (also negative control). *p ⁇ 0.05 vs. NRS.
  • FIGs.6A, 6B Primary chancre development in rabbits immunized with TprK conserved NH2-termial region (AA 37-273 of, for example, SEQ ID NO: 40) (6A) and unimmunized rabbits (6B). Pictures were taken 23 days post- challenge. Morgan et al., Infect Immun.2002;70(12):6811-6. This work was performed with the previously commercially available Ribi adjuvant.
  • FIG. 7. Phagocytosis of T.
  • FIG.8 Schematic representation of the experiments. Sera collected from immunized animals will be used to evaluate B-cell epitopes (procedure #1, ELISA) and ability to opsonize (procedure #2, opsonophagocytosis assay) or neutralize (procedure #3, neutralization assay) T. pallidum.
  • PBMCs were used to evaluate INF-y production upon stimulation (procedure #4, ELISA after stimulation) to evaluate induction of a Th1-type response.
  • ELISA, opsonophagocytosis and neutralization assays were repeated with monoclonal antibodies (procedure #5).
  • TprC sequences of Chicago and Bal73- 1 strains are identical to Nichols. Together, the MexicoA, Sea81-4, and Nichols sequences encompass the known spectrum antigenic variability in these proteins.
  • Peptides 1, 14, 18, 26-30, 46, 47, 51, 54-56 are all predicted to be found in TprC external loops.
  • Treponemal cell burden was estimated by real-time quantitative PCR in lesion aspirates from rabbits immunized with TprK and TprC, compared to control animals that were not immunized but challenged with either the Nichols or SS14 strain. Rabbits immunized with TprK and TprC have significantly fewer treponemes in their lesions compared to controls infected with the Nichols strain, supporting vaccine efficacy.
  • FIG.13 Visual assessment of delayed-type hypersensitivity (DTH) at 48 hours after challenge of immunized rabbits. Control rabbits do not exhibit DTH at injection sites, but rabbits immunized with TprK and TprC do. This result supports that immunization induced a T-cell response in animals.
  • FIGs.14A, 14B Visual assessment of delayed-type hypersensitivity (DTH) at 48 hours after challenge of immunized rabbits. Control rabbits do not exhibit DTH at injection sites, but rabbits immunized with TprK and TprC do. This result supports that immunization induced a T-cell response in animals.
  • FIG. 15 Example of epitope mapping using sera from TprC-immunized animals.
  • FIGs. 16A, 16B Example of epitope mapping using sera from TprK-immunized animals.
  • sera from rabbits immunized with the TprK antigen were evaluated using an enzyme-linked immunosorbent assay (ELISA) to define which epitopes were predominantly recognized on regions of the protein mapping at the host-pathogen interface.
  • Sera were tested against a pool of peptides corresponding to conserved external loops (CExL) of the vaccine candidate.
  • CExL1 (16A) and CEXL2 (16B) are more strongly recognized.
  • FIG.17 Alignment of amino acid sequences of the predicted protein sequences encoded at the tprD, tprC, tprF, and tprI loci to show where the peptides identified here map on the protein sequence and the identity level across Tpr antigens.
  • the TprF truncated proteins in syphilis strains are also included in the alignment for clarity purposes.
  • DVR Discrete variable regions.
  • FIG.18 Sequences for T. pallidum strains aligned in FIG.17.
  • FIG.19 TprK and TprC sequences. DETAILED DESCRIPTION
  • the current disclosure provides T. pallidum vaccine epitopes specifically designed to be recognized by B cells to fight infection.
  • An epitope includes specific amino acids that contact the binding portions of an immunoglobulin or antibody.
  • Epitope determinants can include chemically active surface groupings of molecules such as amino acids, sugar side chains, phosphoryl or sulfonyl groups, or any combination thereof, and can have specific three-dimensional structural characteristics, and/or specific charge characteristics.
  • An “epitope” includes any determinant capable of being bound by an antigen-binding protein, such as a B cell receptor (BCR).
  • An epitope is a region of molecule that is bound by a binding protein that targets that region of a molecule, and when that region of a molecule is a protein, includes specific residues that directly contact the binding protein.
  • an “epitope” denotes the binding site on a protein target bound by a corresponding binding domain.
  • the binding domain binds to a linear epitope, (e.g., an epitope including a stretch of 5 to 12 consecutive amino acids), to a three-dimensional structure formed by the spatial arrangement of several short stretches of the protein target, or a combination thereof.
  • a linear epitope e.g., an epitope including a stretch of 5 to 12 consecutive amino acids
  • Three-dimensional epitopes recognized by a binding domain e.g., by the epitope recognition site of a BCR or BCR fragment, can be thought of as three-dimensional surface features of an epitope molecule. These features fit precisely with the corresponding binding site of the binding domain. Accordingly, binding between the binding domain and its target protein is facilitated.
  • an epitope can be considered to have two levels: (i) the “covered patch” which can be thought of as the shadow a BCR or a binding domain would cast; and (ii) the individual participating side chains and backbone residues. Binding is then due to the aggregate of ionic interactions, hydrogen bonds, and hydrophobic interactions.
  • B cells mediate the humoral immune system through antibody secretion that neutralizes antigens. B cells are stimulated when the antigen receptor, which is part of the paratope, recognizes antigenic epitopes. Epitopes recognized by B cells can be classified into two types: continuous and discontinuous epitopes.
  • Continuous epitopes are short peptide fragments (15 amino acids in size) of an antigen protein that are specifically identified by certain antibodies. Discontinuous epitopes include amino acid residues that are not sequential in their primary structure but involve a folding mechanism that forms into a region that is close together.
  • the T. pallidum strain includes a Nichols strain, a Bal3 strain, a Sea81-4 strain, UW249 strain, a MexicoA strain, an Iraq strain, a SamoaD strain, a Gauthier strain, and a TP0316 strain.
  • the T. pallidum strain includes a Nichols strain, a Bal3 strain, a Sea81-4 strain, UW249 strain, a MexicoA strain, an Iraq strain, a SamoaD strain, a Gauthier strain, and a TP0316 strain.
  • the T. pallidum strain includes a Nichols strain, a Bal3 strain, a Sea81-4 strain, UW249 strain, a MexicoA strain, an Iraq strain,
  • a T. pallidum epitope includes a sequence set forth in Table 1. Table 1. TprK- and TprC-specific Epitopes. Variants of the same epitope are listed with the same External Loop (ExL) number. Epitope Specificity and ExL Sequence SEQ ID NO [0035] As indicated, in particular implementations, the syphilis vaccine includes a subunit vaccine.
  • a subunit vaccine can refer to a vaccine that includes only a subunit (e.g., a single protein or protein fragment) of the pathogen that stimulates an immune response against the pathogen. For instance, a subunit vaccine omits a whole live or killed pathogen.
  • a subunit vaccine omits a whole live or killed pathogen.
  • the term “concatenate” is broadly used to describe linking together into a chain or series. It is used to describe the linking together of nucleotide or amino acid sequences into a single nucleotide or amino acid sequence, respectively.
  • the term “concatemerize” should be interpreted to recite: “concatenate.”
  • a “concatemer” is a molecule containing multiple copies of a sequence linked in series.
  • a concatemer includes 2, 3, 4, 5, 6, 7, 8, 9, or more copies of any of SEQ ID NOs: 1-12.
  • a concatemer includes 3 copies of SEQ ID NO: 1.
  • a concatemer includes 2 copies of SEQ ID NO: 2.
  • a concatemer includes 4 copies of SEQ ID NO: 3.
  • a concatemer includes 3 copies of SEQ ID NO: 4.
  • a concatemer includes 5 copies of SEQ ID NO: 5.
  • a concatemer includes 6 copies of SEQ ID NO: 6.
  • a concatemer includes 7 copies of SEQ ID NO: 7.
  • a concatemer includes 9 copies of SEQ ID NO: 8. In particular implementations, a concatemer includes 8 copies of SEQ ID NO: 9. In particular implementations, a concatemer includes 2 copies of SEQ ID NO: 10. In particular implementations, a concatemer includes 3 copies of SEQ ID NO: 11. In particular implementations, a concatemer includes 3 copies of SEQ ID NO: 12. [0038] In particular implementations, a concatemer includes 2, 3, 4, 5, 6, 7, 8, 9, or more copies of any of cExL1, cExL2, or cExL3.
  • a concatemer includes cExL1-cExL1-cExL1, cExL1-cExL1-cExL1, cExL1-cExL1-cExL1-cExL1-cExL1, cExL1-cExL1-cExL1-cExL1- cExL1- cExL1- cExL1- cExL1, cExL2-cExL2, cExL2-cExL2-cExL2, cExL2-cExL2-cExL2, cExL2-cExL2-cExL2-cExL2, cExL3-cExL3, cExL3-cExL3-cExL3-cExL3, or cExL3-cExL3-cExL3-cExL3-cExL3-cExL3-cExL3.
  • a concatemer includes 2, 3, 4, 5, 6, 7, 8, 9, or more copies of any of ExL1, ExL3, ExL4, ExL6, ExL9, or ExL10.
  • a concatemer includes ExL1- ExL1, ExL1- ExL1- ExL1- ExL1, ExL1- ExL1- ExL1- ExL1- ExL1- ExL1, ExL1- ExL1- ExL1- ExL1- ExL1- ExL1- ExL1- ExL1- ExL1- ExL1- ExL1, ExL3- ExL3, ExL3- ExL3- ExL3, ExL3- ExL3- ExL3- ExL3, ExL3- ExL3- ExL3- ExL3- ExL3, ExL3- ExL3- ExL3- ExL3- ExL3, ExL3- ExL3- ExL3- ExL3- ExL3, ExL3- ExL3- ExL3- ExL3- ExL3, ExL3- ExL3- Ex
  • a concatemer includes a chimeric concatemer.
  • a chimeric concatemer includes at least 2 of the sequences selected from SEQ ID NOs: 1-12.
  • a chimeric concatemer includes 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 copies of a first sequence selected from any one of SEQ ID NOs: 1-12 and 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 copies of a second sequence selected from any one of SEQ ID NOs: 1-12, wherein the first and second sequence are not the same.
  • the chimeric concatemer can include 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 copies of a third, fourth, fifth, or more sequences.
  • a chimeric concatemer includes SEQ ID NO: 1, SEQ ID NO: 2, and SEQ ID NO: 3 operatively linked together.
  • a chimeric concatemer includes SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 11, and SEQ ID NO: 12 operatively linked together.
  • a chimeric concatemer includes SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 11, and/or SEQ ID NO: 12 operatively linked together.
  • a chimeric concatemer includes SEQ ID NO: 5, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 11, and SEQ ID NO: 12 operatively linked together.
  • a chimeric concatemer includes SEQ ID NO: 5, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 11, and SEQ ID NO: 12 operatively linked together.
  • a chimeric concatemer includes SEQ ID NO: 5, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 11, and SEQ ID NO: 12 operatively linked together.
  • a chimeric concatemer includes cExL1-cExL2-cExL1, cExL1-cExL2- cExL3, cExL3-cExL2-cExL3, cExL2-cExL1-cExL2, cExL1-cExL3-cExL3, cExL1-cExL2-cExL2-cExL1, cExL1-cExL1-cExL2- cExL3, cExL1-cExL3-cExL1-cExL3, cExL1-cExL1-cExL1-cExL2, cExL2-cExL3-cExL3-cExL3, cExL2-cExL2-cExL1-cExL1-cExL1-cExL2,
  • a chimeric concatemer includes ExL1-ExL9-ExL10, ExL3-ExL6-ExL3, ExL4- ExL1-ExL1, ExL6-ExL1-ExL6, ExL9-ExL9-ExL9, ExL10-ExL3-ExL4, ExL1-ExL1-ExL4-ExL4, ExL3-ExL4-ExL3-ExL4, ExL4-ExL1-ExL9-ExL1, ExL6-ExL3-ExL10-ExL1, ExL9-ExL4-ExL3-ExL1, ExL10-ExL6-ExL6-ExL10, ExL1-ExL3- ExL4-ExL6-ExL9- ExL10, or ExL9-ExL4-ExL3-ExL1-ExL6-ExL10.
