WO2022251216A1 - Épitopes de lymphocytes t et compositions associées utiles dans la prévention, le diagnostic et le traitement de bêta-coronavirus - Google Patents

Épitopes de lymphocytes t et compositions associées utiles dans la prévention, le diagnostic et le traitement de bêta-coronavirus Download PDF

Info

Publication number
WO2022251216A1
WO2022251216A1 PCT/US2022/030726 US2022030726W WO2022251216A1 WO 2022251216 A1 WO2022251216 A1 WO 2022251216A1 US 2022030726 W US2022030726 W US 2022030726W WO 2022251216 A1 WO2022251216 A1 WO 2022251216A1
Authority
WO
WIPO (PCT)
Prior art keywords
beta
cov
coronavirus
sars
seq
Prior art date
Application number
PCT/US2022/030726
Other languages
English (en)
Inventor
Anne De Groot
William Martin
Original Assignee
Epivax, Inc.
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Epivax, Inc. filed Critical Epivax, Inc.
Publication of WO2022251216A1 publication Critical patent/WO2022251216A1/fr

Links

Classifications

    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • A61P31/12—Antivirals
    • A61P31/14—Antivirals for RNA 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
    • A61K39/12—Viral antigens
    • 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
    • A61K2039/54—Medicinal preparations containing antigens or antibodies characterised by the route of administration
    • 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
    • A61K2039/55561—CpG containing adjuvants; Oligonucleotide containing 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
    • A61K2039/70—Multivalent vaccine
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2770/00—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA ssRNA viruses positive-sense
    • C12N2770/00011—Details
    • C12N2770/20011—Coronaviridae
    • C12N2770/20022—New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2770/00—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA ssRNA viruses positive-sense
    • C12N2770/00011—Details
    • C12N2770/20011—Coronaviridae
    • C12N2770/20034—Use of virus or viral component as vaccine, e.g. live-attenuated or inactivated virus, VLP, viral protein

Definitions

  • the present disclosure generally relates to novel T-cell epitope-based compounds and compositions, including vaccines, effective against beta-coronavirus infection such as infection by Severe Acute Respiratory Syndrome (SARS), Middle East respiratory syndrome coronavirus (MERS-CoV), or Severe Acute Respiratory Syndrome Coronavirus 2 (SARS- CoV-2), and/or related diseases caused by such beta-coronaviruses, in a subject in need thereof.
  • SARS Severe Acute Respiratory Syndrome
  • MERS-CoV Middle East respiratory syndrome coronavirus
  • SARS- CoV-2 Severe Acute Respiratory Syndrome Coronavirus 2
  • T-cell epitope compounds and compositions include immunogenic T-cell epitope polypeptides (including concatemeric polypeptides and chimeric