  • a chimeric concatemer includes cExL1-ExL1-cExL1, cExL2-ExL3-cExL2, cExL3-ExL4-ExL6, ExL1-ExL6-cExL2, ExL3-ExL9-ExL1, ExL4-ExL10-ExL4, ExL6-cExL1-cExL3, ExL9-cExL2-cExL1, ExL10-cExL3-ExL10, cExL1-cExL1-ExL1-ExL1,cExL2-ExL3-ExL4-cExL3, cExL3-ExL6-ExL10-ExL9, ExL1-cExL1- ExL6-cExL3, ExL3-cExL1-ExL9-cExL1, ExL4-ExL4-ExL10-ExL4, ExL6-cExL3-cExL3-cExL1, ExL9-ExL
  • syphilis epitopes and vaccines that can be used to elicit or increase an immune response to syphilis.
  • the syphilis epitopes described herein can be used in a vaccine to prevent or decrease the severity of a syphilis infection.
  • the syphilis vaccine includes an epitope described in SEQ ID NOs: 1-12.
  • the syphilis vaccine includes an epitope described in SEQ ID NOs: 1-12 expressed on a delivery scaffold.
  • the syphilis vaccine includes an epitope described in SEQ ID NOs: 1-12 expressed as a concatemer.
  • the syphilis epitope is presented on a delivery scaffold.
  • the delivery scaffold includes a scaffold protein, a virus-like particle, or a nanoparticle.
  • a vaccine as described herein includes one or more of the syphilis epitopes described herein and an adjuvant. Adjuvants are described in more detail elsewhere herein.
  • a delivery scaffold can confer desired properties to the epitope, for example, a delivery scaffold may improve the immunogenicity of an antigen, e.g., by altering the structure of the antigen, by altering uptake and processing of the antigen, and/or by allowing the antigen to bind to a binding partner.
  • a delivery scaffold includes an epitope- scaffold protein, a virus-like particle, or a nanoparticle.
  • An epitope expressed on a delivery scaffold forms an epitope- scaffold molecule.
  • an epitope-scaffold molecule includes an epitope expressed on an epitope-scaffold protein, an epitope expressed on a virus-like particle, or an epitope expressed on a nanoparticle.
  • An epitope-scaffold protein is a chimeric protein that includes an epitope sequence fused to a heterologous "acceptor" scaffold protein. Design of the epitope-scaffold is performed, for example, computationally in a manner that preserves the native structure and conformation of the epitope when it is fused onto the heterologous scaffold protein.
  • the use of such scaffold proteins is well known in the art and such methods and techniques are described in WO 2011/050168 and US 2010/0068217 and the skilled person can follow methods described therein and apply them to the present invention.
  • Scaffold proteins are useful for creating immunogens of the present invention in that they hold contact residues in the immunogen in the proper spatial orientation to facilitate interaction between such residues and contact residues of the antigen.
  • “Superposition” epitope-scaffolds are based on scaffold proteins having an exposed segment with similar conformation as the target epitope—the backbone atoms in this “superposition-region” can be structurally superposed onto the target epitope with minimal root mean square deviation (RMSD) of their coordinates.
  • RMSD root mean square deviation
  • Suitable scaffolds are identified by computationally searching through a library of protein crystal structures; epitope-scaffolds are designed by putting the epitope residues in the superposition region and making additional mutations on the surrounding surface of the scaffold to prevent clash or other interactions with the antibody.
  • “Grafting” epitope-scaffolds utilize scaffold proteins that can accommodate replacement of an exposed segment with the crystallized conformation of the target epitope. For each suitable scaffold identified by computationally searching through all protein crystal structures, an exposed segment is replaced by the target epitope and the surrounding sidechains are redesigned (mutated) to accommodate and stabilize the inserted epitope.
  • the epitope-scaffold protein can include an appropriate protein from nature or synthetic.
  • the epitope-scaffold protein can be derived from bacterial proteins, viral proteins, or eukaryotic proteins.
  • the epitope-scaffold protein includes a Treponema pallidum protein (Tp0751), small-synthetic ⁇ -barrel protein, Protein Z from S.
  • Treponema pallidum protein (Tp0751), also referred to as pallilysin, is a protein from Treponema pallidum that displays a propensity to interact with the extracellular matrix. Structural analysis revealed an eight-stranded beta-barrel with a profile of short, conserved regions consistent with a non-canonical lipocalin fold.
  • Tp0751 includes the sequence: GSMASHGNAPPAPVGGAAQTHTQPPVQTAMRIALWNRATHGEQGALQHLLAGLWIQTEISPNSGDIHPLLFFDREHA EITFSRASVQEIFLVDSAHTHRKTVSFLTRNTAISSIRRRLEVTFESHAVIHVRAVEDVARLKIGSTSMWDGQYTRYHA GPASAPSPAAA (SEQ ID NO: 42).
  • a small beta barrel is a protein structural domain, highly conserved throughout evolution and hence exhibits a broad diversity of functions. Small beta barrels function as a single domain protein, or as part of a multi-domain protein.
  • the epitope-scaffold protein includes a small-synthetic ⁇ - barrel protein.
  • the small-synthetic ⁇ -barrel includes the sequence: MEQKPGTLMVYVVVGYNTDNTVDVVGGAQYAVSPYLFLDVGYGWNNSSLNFLEVGGGVSYKVSPDLEPYVKAGFE YNTDNTIKPTAGAGALYRVSPNLALMVEYGWNNSSLQKVAIGIAYKVKD (SEQ ID NO: 60).
  • Staphylococcal protein A is a cell-wall-bound pathogenicity factor from the bacterium Staphylococcus aureus.
  • the protein A specific binding protein, or Z-domain contains three well-defined alpha-helices corresponding to polypeptide segments Lys7 to Leu17 (helix 1), Glu24 to Asp36 (helix 2), and Ser41 to Ala54 (helix 3).
  • protein A includes the sequence: MKKKKIYSIRKLGVGIASVTLGTLLISGGVTPAANAAQHDEAQQNAFYQVLNMPNLNADQRNGFIQSLKDDPSQSANV LGEAQKLNDSQAPKADAQQNKFNKDQQSAFYEILNMPNLNEEQRNGFIQSLKDDPSQSTNVLGEAKKLNESQAPKA DNNFNKEQQNAFYEILNMPNLNEEQRNGFIQSLKDDPSQSANLLAEAKKLNDAQAPKADNKFNKEQQNAFYEILHLP NLTEEQRNGFIQSLKDDPSVSKEILAEAKKLNDAQAPKEEDNNKPGKEDGNKPGKEDGNKPGKEDNKKPGKEDGNK PGKEDNKKPGKEDGNK PGKEDNKKPGKEDGNK PGKEDNKKPGKEDGNK PGKEDNKKPGKEDGNK PGKEDNKKPGKEDGNK PGKEDNKKPGKEDGNK PGKEDNKKPGKEDGNK PGKE
  • the Z-domain of protein A includes the sequence: ADNKFNKEQQNAFYEILHLPNLNEEQRNGFIQSLKDDPSQSANLLAEAKKLNDAQAPK (SEQ ID NO: 44).
  • Cag-Z from Heliobacter pylori regulates Cag ⁇ -mediated CagA transport.
  • the protein includes a single compact L-shaped domain, composed of seven alpha-helices including 70% of the total residues. The presence of a disordered C-terminal tail and the nature of the molecular surface suggest that CagZ may participate in the interaction of effector proteins with one or more components of the H. pylori type IV secretion system on the cytoplasmic side of the inner membrane.
  • CagZ [Heliobacter pylori] includes the sequence: MELGFNETERQKILDSNKSLMGNANEVRDKFIQNYAASLKDSNDPQDFLRRVQELRINMQKNFISFDAYYNYLNNLVL ASYNRCKQEKTFAESTIKNELTLGEFVAEISDNFNNFMCDEVARISDLVASYLPREYLPPFIDGNMMGVAFQILGIDDF GKKLNEIVQDIGTKYIILSKNKTYLTSLERAKLITQLKLNLE (SEQ ID NO: 45).
  • Equine infectious anemia virus is a lentivirus having a single-stranded RNA genome of 8.2 kb and contains three major genes, gag (structural), pol (enzymes), and env (surface glycoproteins), which are flanked by long terminal repeats (LTRs).
  • the protein p26, encoded by the gag gene, is a capsid protein with high antigenicity.
  • p26 [equine infectious anemia virus] includes the sequence: MGDPLTWSKALKKLEKVTVSGSQKLTTGNCNWALSLVDLFHDTNFTKEKDWQLRDVIPLLEDVSQTLSGLEREAFEK TWWAISAVKMGLQINNAGDGKASFQLLKVKYEKKATGKRQPEPPEEYPIMIDGAGNRNFRPLTPRGYTTWVNTIQQN NLLNEASVNLFGILSVDCTSEEMNAFLDVVPGQAGQKQVLLDLLDKIAEDWDNRHPLPNPPLVAPAQGPIPMTARFIR GLGVPRERQMEPAFDQFRQTYRQWIIEAMTEGIKIMIGKPKAQNIRQGPKEPYPEFVDRLLSQIKSEGHPTEITKFLTD TLTIQNANEECKNAMRHLRPEDTLEEKLYACRDIGTTKQKMMLFAKALQAGLAGSMKGGIIKGGPPRAKQTCYNC
  • the transmembrane (T) domain of diphtheria toxin (DTT) is known to participate in the pH-dependent translocation of the catalytic C domain of the toxin across the endosomal membrane.
  • the transmembrane domain includes nine alpha-helices, two pairs of which are unusually apolar.
  • the T domain of diptheria toxin includes the sequence: SCINLDWDVIRDKTKTKIESLKEHGPIKNKMSESPNKTVSEEKAKQYLEEFHQTALEHPELSELKTVTGTNPVFAGANY AAWAVNVAQVIDSETADNLEKTTAALSILPGIGSVMGIADGAVHHNTEEIVAQSIALSSLMVAQAIPLVGELVDIGFAAY NFVESIINLFQVVHNSYNRPA (SEQ ID NO: 47).
  • the epitope can be placed anywhere in the epitope-scaffold protein, as long as the resulting epitope-scaffold protein can be specifically bound by molecules of the immune system (e.g., antibodies) or properly processed for presentation on antigen presenting cells.
  • molecules of the immune system e.g., antibodies
  • Methods for determining if a particular epitope- scaffold protein is specifically bound by its cognate binding molecule are disclosed herein and known to the person of ordinary skill in the art (see, for example, International Application Pub. Nos. WO 2006/091455 and WO 2005/111621).
  • an antibody-antigen complex or MHC-epitope complex can be assayed using a number of well-defined diagnostic assays including conventional immunoassay formats to detect and/or quantitate antigen- specific antibodies.
  • diagnostic assays include, for example, enzyme immunoassays, e.g., ELISA, cell-based assays, flow cytometry, radioimmunoassays, and immunohistochemical staining. Numerous competitive and non-competitive protein binding assays are known in the art and many are commercially available.
  • the epitope can be expressed on a virus-like particle (VLP).
  • VLP virus-like particle
  • a VLP is a non- replicating, viral shell, derived from any of several viruses.
  • VLPs are generally composed of one or more viral proteins, such as, those proteins referred to as capsid, coat, shell, surface and/or envelope proteins, or particle-forming polypeptides derived from these proteins. VLPs lack the viral components that are required for virus replication and thus represent a highly attenuated form of a virus.
  • the VLP can display a polypeptide (e.g., a Treponema pallidum epitope) that is capable of eliciting an immune response when administered to a subject.
  • VLPs and methods of their production are known and familiar to the person of ordinary skill in the art, and viral proteins from several viruses are known to form VLPs. VLPs can form spontaneously upon recombinant expression of the protein in an appropriate expression system.
  • VLPs can be detected by any suitable technique.
  • suitable techniques known in the art for detection of VLPs in a medium include, e.g., electron microscopy techniques, dynamic light scattering (DLS), selective chromatographic separation (e.g., ion exchange, hydrophobic interaction, and/or size exclusion chromatographic separation of the VLPs) and density gradient centrifugation.
  • VLPs can be isolated by known techniques, e.g., density gradient centrifugation and identified by characteristic density banding. See, for example, Baker et al. (1991) Biophys. J.60:1445-1456; and Hagensee et al. (1994) J.
  • the virus-like particle is derived from a virus selected from adeno-associated virus (AAV), Budgerigar fledgling disease virus (BFDV), Bluetongue virus (BTV), Ebola, Enterovirus 71, Goose hemorrhagic polyoma virus (GHPV), Hepatitis B virus (HBV), Hepatitis C virus (HCV), Hepatitis ⁇ virus (HDV), Hepatitis E virus (HEV), human immunodeficiency virus (HIV), Human papillomavirus (HPV), Infectious bursal disease virus (IBDV), influenza (e.g., Influenza A, Influenza A H1N1, Influenza A H3N2, avian influenza (H5N3)), Marburg virus (
  • the virus-like particle is derived from HEV, HIV, HPV, NDV, polyomavirus, or parvovirus.
  • the virus-like particle is an L1 capsid protein of HPV type 16.
  • the epitope is expressed on the DE loop of the L1 capsid protein of HPV type 16.
  • a VLP can be any size but viruses typically range in size from 20 nm – 200 nm.
  • a VLP can be 0 – 500 nm in size, 20-200 nm in size, 30-100 nm in size, or 50-60 nm in size.
  • a VLP is 50-60 nm in size.
  • a VLP is 55 nm in size.
  • a VLP that is 55 nm in size has optimal drainage to the lymph nodes.
  • an epitope can be expressed on a nanoparticle.
  • the term nanoparticle includes particles of spherical, elliptical, elongated shape or irregular structure.
  • Nanoparticles can include one or more materials. Nanoparticles also include nanocapsules and polyplex particles. Nanocapsules are nanoparticles containing a reservoir, for example an oil reservoir, which is surrounded by a polymer wall (Maria J. Alonso, Microparticulate systems for the delivery of proteins and vaccines, S. Cohen And Howard Bernstein, Marcel Dekker, New York 1996, p206).
  • a nanoparticle has dimensions controlled on the nanometer scale.
  • a nanoparticle typically has at least one dimension less than 1000 nm or 1 to 1000 nm.
  • Still a nanoparticle can refer to particles with larger dimensions.
  • a nanoparticle can have a diameter of 5000 nm, 2000 nm, 2500 nm, 1000 nm, 900 nm, 700 nm, 500 nm, 250 nm, 100 nm, 50 nm, 30 nm, 10 nm, 1 nm, or any size therebetween.
  • a nanoparticle is 0 – 500 nm in size, 20-200 nm in size, 30-100 nm in size, or 50-60 nm in size.
  • a nanoparticle is 50- 60 nm in size. In particular implementations, a nanoparticle is 55 nm in size.
  • a nanoparticle can include one or more materials, for instance.
  • the nanoparticle is made of self-assembling proteins including ferritin, lumazine, and encapsulin.
  • the epitope is linked to a subunit of the nanoparticle (such as ferritin, lumazine, or encapsulin).
  • the fusion protein self-assembles into a nanoparticle under appropriate conditions.
  • the epitope can be linked to the nanoparticle using linker chemistry or a peptide linker.
  • a self-assembling protein can be used in a nanoparticle.
  • Self-assembling proteins include ferritin, lumazine, and encapsulin.
  • Ferritin is a protein whose primary function is intracellular iron storage.
  • Ferritin is a protein whose main function is intracellular iron storage.
  • Ferritin is composed of 24 subunits, each composed of four alpha-helical bundles that self-assemble into quaternary structures with octahedral symmetry (Cho KJ et al. J Mol Biol.2009; 390: 83). -98).
  • LS is an enzyme present in a variety of organisms including archaea, bacteria, fungi, and plants (Weber S.E. Flavins and Flavoproteins. Methods and Protocols, Series: Methods in Molecular Biology.2014).
  • the LS monomer is 150 amino acids long and includes a beta-sheet with tandem alpha-helical flanking on its sides.
  • a number of different quaternary structures have been reported for LS, showing morphological diversity ranging from homopentamers to symmetrical assembly of 12 pentamers forming 150 ⁇ diameter capsids.
  • LS cages of more than 100 subunits have also been described (Zhang X. et al.
  • Encapsulins novel protein cage nanoparticles isolated from the thermophilic bacterium Thermotoga maritima, can also be used as a platform to present antigens on the surface of self-assembling nanoparticles.
  • Nanoparticle can also be used to describe other types and materials of particles including nanoshells, nanobeads, or nanodots. Nanoparticles can be made from organic matter and/or inorganic matter. They can be made of any suitable materials that allow for the conjugation of epitopes disclosed herein to their surface. Examples of suitable materials include: ceramics, gold, carbon, glass (silica), polymers, and magnetic materials.
  • Suitable polymers include polystyrene, poly-(methyl methacrylate), poly-(lactic acid), (poly-(lactic-co -glycolic acid)), polyesters, polyethers, polyolef ⁇ ns, polyalkylene oxides, polyamides, polyurethanes, polysaccharides, celluloses, polyisoprenes, methylstyrene, acrylic polymers, thoria sol, latex, nylon, Teflon cross- linked dextrans (e.g., Sepharose), chitosan, agarose, and cross-linked micelles. Additional examples include carbon graphited, titanium dioxide, and paramagnetic materials.
  • microparticles can be made of one or more materials.
  • a nanoparticle includes a gold nanoparticle, a carbon nanoparticle or a silica nanoparticle.
  • the proteins and vaccines disclosed herein are produced from a gene using a protein expression system.
  • Protein expression systems can utilize DNA constructs (e.g., chimeric genes, expression cassettes, expression vectors, recombination vectors) including a nucleic acid sequence encoding the protein or proteins of interest operatively linked to appropriate regulatory sequences.
  • DNA constructs are not naturally-occurring DNA molecules and are useful for introducing DNA into host-cells to express selected proteins of interest.
  • a DNA construct that encodes a vaccine protein can be inserted into cells (e.g., bacterial, mammalian, insect, etc.), which can produce the vaccine protein encoded by the DNA construct.
  • Operatively linked refers to the linking of DNA sequences (including the order of the sequences, the orientation of the sequences, and the relative spacing of the various sequences) in such a manner that the encoded protein is expressed. Methods of operatively linking expression control sequences to coding sequences are well known in the art.
  • Expression control sequences are DNA sequences involved in any way in the control of transcription or translation. Suitable expression control sequences and methods of making and using them are well known in the art. Expression control sequences generally include a promoter.
  • the promoter may be inducible or constitutive. It may be naturally occurring, may be composed of portions of various naturally occurring promoters, or may be partially or totally synthetic.
  • the promoter may include, or be modified to include, one or more enhancer elements.
  • the promoter includes a plurality of enhancer elements. Promoters including enhancer elements can provide for higher levels of transcription as compared to promoters that do not include them.
  • the coding sequences can be operatively linked to a 3’ untranslated sequence.
  • the 3’ untranslated sequence can include a transcription termination sequence and a polyadenylation sequence.
  • the 3’ untranslated region can be obtained, for example, from the flanking regions of genes.
  • a 5’ untranslated leader sequence can also be employed.
  • the 5’ untranslated leader sequence is the portion of an mRNA that extends from the 5’ CAP site to the translation initiation codon.
  • a “hisavi” tag can be added to the N-terminus or C-terminus of a gene by the addition of nucleotides coding for the Avitag amino acid sequence, “GLNDIFEAQKIEWHE” (SEQ ID NO: 48), as well as the 6xhistidine tag “HHHHHH” (SEQ ID NO: 49).
  • the Avitag avidity tag can be biotinylated by a biotin ligase to allow for biotin-avidin or biotin-streptavidin based interactions for protein purification, as well as for immunobiology (such as immunoblotting or immunofluorescence) using anti-biotin antibodies.
  • the 6xhistidine tag allows for protein purification using Ni- 2+ affinity chromatography.
  • Other tags include: Flag tag (DYKDDDDK; SEQ ID NO: 50), Xpress tag (DLYDDDDK; SEQ ID NO: 51), Calmodulin tag (KRRWKKNFIAVSAANRFKKISSSGAL; SEQ ID NO: 52), Polyglutamate tag, HA tag (YPYDVPDYA; SEQ ID NO: 53), Myc tag (EQKLISEEDL; SEQ ID NO: 54), Strep tag (which refers the original STREP ® tag (WRHPQFGG; SEQ ID NO: 55), STREP ® tag II (WSHPQFEK SEQ ID NO: 56 (IBA Institut fur Bioanalytik, Germany); see, e.g., US 7,981,632), Softag 1 (SLAELLNAGLGGS; SEQ ID NO: 57), Softag 3 (TQDPSRVG; SEQ ID NO: 58), and V5 tag (G
  • syphilis vaccines can be produced using, for example, human suspension cells and/or the Daedalus expression system as described in Pechman et al., Am J Physiol 294: R1234-R1239, 2008.
  • the Daedalus system utilizes inclusion of minimized ubiquitous chromatin opening elements in transduction vectors to reduce or prevent genomic silencing and to help maintain the stability of decigram levels of expression. This system can bypass tedious and time-consuming steps of other protein production methods by employing the secretion pathway of serum-free adapted human suspension cell lines, such as 293 Freestyle.
  • the concatemer-encoding sequence is obtained by joining pre-synthesized DNA fragments by overlapping PCR, which are then cloned into an expression vector.
  • the expression vector includes the E. coli expression vector pET-32.
  • the concatemer-encoding sequence includes a tag for purification.
  • the tag is a 5xHis-tag.
  • an epitope-VLP epitope expressed on a VLP is produced by the epitope(s) sequence being inserted into a p16L1L2 vector using primer extension to replace a portion of the delivery scaffold followed by Infusion cloning.
  • VLPs will be produced according to the protocol developed by Buck and colleagues (“Buck”; Buck et al., Journal of virology.2004, 78(2):751-7; and Cardone et al., MBio.2014, 5(4):e01104-14). Briefly, plasmids are used to transfect 7x10 6 cells.
  • the cells are human embryonic kidney (HEK) cell line 293T, previously modified to express high levels of SV40 LT antigen to drive L1 expression from the p16L1L2 vector.
  • HEK human embryonic kidney
  • VLPs will be given time to mature by incubation of the cell lysates at an incubation temperature.
  • the incubation temperature is 30°C-50°C. In particular implementations, the incubation temperature is 37°C.
  • VLPs can be harvested by centrifugation. In particular implementations, centrifugation includes for a select time at a select temperature with a select speed. In particular implementations, the select time is 0-12 hours, 1-5 hours, or 2-4 hours. In particular implementations, the select time is 3.5 hours. In particular implementations, the select temperature is 0-50°C, 5-30°C, 10-20°C, or 15-18°C. In particular implementations, the select temperature s 16°C.
  • the select speed is 10,000xg to 900,000xg, 100,000xg to 500,000xg, or 200,000xg to 300,000xg. In particular implementations, the select speed is 234,000xg.
  • centrifugation includes ultracentrifugation through an Optiprep (60% iodixanol) step gradient for 3.5 hours at 16°C at 234,000xg.
  • the DNA constructs can be introduced by transfection, a technique that involves introduction of foreign DNA into the nucleus of eukaryotic cells.
  • the proteins can be synthesized by transient transfection (e.g., in which the DNA does not integrate with the genome of the eukaryotic cells, but the genes are expressed for 24-96 hours).
  • transient transfection e.g., in which the DNA does not integrate with the genome of the eukaryotic cells, but the genes are expressed for 24-96 hours.
  • Various methods can be used to introduce the foreign DNA into the host-cells, and transfection can be achieved by chemical-based means including by the calcium phosphate, by dendrimers, by liposomes, and by the use of cationic polymers.
  • Non-chemical methods of transfection include electroporation, sono-poration, optical transfection, protoplast fusion, impalefection, and hydrodynamic delivery.
  • transfection can be achieved by particle-based methods including gene gun techniques, wherein the DNA construct is coupled to a nanoparticle of an inert solid which is then "shot" directly into the target- cell's nucleus.
  • particle-based transfection methods include magnet assisted transfection and impalefection.
  • Nucleic acid sequences encoding proteins disclosed herein can be derived by those of ordinary skill in the art. Nucleic acid sequences can also include one or more of various sequence polymorphisms, mutations, and/or sequence variants (e.g., splice variants or codon optimized variants). In particular implementations, the sequence polymorphisms, mutations, and/or sequence variants do not affect the function of the encoded protein.
  • syphilis therapeutics can be formulated alone or in combination into compositions for administration to subjects.
  • the syphilis therapeutics include immunogenic compositions.
  • An immunogenic composition refers to an agent that stimulates an innate and/or an adaptive immune response in a subject.
  • Salts and/or pro-drugs of syphilis therapeutics can also be used.
  • a pharmaceutically acceptable salt includes any salt that retains the activity of the syphilis therapeutic and is acceptable for pharmaceutical use.
  • a pharmaceutically acceptable salt also refers to any salt which may form in vivo as a result of administration of an acid, another salt, or a prodrug which is converted into an acid or salt.
  • Suitable pharmaceutically acceptable acid addition salts can be prepared from an inorganic acid or an organic acid. Examples of such inorganic acids are hydrochloric, hydrobromic, hydroiodic, nitric, carbonic, sulfuric and phosphoric acid. Appropriate organic acids can be selected from aliphatic, cycloaliphatic, aromatic, arylaliphatic, heterocyclic, carboxylic and sulfonic classes of organic acids.
  • Suitable pharmaceutically acceptable base addition salts include metallic salts made from aluminum, calcium, lithium, magnesium, potassium, sodium and zinc or organic salts made from N,N'-dibenzylethylene-diamine, chloroprocaine, choline, diethanolamine, ethylenediamine, N-methylglucamine, lysine, arginine and procaine.
  • a prodrug includes an active ingredient which is converted to a therapeutically active compound after administration, such as by cleavage of a syphilis therapeutic or by hydrolysis of a biologically labile group.
  • compositions disclosed herein include a syphilis therapeutic of at least 0.1% w/v or w/w of the composition; at least 1% w/v or w/w of composition; at least 10% w/v or w/w of composition; at least 20% w/v or w/w of composition; at least 30% w/v or w/w of composition; at least 40% w/v or w/w of composition; at least 50% w/v or w/w of composition; at least 60% w/v or w/w of composition; at least 70% w/v or w/w of composition; at least 80% w/v or w/w of composition; at least 90% w/v or w/w of composition; at least 95% w/v or w/w of composition; or at least 99% w/v or w/w of composition.
  • Exemplary generally used pharmaceutically acceptable carriers include any and all absorption delaying agents, antioxidants, binders, buffering agents, bulking agents or fillers, chelating agents, coatings, disintegration agents, dispersion media, gels, isotonic agents, lubricants, preservatives, salts, solvents or co-solvents, stabilizers, surfactants, and/or delivery vehicles.
  • Exemplary antioxidants include ascorbic acid, methionine, and vitamin E.
  • Exemplary buffering agents include citrate buffers, succinate buffers, tartrate buffers, fumarate buffers, gluconate buffers, oxalate buffers, lactate buffers, acetate buffers, phosphate buffers, histidine buffers, and/or trimethylamine salts.
  • An exemplary chelating agent is EDTA.
  • Exemplary isotonic agents include polyhydric sugar alcohols including trihydric or higher sugar alcohols, such as glycerin, erythritol, arabitol, xylitol, sorbitol, or mannitol.
  • Exemplary preservatives include phenol, benzyl alcohol, meta-cresol, methyl paraben, propyl paraben, octadecyldimethylbenzyl ammonium chloride, benzalkonium halides, hexamethonium chloride, alkyl parabens such as methyl or propyl paraben, catechol, resorcinol, cyclohexanol, and 3-pentanol.
  • Stabilizers refer to a broad category of excipients which can range in function from a bulking agent to an additive which solubilizes the syphilis therapeutic or helps to prevent denaturation or adherence to the container wall.
  • Typical stabilizers can include polyhydric sugar alcohols; amino acids, such as arginine, lysine, glycine, glutamine, asparagine, histidine, alanine, ornithine, L-leucine, 2-phenylalanine, glutamic acid, and threonine; organic sugars or sugar alcohols, such as lactose, trehalose, stachyose, mannitol, sorbitol, xylitol, ribitol, myoinisitol, galactitol, glycerol, and cyclitols, such as inositol; PEG; amino acid polymers; sulfur-containing reducing agents, such as urea, glutathione, thioctic acid, sodium thioglycolate, thioglycerol, ⁇ -monothioglycerol, and sodium thiosulfate; low molecular weight polypeptides (i.
  • Stabilizers are typically present in the range of from 0.1 to 10,000 parts by weight based on therapeutic weight.
  • the compositions disclosed herein can be formulated for administration by, for example, injection, inhalation, infusion, perfusion, lavage, or ingestion.
  • compositions disclosed herein can further be formulated for intravenous, intradermal, intraarterial, intranodal, intralymphatic, intraperitoneal, intralesional, intraprostatic, intravaginal, intrarectal, topical, intrathecal, intramuscular, intravesicular, oral and/or subcutaneous administration and more particularly by intravenous, intradermal, intraarterial, intranodal, intralymphatic, intraperitoneal, intralesional, intraprostatic, intravaginal, intrarectal, intrathecal, intramuscular, intravesicular, and/or subcutaneous injection.
  • compositions can be formulated as aqueous solutions, such as in buffers including Hanks' solution, Ringer's solution, or physiological saline.
  • the aqueous solutions can include formulatory agents such as suspending, stabilizing, and/or dispersing agents.
  • the formulation can be in lyophilized and/or powder form for constitution with a suitable vehicle, e.g., sterile pyrogen-free water, before use.
  • a suitable vehicle e.g., sterile pyrogen-free water
  • the compositions can be formulated as tablets, pills, dragees, capsules, liquids, gels, syrups, slurries, suspensions and the like.
  • suitable excipients include binders (gum tragacanth, acacia, cornstarch, gelatin), fillers such as sugars, e.g., lactose, sucrose, mannitol and sorbitol; dicalcium phosphate, starch, magnesium stearate, sodium saccharine, cellulose, magnesium carbonate; cellulose preparations such as maize starch, wheat starch, rice starch, potato starch, gelatin, gum tragacanth, methyl cellulose, hydroxypropylmethyl-cellulose, sodium carboxy-methylcellulose, and/or polyvinylpyrrolidone (PVP); granulating agents; and binding agents.
  • binders such as sugars, e.g., lactose, sucrose, mannitol and sorbitol
  • dicalcium phosphate starch, magnesium stearate, sodium saccharine, cellulose, magnesium carbonate
  • cellulose preparations such as maize starch, wheat starch, rice starch
  • disintegrating agents can be added, such as corn starch, potato starch, alginic acid, cross-linked polyvinylpyrrolidone, agar, or alginic acid or a salt thereof such as sodium alginate.
  • solid dosage forms can be sugar-coated or enteric-coated using standard techniques. Flavoring agents, such as peppermint, oil of wintergreen, cherry flavoring, orange flavoring, etc. can also be used.
  • Compositions can be formulated as an aerosol. In particular implementations, the aerosol is provided as part of an anhydrous, liquid or dry powder inhaler.
  • Aerosol sprays from pressurized packs or nebulizers can also be used with a suitable propellant, e.g., dichlorodifluoromethane, trichlorofluoromethane, dichlorotetrafluoroethane, carbon dioxide or other suitable gas.
  • a dosage unit may be determined by providing a valve to deliver a metered amount.
  • Capsules and cartridges of gelatin for use in an inhaler or insufflator may also be formulated including a powder mix of syphilis therapeutic composition and a suitable powder base such as lactose or starch.
  • Compositions can also be formulated as depot preparations.
  • compositions can be formulated as sustained-release systems utilizing semipermeable matrices of solid polymers including at least one syphilis therapeutic.
  • sustained-release systems may, depending on their chemical nature, release one or more syphilis therapeutics following administration for a few weeks up to over 100 days.
  • Depot preparations can be administered by injection; parenteral injection; instillation; or implantation into soft tissues, a body cavity, or occasionally into a blood vessel with injection through fine needles.
  • Depot formulations can include a variety of bioerodible polymers including poly(lactide), poly(glycolide), poly(caprolactone) and poly(lactide)-co(glycolide) (PLG) of desirable lactide:glycolide ratios, average molecular weights, polydispersities, and terminal group chemistries. Blending different polymer types in different ratios using various grades can result in characteristics that borrow from each of the contributing polymers.
  • solvents for example, dichloromethane, chloroform, ethyl acetate, triacetin, N-methyl pyrrolidone, tetrahydrofuran, phenol, or combinations thereof
  • Other useful solvents include water, ethanol, dimethyl sulfoxide (DMSO), N-methyl- 2-pyrrolidone (NMP), acetone, methanol, isopropyl alcohol (IPA), ethyl benzoate, and benzyl benzoate.
  • Exemplary release modifiers can include surfactants, detergents, internal phase viscosity enhancers, complexing agents, surface active molecules, co-solvents, chelators, stabilizers, derivatives of cellulose, (hydroxypropyl)methyl cellulose (HPMC), HPMC acetate, cellulose acetate, pluronics (e.g., F68/F127), polysorbates, Span® (Croda Americas, Wilmington, Delaware), poly(vinyl alcohol) (PVA), Brij® (Croda Americas, Wilmington, Delaware), sucrose acetate isobutyrate (SAIB), salts, and buffers.
  • surfactants e.g., hydroxypropyl)methyl cellulose (HPMC), HPMC acetate, cellulose acetate, pluronics (e.g., F68/F127), polysorbates, Span® (Croda Americas, Wilmington, Delaware), poly(vinyl alcohol) (PVA), Brij® (Croda Americas, Wilmington, Delaware), suc
  • Excipients that partition into the external phase boundary of microparticles such as surfactants including polysorbates, dioctylsulfosuccinates, poloxamers, PVA, can also alter properties including particle stability and erosion rates, hydration and channel structure, interfacial transport, and kinetics in a favorable manner.
  • Additional processing of the disclosed sustained release depot formulations can utilize stabilizing excipients including mannitol, sucrose, trehalose, and glycine with other components such as polysorbates, PVAs, and dioctylsulfosuccinates in buffers such as Tris, citrate, or histidine.
  • compositions include a vaccine adjuvant and/or a secondary syphilis treatment. Examples of secondary syphilis treatments are described elsewhere herein.
  • the compositions disclosed herein may be combined with one or more adjuvants to enhance the immune response. In other implementations, the compositions are prepared without adjuvants, and thus are available for administration as non-adjuvant compositions.
  • a squalene-based adjuvant can be used.
  • Squalene is part of the group of molecules known as triterpenes, which are all hydrocarbons with 30 carbon molecules. Squalene can be derived from certain plant sources, such as rice bran, wheat germ, amaranth seeds, and olives, as well as from animal sources, such as shark liver oil.
  • the squalene-based adjuvant is MF59® (Novartis, Basel, Switzerland).
  • MF59 Novartis, Basel, Switzerland
  • AddavaxTM InvivoGen, San Diego, CA.
  • MF59 has been FDA approved for use in an influenza vaccine, and studies indicate that it is safe for use during pregnancy (Tsai T, et al.
  • squalene-based adjuvants can include 0.1%-20% (v/v) squalene oil.
  • squalene-based adjuvants can include 5%(v/v) squalene oil.
  • a squalene-based adjuvant includes the RIBI Adjuvant System ® (RAS®) (Corxia Corporation, Seattle, WA).
  • RAS® (Corxia Corporation) is a commercial adjuvant system that uses a small quantity of metabolizable squalene oil and Tween 80 surfactant in which the antigen is incorporated before emulsion in water.
  • the adjuvant may be alum.
  • Alum refers to a family of salts that contain a monovalent cation, and a trivalent metal, such as aluminum or chromium.
  • Aluminum hydroxide, aluminum phosphate, and potassium aluminum sulphate (alum) can each be used as an adjuvant in a vaccine.
  • vaccines can include alum in the amounts of 1-1000 ⁇ g/dose or 0.1mg-10mg/dose.
  • Saponin-containing adjuvants can also be combined with the immunogens disclosed herein.
  • Saponins are glycosides derived from the bark of the Quillaja saponaria Molina tree. Usually, saponins are obtained using a multi- step purification process resulting in the formation of several fractions.
  • saponin fraction from Quillaja saponaria Molina is used generically to describe a semi-purified or specified saponin fraction of Quillaja saponaria, or a substantially purified fraction thereof.
  • Liposome-based adjuvants may act as both delivery systems for subunit antigens and as immunopotentiators, and they are highly versatile adjuvants, as they can be tailored through (i) the choice of lipid composition, (ii) the inclusion of immunostimulating compounds, (iii) the choice of formulation method and (iv) the mode of antigen and immunostimulator association.
  • the adjuvant is a liposome-based adjuvant.
  • a vaccine adjuvant can include any kind of Toll-like receptor ligand (TLR-based) or combinations thereof (e.g.
  • CpG, Cpg-28 (a TLR9 agonist), polyriboinosinic polyribocytidylic acid (Poly(I:C)), ⁇ - galactoceramide, MPLA, Motolimod (VTX-2337, a novel TLR8 agonist developed by VentiRx), IMO-2055 (EMD1201081), TMX-101 (imiquimod), MGN1703 (a TLR9 agonist), G100 (a stabilized emulsion of the TLR4 agonist glucopyranosyl lipid A), Entolimod (a derivative of Salmonella flagellin also known as CBLB502), Hiltonol (a TLR3 agonist), and Imiquimod), and/or inhibitors of heat-shock protein 90 (Hsp90), such as 17-DMAG (17- dimethylaminoethylamino-17-demethoxygeldanamycin).
  • Hsp90 heat-shock protein 90
  • an adjuvant can include a TLR4 agonist glucopyranosyl lipid A (GLA) formulated in stable emulsion (GLA SE; Odegard et al. (2016) Vaccine 34(1): 101-109).
  • GLA SE TLR4 agonist glucopyranosyl lipid A
  • an adjuvant includes resiquimod (TLR 7/8 agonist), muramyl dipeptide (MDP, a NOD2 agonist), synthetic diacylated lipoprotein FSL-1 (TLR2/TLR6 agonist), or GLA.
  • TLR7/8 agonist muramyl dipeptide
  • MDP muramyl dipeptide
  • TLR2/TLR6 agonist synthetic diacylated lipoprotein FSL-1
  • one or more STING agonists are used as a vaccine adjuvant.
  • STING is an abbreviation of "stimulator of interferon genes", which is also known as “endoplasmic reticulum interferon stimulator (ERIS)", “mediator of IRF3 activation (MITA)”, “MPYS” or “transmembrane protein 173 (TM173)”.
  • ERIS endoplasmic reticulum interferon stimulator
  • MIMA immediate-reactive reticulum interferon stimulator
  • MPYS transmembrane protein 173
  • TM173 transmembrane protein 173
  • STING agonists include cyclic molecules with one or two phosphodiester linkages, and/or one or two phosphorothioate diester linkages, between two nucleotides.
  • nucleotide linkages (abbreviated as (3',3')); (3',5')-(2',5') nucleotide linkages (abbreviated as (3',2')); (2',5')-(3',5') nucleotide linkages (abbreviated as (2',3')); and (2',5')-(2',5') nucleotide linkages (abbreviated as (2',2')).
  • Nucleotide refers to any nucleoside linked to a phosphate group at the 5', 3' or 2' position of the sugar moiety.
  • STING agonists include c-AIMP; (3’,2’)c-AIMP; (2’,2’)c-AIMP; (2’,3’)c-AIMP; c- AIMP(S); c-(dAMP-dIMP); c-(dAMP-2’FdIMP); c-(2’FdAMP-2’FdIMP); (2’,3’)c-(AMP-2’FdIMP); c-[2’FdAMP(S)- 2’FdIMP(S)]; c-[2’FdAMP(S)-2’FdIMP(S)](POM)2; and DMXAA. Additional examples of STING agonists are described in WO2016/145102.
  • the adjuvant used in a composition of the invention herein is an attenuated lipid A derivative (ALD).
  • ALDs are lipid A-like molecules that have been altered or constructed to lower pyrogenicity, local Shwarzman reactivity and toxicity.
  • Useful ALDs include monophosphoryl lipid A (MLA) and 3-deacylated monophosphoryl lipid A (3D-MLA). See for example U.S. Pat. No.4,436,727 issued Mar.13, 1984, assigned to Ribi ImmunoChem Research, Inc., which discloses monophosphoryl lipid A and its manufacture.
  • an adjuvant includes a Ribi-like adjuvant.
  • a Ribi-like adjuvant includes mono-phosphoryl lipid A (MPLA, a TLR4 agonist) from S. minnesota, synthetic trehalose dimycolate (TDM), and cell wall skeleton (CWS, a TLR2 and TLR4 agonist) from M. bovis.
  • MPLA mono-phosphoryl lipid A
  • TDM synthetic trehalose dimycolate
  • CWS cell wall skeleton
  • a Ribi-like adjuvant includes mono-phosphoryl lipid A (MPLA, a TLR4 agonist) from S. minnesota, synthetic trehalose dimycolate (TDM), and muramyl dipeptide (MDP).
  • MPLA mono-phosphoryl lipid A
  • TDM synthetic trehalose dimycolate
  • MDP muramyl dipeptide
  • an adjuvant can include a carbomer-lecithin-based adjuvant (e.g., AdjuplexTM, Millipore Sigma, Burlington, MA; Wegmann et al. (2015) Clin Vaccine Immunol. CVI-00736).
  • Other immune stimulants can also be used as vaccine adjuvants.
  • exemplary small molecule immune stimulants include TGF- ⁇ inhibitors, SHP-inhibitors, STAT-3 inhibitors, and/or STAT-5 inhibitors.
  • exemplary siRNA capable of down-regulating immune-suppressive signals or oncogenic pathways can be used whereas any plasmid DNA (such as minicircle DNA) encoding immune-stimulatory proteins can also be used.
  • the immune stimulant may be a cytokine and or a combination of cytokines, such as IL-2, IL-12 or IL-15 in combination with IFN- ⁇ , IFN- ⁇ or IFN- ⁇ , or GM-CSF, or any effective combination thereof, or any other effective combination of cytokines.
  • cytokines stimulate TH1 responses
  • cytokines that stimulate TH2 responses may also be used, such as IL-4, IL-10, IL-11, or any effective combination thereof.
  • combinations of cytokines that stimulate TH1 responses along with cytokines that stimulate TH2 responses may be used.
  • the adjuvant induces a strong Th1-type response.
  • Molecules that induce a strong Th1-type response include TLR2, TLR4, and NOD2 agonists.
  • Molecules that induce a strong Th1-type response include MPLA, TDM, synthetic diacylated lipoprotein FSL-1 (TLR2/TLR6 agonist), and muramyl dipeptide (MDP, a NOD2 agonist).
  • the adjuvant includes GLA, risiquimod, MDP, FSL-1, and/or Qui-A.
  • the adjuvant includes TiterMax® Gold (Titermax USA, Inc., Norcross, GA).
  • Any composition disclosed herein can advantageously include any other pharmaceutically acceptable carriers which include those that do not produce significantly adverse, allergic, or other untoward reactions that outweigh the benefit of administration.
  • compositions disclosed herein include treating subjects (e.g., humans, veterinary animals (dogs, cats, reptiles, birds) livestock (e.g., horses, cattle, goats, pigs, chickens) and research animals (e.g., monkeys, rats, mice, fish) with compositions disclosed herein. Treating subjects includes delivering therapeutically effective amounts.
  • Therapeutically effective amounts include those that provide effective amounts, prophylactic treatments and/or therapeutic treatments.
  • An “effective amount” is the amount of a composition necessary to result in a desired physiological change in the subject.
  • an effective amount can provide an immunogenic effect.
  • Effective amounts are often administered for research purposes.
  • Effective amounts disclosed herein can cause a statistically-significant effect in an in vitro assay, an animal model or clinical study relevant to the assessment of an infection’s development, progression, and/or resolution, as well as the effects of the infection.
  • An immunogenic composition can be provided in an effective amount, wherein the effective amount stimulates an immune response.
  • a prophylactic treatment includes a treatment administered to a subject who does not display signs or symptoms of an infection or displays only early signs or symptoms of an infection such that treatment is administered for the purpose of diminishing or decreasing the risk of developing the infection.
  • a prophylactic treatment functions as a preventative treatment against an infection and/or the potential effects of an infection.
  • Particular uses of the compositions include use as prophylactic vaccines. Vaccines increase the immunity of a subject against a particular infection. Therefore, "syphilis vaccine” can refer to a treatment that increases the immunity of a subject against syphilis.
  • a vaccine may be administered prophylactically, for example to a subject that is immunologically naive (e.g., no prior exposure or experience with syphilis).
  • a vaccine may be administered therapeutically to a subject who has been exposed to Treponema pallidum or syphilis.
  • a vaccine can be used to ameliorate a symptom and/or complication associated with syphilis, examples of each of which are described elsewhere herein.
  • a syphilis vaccine is a therapeutically effective composition including a Treponema pallidum epitope that induces an immune response in a subject against syphilis.
  • the immune system generally is capable of producing an innate immune response and an adaptive immune response.
  • An innate immune response generally can be characterized as not being substantially antigen specific and/or not generating immune memory.
  • An adaptive immune response can be characterized as being substantially antigen specific, maturing over time (e.g., increasing affinity and/or avidity for antigen), and in general can produce immunologic memory. Even though these and other functional distinctions between innate and adaptive immunity can be discerned, the skilled artisan will appreciate that the innate and adaptive immune systems can be integrated and therefore can act in concert. [0134]
  • administration of a syphilis vaccine can further include administration of one or more adjuvants.
  • adjuvant refers to material that enhances the immune response to a vaccine antigen and is used herein in the customary use of the term. The precise mode of action is not understood for all adjuvants, but such lack of understanding does not prevent their clinical use for a wide variety of vaccines.
  • "Immune response” refers to a response of the immune system to produce Treponema pallidum neutralizing antibodies in response to B cells binding T. pallidum epitopes.
  • the immune response causes production of antibodies to neutralize T. pallidum.
  • an immune response to a syphilis vaccine can be an innate and/or adaptive response.
  • an adaptive immune response can be a "primary immune response" which refers to an immune response occurring on the first exposure of a "naive" subject to T. pallidum or vaccine.
  • a primary immune response for example, after a lag or latent period of from 3 to 14 days depending on, for example, the composition, dose, and subject, antibodies to T. pallidum can be produced.
  • IgM production lasts for several days followed by IgG production and the IgM response can decrease.
  • Antibody production can terminate after several weeks but memory cells can be produced.
  • an adaptive immune response can be a "secondary immune response", “anamnestic response,” or “booster response” which refer to the immune response occurring on a second and subsequent exposure of a subject to T. pallidum or vaccine.
  • a secondary immune response memory cells respond to the T. pallidum or vaccine and therefore the secondary immune response can differ from a primary immune response qualitatively and/or quantitatively.
  • the lag period of a secondary antibody response can be shorter, the peak antibody titer can be higher, higher affinity antibody can be produced, and/or antibody can persist for a greater period of time.
  • an immune response against T. pallidum will include antibody production against T. pallidum.
  • a "therapeutic treatment” includes a treatment administered to a subject who displays symptoms or signs of an infection and is administered to the subject for the purpose of diminishing or eliminating those signs or symptoms of the infection or effects of the infection.
  • the therapeutic treatment can reduce, control, or eliminate the presence or activity of the infection and/or reduce, control or eliminate side effects of the infection.
  • a therapeutic treatment can reduce, control, or eliminate a primary infection with T. pallidum.
  • a therapeutic treatment can reduce or eliminate the symptoms of syphilis.
  • a therapeutically effective amount reduces or prevents transmission of T. pallidum and/or syphilis.
  • a therapeutically effective amount alleviates or reduces the severity or occurrence of symptoms and/or complications associated with T. pallidum infection.
  • exemplary symptoms include skin rash (e.g., on palms of the hands and soles of the fee), sores (e.g., mucous patches in mouth, vagina, or penis), warty patches, fever, general ill feeling, loss of appetite, muscle and joint pain, swollen lymph nodes, vision changes, hair loss, heart damage, tumors, and central nervous system disorders (e.g., neurosyphilis).
  • a therapeutically effective amount reduces the duration of infection with syphilis for a subject as compared to a subject that has not received a syphilis vaccine disclosed herein.
  • a therapeutically effective amount reduces the time to sustained non-detectable syphilis in the sera, blood, or urine in a patient infected with the bacteria as compared to a subject that has not received a syphilis vaccine disclosed herein.
  • a therapeutically effective amount reduces the spread and symptoms as compared to a subject that has not received a syphilis vaccine disclosed herein.
  • administering may accomplish more than one treatment type.
  • therapeutically effective amounts also referred to herein as doses
  • doses can be initially estimated based on results from in vitro assays and/or animal model studies. Such information can be used to more accurately determine useful doses in subjects of interest.
  • the actual dose amount administered to a particular subject can be determined by a physician, veterinarian or researcher taking into account parameters such as physical and physiological factors including target, body weight, severity of infection, stage of infection, effects of infection, previous or concurrent therapeutic interventions, idiopathy of the subject and route of administration.
  • Useful doses can range from 0.1 to 5 ⁇ g/kg or from 0.5 to 1 ⁇ g /kg.
  • a dose can include 1 ⁇ g /kg, 15 ⁇ g /kg, 30 ⁇ g /kg, 50 ⁇ g/kg, 55 ⁇ g/kg, 70 ⁇ g/kg, 90 ⁇ g/kg, 150 ⁇ g/kg, 350 ⁇ g/kg, 500 ⁇ g/kg, 750 ⁇ g/kg, 1000 ⁇ g/kg, 0.1 to 5 mg/kg or from 0.5 to 1 mg/kg.
  • a dose can include 1 mg/kg, 10 mg/kg, 30 mg/kg, 50 mg/kg, 70 mg/kg, 100 mg/kg, 300 mg/kg, 500 mg/kg, 700 mg/kg, 1000 mg/kg or more.
  • Therapeutically effective amounts can be achieved by administering single or multiple doses during the course of a treatment regimen (e.g., daily, every other day, every 3 days, every 4 days, every 5 days, every 6 days, weekly, every 2 weeks, every 3 weeks, monthly, every 2 months, every 3 months, every 4 months, every 5 months, every 6 months, every 7 months, every 8 months, every 9 months, every 10 months, every 11 months or yearly).
  • the syphilis vaccine described herein can be administered in combination or alternation with a secondary syphilis treatment.
  • the secondary syphilis treatment includes antibiotics such as penicillin (e.g., Benzathine penicillin G), amoxicillin, ampicillin, doxycycline, tetracycline, or ceftriaxone, or combinations thereof.
  • the syphilis vaccine disclosed herein can be used as a booster vaccine, to increase or modify or alter immune responses induced by a prior syphilis vaccine.
  • the syphilis vaccine disclosed herein can be used as a booster vaccine following infection and recovery from syphilis.
  • the syphilis vaccine described herein can be administered on top of the current standard of care for syphilis patients, or in combination or alternation with any other compound or therapy that the healthcare provider deems beneficial for the patient.
  • the combination and/or alternation therapy can be therapeutic, adjunctive, or palliative.
  • the pharmaceutical compositions described herein can be administered by, without limitation, injection, inhalation, infusion, perfusion, lavage or ingestion.
  • Routes of administration can include intravenous, intradermal, intramucosal, intraarterial, intraparenteral, intranasal, intranodal, intralymphatic, intraperitoneal, intralesional, intraprostatic, intravaginal, intrarectal, topical, intrathecal, intramuscular, intravesicular, oral, subcutaneous, and/or sublingual administration and more particularly intramuscular, subcutaneous, and/or intradermal injection. [0151] (V) Kits.
  • kits including one or more containers including one or more of the Treponema pallidum epitope sequences described herein, nucleic acids encoding Treponema pallidum epitope sequences, vectors, Treponema pallidum epitope sequences on a delivery scaffold, modified cells (e.g., cells modified to express a vaccine disclosed herein), and/or compositions and/or adjuvants, anti-infective agents, or secondary vaccines described herein.
  • Associated with such container(s) can be a notice in the form prescribed by a governmental agency regulating the manufacture, use, or sale of pharmaceuticals or biological products, which notice reflects approval by the agency of manufacture, use, or sale for human administration.
  • Example Clauses 1. A An epitope-scaffold molecule including an epitope having a sequence as set forth in at least one of SEQ ID NOs: 1-12 or a sequence having at least 98% sequence identity to the sequence as set forth in at least one of SEQ ID NOs.1-12; and a delivery scaffold. 2.
  • the epitope-scaffold molecule of clause 1 or 2 wherein the epitope includes the sequence as set forth in SEQ ID NO: 2 or a sequence having at least 98% sequence identity to the sequence as set forth in SEQ ID NO: 2.
  • the epitope-scaffold molecule of any one of clauses 1-7 wherein the epitope includes the sequence as set forth in SEQ ID NO: 7 or a sequence having at least 98% sequence identity to the sequence as set forth in SEQ ID NO: 7.
  • the epitope-scaffold molecule of any one of clauses 1-8 wherein the epitope includes the sequence as set forth in SEQ ID NO: 8 or a sequence having at least 98% sequence identity to the sequence as set forth in SEQ ID NO: 8.
  • 10. The epitope-scaffold molecule of any one of clauses 1-9 wherein the epitope includes the sequence as set forth in SEQ ID NO: 9 or a sequence having at least 98% sequence identity to the sequence as set forth in SEQ ID NO: 9. 11.
  • the scaffold protein includes a protein from T. pallidum, S. aureus, H. pylori, equine infectious anemia virus, or diphtheria toxin.
  • the scaffold protein includes a T. pallidum protein (Tp0751) from T. pallidum, a Protein A from S. aureus, a Cag-Z from H.
  • pylori a p26 capsid protein from equine infectious anemia virus, or a transmembrane domain of diphtheria toxin (DTT).
  • DTT diphtheria toxin 17.
  • the scaffold protein includes a synthetic protein.
  • the synthetic protein includes a small-synthetic ⁇ -barrel protein. 19.
  • 32. A concatenated epitope including at least two copies of a sequence selected from any of SEQ ID NOs: 1-12 or a sequence having at least 98% sequence identity to any of SEQ ID NOs: 1-12. 33.
  • the concatenated epitope includes: 2, 3, 4, 5, 6, 7, 8, 9, or more copies of SEQ ID NO: 1 or a sequence having at least 98% sequence identity to SEQ ID NO: 1; 2, 3, 4, 5, 6, 7, 8, 9, or more copies of SEQ ID NO: 2 or a sequence having at least 98% sequence identity to SEQ ID NO: 2; 2, 3, 4, 5, 6, 7, 8, 9, or more copies of SEQ ID NO: 3 or a sequence having at least 98% sequence identity to SEQ ID NO: 3; 2, 3, 4, 5, 6, 7, 8, 9, or more copies of SEQ ID NO: 4 or a sequence having at least 98% sequence identity to SEQ ID NO: 4; 2, 3, 4, 5, 6, 7, 8, 9, or more copies of SEQ ID NO: 5 or a sequence having at least 98% sequence identity to SEQ ID NO: 5; 2, 3, 4, 5, 6, 7, 8, 9, or more copies of SEQ ID NO: 6 or a sequence having at least 98% sequence identity to SEQ ID NO: 6; 2, 3, 4, 5, 6, 7, 8, 9, or more copies of SEQ ID NO: 6; 2, 3, 4,
  • a composition including: (i) an epitope including the sequence as set forth in at least one of SEQ ID NOs: 1-12 or a sequence having at least 98% sequence identity to the sequence as set forth in at least one of SEQ ID NOs: 1-12, an epitope-scaffold molecule including the sequence as set forth in at least one of SEQ ID NOs: 1-12 or a sequence having at least 98% sequence identity to the sequence as set forth in at least one of SEQ ID NOs: 1-12 and a delivery scaffold, or a concatenated epitope including the sequence as set forth in at least two of SEQ ID NOs: 1-12 or a sequence having at least 98% sequence identity to the sequence as set forth in of SEQ ID NOs: 1-12; and (ii) a pharmaceutically acceptable carrier.
  • composition of clause 36 further including an adjuvant.
  • adjuvant includes a squalene-based adjuvant, an aluminum- based adjuvant, a saponin-based adjuvant, a liposome-based adjuvant, a Toll-like receptor (TLR)-based adjuvant, a STING agonist, an attenuated lipid A derivative (ALD), or a carbomer-lecithin-based adjuvant.
  • the aluminum-based adjuvant includes aluminum hydroxide, aluminum phosphate, or potassium aluminum sulphate (alum).
  • TLR-based adjuvant includes resiquimod, muramyl dipeptide, synthetic diacylated lipoprotein FSL-1, glucopyranosyl lipid A (GLA).
  • GLA glucopyranosyl lipid A
  • the TLR-based adjuvant includes GLA formulated in stable emulsion (GLA SE).
  • GLA SE stable emulsion
  • ALD monophosphoryl lipid A
  • MLA-Alum MLA-Alum.
  • a syphilis vaccine including an epitope including a sequence as set forth in at least one of SEQ ID NOs: 1- 12 or a sequence having at least 98% sequence identity to at least one of SEQ ID NOs: 1-12.
  • Example 1 Diverse and conserveed Sequences of TprC and TprK Variants. Two antigens have been identified among T. pallidum surface-exposed proteins belonging to the Tpr family of virulence factors that confer substantial protection to syphilis in the rabbit model. Immunity to these antigens promoted pathogen clearance by opsonophagocytosis and attenuated chancre development at challenge sites compared to controls.
  • T. pallidum outer membrane hosts very few and a very low density of surface-exposed integral membrane proteins (Radolf et al., Proc Natl Acad Sci USA.1989, 86(6):2051-5). Finding these rare outer membrane proteins (OMPs) is compared to "finding the holy grail" in the field, as OMP identification would allow a deeper understanding of the host-pathogen interaction dynamics. Progress toward OMP identification has been hindered by limitations such as the inability to steadily culture this pathogen, even though this might change in the future.
  • T. pallidum can be propagated in rabbits by intratesticular passage every 10-12 days, low treponemal yields from infected animals (10 7 - 10 10 cells per animal, depending on the strain) and the presence of contaminating rabbit material make it difficult to evaluate the bacterial surface.
  • the uncommon fragility of T. pallidum outer membrane makes direct identification of OMPs very difficult using approaches such as surface immunofluorescence (Penn et al., J Gen Microbiol.1985, 131(Pt 9):2349-57; and Blanco et al., Emerg Infect Dis.1997, 3(1):11-20). Lack of genetic tools for this pathogen further limits the array of techniques applicable for OMP identification. Mining of the T.
  • T. pallidum repeat (Tpr) antigens that are the focus of this example are showing significant protection to the infection.
  • T. pallidum has eliminated genes encoding important metabolic activities (such as oxidative phosphorylation, synthesis of fatty acids, nucleotides, and most amino acids) (Norris and Sell, J Immunol. 1984, 133(5):2686-92), but has retained the tpr-encoding genes likely because they play a key role in T. pallidum biology and disease pathogenesis.
  • the members of the Tpr family are divided into three subfamilies according to sequence homology. Subfamily I member TprC and Subfamily III member TprK are the focus of this example. Additionally, as T.
  • pallidum has a very low-density of integral proteins in its outer membrane, evidence that the tprK, and tprC, genes are highly expressed in this pathogen (“Giacani 3”; Smajs et al., J Bacteriol. 2005, 187(5):1866-74) further supports their selection for vaccine design.
  • a three-dimensional model of both TprC generated with homology- based modeling programs such as I-TASSER (Yang et al., Nature methods.2015, 12(1):7-8), predict these proteins to be typical ⁇ -barrel outer membrane proteins (OMPs) with 11 external loops (ExL, FIG.1A). Relying on 3D models is still utilized because crystallization has not been accomplished for any Tpr protein to date.
  • FIG.2 shows a multiple sequence alignment of the COOH-terminal region (AA 405-600) of five variants of TprC.
  • TprC variants are the result of combinations of a small number of antigenically different "external loop" sequences.
  • Three TprC protein variants cover the whole spectrum of known loop variants in this protein ("Centurion- Lara 3”).
  • TprK is also a bona fide OMP of T.
  • TprK is the most highly expressed (“Giacani 3”) and most immunogenic of the Tprs, containing B- cell epitopes (“Giacani 3”; and Leader et al., Infect Immun. 2003, 71(10):6054-7). This protein is the T. pallidum to undergo intra- strain antigenic variation during the course of infection. [0160] Generation of diversity in TprK occurs through segmental gene conversion in seven discrete variable (V) regions named V1-V7 (FIG.3), that are separated from each other by conserved sequences (“Giacani 1”).
  • V discrete variable
  • TprK models support that this protein has a ⁇ -barrel structure with 11 surface-exposed loops, seven of which are predicted to harbor the variable regions (“Giacani 2”) (“V regions” in FIG. 3), while the remaining four, all located in the protein NH2- terminus, are conserved (FIG. 3).
  • the TprK conserved region is already known to contain targets for opsonizing antibodies and to be protective in immunization/challenge experiments.
  • the role of the TprK V region in allowing immune evasion during infection was repeatedly shown (“Giacani 2”; LaFond et al., Infect Immun.2006, 74(3):1896- 906; LaFond et al., J Bacteriol.
  • MPLA mono-phosphoryl lipid A
  • TDM synthetic trehalose dimycolate
  • CWS cell wall skeleton
  • adjuvants were tested in combination with the antigens representing the small conserved regions of TprK and TprC.
  • This list also included adjuvants for human use, such as alum, in addition to several custom-made emulsion-based adjuvants containing various combinations of immunomodulatory components such as glucopyranosyl lipid (GL), risiquimod, muramyl dipeptide (MDP, a NOD2 agonist), FSL-1, and Qui- A (a saponin adjuvant), as well as the commercially available Titermax Gold.
  • GL glucopyranosyl lipid
  • MDP muramyl dipeptide
  • FSL-1 FSL-1
  • Qui- A a saponin adjuvant
  • FIGs.4A, 4B shows that i) antigen-stimulated PBMCs from protected rabbits produced significantly higher IFN ⁇ levels compared to PBMCs from rabbits that received a "non-protective" adjuvant, and ii) that immunization with antigen+Ribi-like significantly reduced the number of ulcerating lesions at challenge sites (which is a correlate of protection).
  • Example 3 Evaluate protective ability of recombinant full-length TprC and TprK antigens to inform vaccine design. Antisera raised against the conserved TprK NH 2 -terminal region (AA 37-273 of, for example, SEQ ID NO: 40, predicted to contain four conserved surface-exposed loops) (“Giacani 2”), are opsonic for T.
  • TprK conserved fragment contains targets of opsonic antibodies, and immunity to these targets promotes early treponemal clearance. Similar results were obtained in opsonophagocytosis assays performed with immune sera to the NH 2 -terminal conserved region shared by TprC variants (AA 21-284 of, for example, SEQ ID NO: 41, containing four predicted surface loops) (FIG. 5). Furthermore, immunization with this TprC conserved peptide provided significant protection against infectious challenge. Upon challenge with the Nichols strain, for example, ulcerative chancres developed in 8% of the inoculation sites in immunized rabbits compared to 83% of the inoculation sites in controls (p ⁇ 0.01) (“Sun”).
  • Example 4 Evaluate the protective ability of chimeric concatemers based on protective B-cell epitopes of TprC and TprK. Epitopes for concatemer construction were selected and include those recognized by mAbs with opsonic and neutralizing ability.
  • TprC and TprK B-cell epitopes that fall within the predicted proteins loops and are seen by immune sera in animals exhibiting significant protection compared to controls were cloned.
  • the concatemer- encoding sequence was obtained by joining pre-synthesized DNA fragments by overlapping PCR, which was then cloned into the E. coli expression vector pET-32 to add a 5xHis-tag for purification. Concatemers were produced as codon-optimized genes for E. coli to maximize likelihood of expression.
  • Tpr antigens are relatively large hydrophobic proteins (60 kDa) that require extensive work to be purified and refolded into an antigen that is similar to its native counterpart.
  • Antigen solubility does not generally improve even when smaller portions of the whole proteins are expressed in E. coli, as hydrophobic regions alternate with hydrophilic ones throughout the primary sequence of these antigens.
  • sequences representing both hydrophilic surface loops and the hydrophobic ⁇ -barrel scaffolding some antibodies are inevitably elicited by portions of the proteins that are not surface exposed and therefore will not opsonize T. pallidum.
  • VLPs are supra-molecular protein structures that carry many of the characteristics of viruses, and that can be exploited in vaccine development strategies. As VLP-based vaccines lack a viral genome and have lost the ability to replicate, they constitute a very safe template for vaccine development. VLPs, including those derived from the HPV16 L1 protein, form a unique repetitive surface structure making them highly immunogenic vaccine templates.
  • VLP-based vaccines mainly target B- cells and induce potent antibody responses following activation of T helper cells and presentation by antigen presenting cells (APCs), making them an ideal vector for T. pallidum antigens. Additional advantages include a size (55 nm) that is optimal for drainage to the lymph nodes.
  • the size of the sequences that can be cloned into the L1 DE loop of chimeric HPV16 L1-based VLPs is limited to 15-20 amino acids in length.
  • B-cell epitope mapping using synthetic peptides representing the Nichols TprC COOH-region with antisera from long-term infected rabbits suggests that the sequences to be cloned do not exceed 20 amino acids (FIG.10).
  • Experimental procedures to obtain chimeric VLPs were performed using data acquired on B-cell epitope mapping for TprC and TprK.
  • plasmids were used to transfect 7x10 6 human embryonic kidney (HEK) cell line 293TT, previously modified to express high levels of SV40 LT antigen to drive L1 expression from the p16L1L2 vector. After 48 hours, cells were collected and lysed using detergent. VLPs were given time to mature by overnight incubation of the cell lysates at 37°C. VLPs were harvested by ultracentrifugation through an Optiprep (60% iodixanol) step gradient for 3.5 hours at 16°C at 234,000xg. L1 antibodies that bind to conformational epitopes were used to recognize properly folded L1. Presence of the T.
  • Example 6 Effect of immunization on lesion development in rabbits infected with the syphilis agent. For some of these experiments, a total of 60 ⁇ g of each antigen was given over the course of 5 immunizations (20 ⁇ g in the initial immunization and 10 ⁇ g in the boost injections). During each immunization, antigens were administered intramuscularly (400 ⁇ l), subcutaneously (400 ⁇ l distributed in four sites), and intradermally (100 ⁇ l in two sites).
  • the epitope-specificity of the Tpr monoclonal antibodies were determined using synthetic peptides, and the mAbs were tested for opsonic and neutralizing ability. As negative controls for these experiments, normal rabbit serum was used. Opsonization assays were conducted as previously described (Shaffer et al., Infect Immun.1993, 61(2):781-4). Briefly, after incubation of treponemes with control or test sera plus rabbit peritoneal macrophages, the un-ingested treponemes were washed off, and macrophages were fixed and stained for T. pallidum. An observer blinded to the test conditions deterined the percentage of macrophages containing ingested T. pallidum on triplicate slides per condition.
  • Macrophages incubated with treponemes + normal rabbit serum, and macrophages incubated without treponemes were the negative controls. Pooled sera from long term- infected rabbits serve as positive controls. Each antiserum was tested in triplicate per macrophage donor, with at least 3 macrophage donors. Neutralization was performed. Treponemes were incubated with heated control or test sera, with and without added active rabbit complement, under microaerophilic conditions for 16 hours, followed by intradermal inoculation onto the backs of normal rabbits. Animals were monitored for 1 month. Neutralization of treponemes is indicated by lack of lesion appearance at an inoculation site. Negative controls include NRS and all sera incubated in the absence of active complement.
  • Positive controls include infection-immune serum relevant to the target strain being used in the assay, incubated with active complement. Each condition is injected into two sites per rabbit, 3 rabbits per assay; 2 assays per test serum. The proportion of lesions appearing at injection sites for each condition was compared to the values for positive and negative controls. Induction of Th1-type immunity was assessed prior to challenge by measurement of IFN ⁇ release in PBMC incubated with the antigens. PBMC was purified by Ficoll-Hypaque and cultured for 8 and 24 hours with the antigens or ConA (positive control). IFN ⁇ release was measured using the LSBio Rabbit IFN ⁇ ELISA kit.
  • Biopsies were harvested from 2 challenge sites per rabbit at the time when lesions in the unimmunized control rabbits were on the brink of ulceration; these were processed for qPCR and reverse transcription (RT)-qPCR to determine total (DNA) and live (mRNA) T. pallidum burden, normalized to the message for the rabbit housekeeping gene hypoxanthine-guanine phosphoribosyltransferase (HPRT) (for RNA quantification) or rabbit cystic fibrosis transmembrane conductance regulator (CFTR) gene (for DNA quantification).
  • HPRT hypoxanthine-guanine phosphoribosyltransferase
  • CFTR cystic fibrosis transmembrane conductance regulator
  • Treponemal cell burden was estimated by counting bacterial cells under the microscope in lesion aspirates from rabbits immunized with TprK and TprC compared to control animals that were not immunized but were challenged with either the Nichols or SS14 strain. Treponemal cell burden was also estimated by real-time quantitative PCR in lesion aspirates from rabbits (FIG.12B) Rabbits immunized with TprK and TprC were observed to have significantly fewer treponemes in their lesions compared to controls infected with the Nichols strain (FIG.12A). Lesion ulceration was lower in rabbits immunized with The TprK and TprC antigens.
  • Immunization induced a T-cell response in animals as shown by the delayed-type hypersensitivity (DTH) at 48 hours after challenge for immunized rabbits (FIG.13).
  • Another way to assess vaccine efficacy is to harvest lymph nodes from vaccinated/challenge animals and inject them into one testis of a naive rabbit. If the na ⁇ ve rabbit develops syphilis more slowly compared to the control rabbits, it means that the immunized animals were protected. Animals that received lymph nodes from rabbits vaccinated with TprC (FIG. 14A) or TprK (FIG.14B) developed syphilis more slowly than controls. [0171] Example 7. Epitope Mapping.
  • nucleic acid and amino acid sequences provided herein are shown using letter abbreviations for nucleotide bases and amino acid residues, as defined in 37 C.F.R. ⁇ 1.831-1.835 and set forth in WIPO Standard ST.26 (implemented on July 1, 2022). Only one strand of each nucleic acid sequence is shown, but the complementary strand is understood as included in implementations where it would be appropriate. [0175] Variants of the sequences disclosed and referenced herein are also included. Guidance in determining which amino acid residues can be substituted, inserted, or deleted without abolishing biological activity can be found using computer programs well known in the art, such as DNASTARTM (Madison, Wisconsin) software.
  • amino acid changes in the protein variants disclosed herein are conservative amino acid changes, i.e., substitutions of similarly charged or uncharged amino acids.
  • a conservative amino acid change involves substitution of one of a family of amino acids which are related in their side chains.
  • suitable conservative substitutions of amino acids are known to those of skill in this art and generally can be made without altering a biological activity of a resulting molecule.
  • Those of skill in this art recognize that, in general, single amino acid substitutions in non-essential regions of a polypeptide do not substantially alter biological activity (see, e.g., Watson et al. Molecular Biology of the Gene, 4th Edition, 1987, The Benjamin/Cummings Pub. Co., p.
  • Naturally occurring amino acids are generally divided into conservative substitution families as follows: Group 1: Alanine (Ala), Glycine (Gly), Serine (Ser), and Threonine (Thr); Group 2: (acidic): Aspartic acid (Asp), and Glutamic acid (Glu); Group 3: (acidic; also classified as polar, negatively charged residues and their amides): Asparagine (Asn), Glutamine (Gln), Asp, and Glu; Group 4: Gln and Asn; Group 5: (basic; also classified as polar, positively charged residues): Arginine (Arg), Lysine (Lys), and Histidine (His); Group 6 (large aliphatic, nonpolar residues): Isoleucine (Ile), Leucine (Leu), Methionine (Met), Valine (Val) and Cysteine (Cys); Group 7 (uncharged polar): Tyrosine (Tyr), Gly, Asn, Gln, Cys,
  • substitution of amino acids whose hydropathic indices are within ⁇ 2 is preferred, those within ⁇ 1 are particularly preferred, and those within ⁇ 0.5 are even more particularly preferred. It is also understood in the art that the substitution of like amino acids can be made effectively on the basis of hydrophilicity.
  • amino acid substitutions may be based on the relative similarity of the amino acid side- chain substituents, for example, their hydrophobicity, hydrophilicity, charge, size, and the like.
  • variants of gene sequences can include codon optimized variants, sequence polymorphisms, splice variants, and/or mutations that do not affect the function of an encoded product to a statistically-significant degree.
  • Variants of the protein, nucleic acid, and gene sequences disclosed herein also include sequences with at least 70% sequence identity, 80% sequence identity, 85% sequence, 90% sequence identity, 95% sequence identity, 96% sequence identity, 97% sequence identity, 98% sequence identity, or 99% sequence identity to the protein, nucleic acid, or gene sequences disclosed herein.
  • “% sequence identity” refers to a relationship between two or more sequences, as determined by comparing the sequences. In the art, "identity” also means the degree of sequence relatedness between protein, nucleic acid, or gene sequences as determined by the match between strings of such sequences.
  • GCG Genetics Computer Group
  • BLASTP BLASTN
  • BLASTX Altschul, et al., J. Mol. Biol. 215:403-410 (1990); DNASTAR (DNASTAR, Inc., Madison, Wisconsin)
  • FASTA program incorporating the Smith-Waterman algorithm (Pearson, Comput. Methods Genome Res., [Proc. Int. Symp.] (1994), Meeting Date 1992, 111-20. Editor(s): Suhai, Sandor. Publisher: Plenum, New York, N.Y.
  • variants also include nucleic acid molecules that hybridize under stringent hybridization conditions to a sequence disclosed herein and provide the same function as the reference sequence.
  • Exemplary stringent hybridization conditions include an overnight incubation at 42 °C in a solution including 50% formamide, 5XSSC (750 mM NaCl, 75 mM trisodium citrate), 50 mM sodium phosphate (pH 7.6), 5XDenhardt's solution, 10% dextran sulfate, and 20 ⁇ g/ml denatured, sheared salmon sperm DNA, followed by washing the filters in 0.1XSSC at 50 °C. Changes in the stringency of hybridization and signal detection are primarily accomplished through the manipulation of formamide concentration (lower percentages of formamide result in lowered stringency); salt conditions, or temperature.
  • washes performed following stringent hybridization can be done at higher salt concentrations (e.g.5XSSC). Variations in the above conditions may be accomplished through the inclusion and/or substitution of alternate blocking reagents used to suppress background in hybridization experiments.
  • Typical blocking reagents include Denhardt's reagent, BLOTTO, heparin, denatured salmon sperm DNA, and commercially available proprietary formulations. The inclusion of specific blocking reagents may require modification of the hybridization conditions described above, due to problems with compatibility.
  • "Specifically binds" refers to an association of a binding domain (of, for example, a CAR binding domain) to its cognate binding molecule with an affinity or K a (i.e., an equilibrium association constant of a particular binding interaction with units of 1/M) equal to or greater than 10 5 M -1 , while not significantly associating with any other molecules or components in a relevant environment sample. Binding domains may be classified as "high affinity” or "low affinity”.
  • "high affinity" binding domains refer to those binding domains with a K a of at least 10 7 M- 1 , at least 10 8 M -1 , at least 10 9 M -1 , at least 10 10 M -1 , at least 10 11 M -1 , at least 10 12 M -1 , or at least 10 13 M -1 .
  • "low affinity" binding domains refer to those binding domains with a K a of up to 10 7 M -1 , up to 10 6 M- 1 , up to 10 5 M -1 .
  • affinity may be defined as an equilibrium dissociation constant (K d ) of binding interaction with units of M (e.g., 10 -5 M to 10 -13 M).
  • a binding domain may have "enhanced affinity," which refers to a selected or engineered binding domains with stronger binding to a cognate binding molecule than a wild type (or parent) binding domain.
  • enhanced affinity may be due to a K a (equilibrium association constant) for the cognate binding molecule that is higher than the reference binding domain or due to a K d (dissociation constant) for the cognate binding molecule that is less than that of the reference binding domain, or due to an off-rate (K off ) for the cognate binding molecule that is less than that of the reference binding domain.
  • each implementation disclosed herein can comprise, consist essentially of or consist of its particular stated element, step, or component.
  • the terms “include” or “including” should be interpreted to recite: “comprise, consist of, or consist essentially of.”
  • the transition term “comprise” or “comprises” means has, but is not limited to, and allows for the inclusion of unspecified elements, steps, ingredients, or components, even in major amounts.
  • the transitional phrase “consisting of” excludes any element, step, ingredient or component not specified.
  • the transition phrase “consisting essentially of” limits the scope of the implementation to the specified elements, steps, ingredients or components and to those that do not materially affect the implementation.
  • the term “about” has the meaning reasonably ascribed to it by a person skilled in the art when used in conjunction with a stated numerical value or range, i.e. denoting somewhat more or somewhat less than the stated value or range, to within a range of ⁇ 20% of the stated value; ⁇ 19% of the stated value; ⁇ 18% of the stated value; ⁇ 17% of the stated value; ⁇ 16% of the stated value; ⁇ 15% of the stated value; ⁇ 14% of the stated value; ⁇ 13% of the stated value; ⁇ 12% of the stated value; ⁇ 11% of the stated value; ⁇ 10% of the stated value; ⁇ 9% of the stated value; ⁇ 8% of the stated value; ⁇ 7% of the stated value; ⁇ 6% of the stated value; ⁇ 5% of the stated value; ⁇ 4% of the stated value; ⁇ 3% of the stated value; ⁇ 2% of the stated value; or ⁇ 1% of the stated value.

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Abstract

Des épitopes de lymphocytes B de Treponema pallidum et des vaccins contre la syphilis à base de ceux-ci sont décrits dans la présente invention. Les épitopes décrits dans la présente invention peuvent être concaténés ou présentés sur un échafaudage d'administration tel qu'une protéine d'échafaudage, une pseudo-particule virale ou une nanoparticule aux fins d'une administration en tant que vaccin. La présente divulgation propose également des procédés de stimulation d'une réponse immunitaire anti-syphilis chez un sujet à l'aide des épitopes et/ou du vaccin décrits dans la présente invention.
PCT/US2024/026293 2023-04-28 2024-04-25 Épitopes de lymphocytes b d'antigènes treponema pallidum destinés à être utilisés dans un vaccin contre la syphilis Pending WO2024226815A2 (fr)

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EP1071819A1 (fr) * 1998-04-10 2001-01-31 The University of Washington Proteines recombinees de treponeme pale et leur utilisation pour former un vaccin contre la syphilis
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US8343498B2 (en) * 2008-10-12 2013-01-01 Massachusetts Institute Of Technology Adjuvant incorporation in immunonanotherapeutics
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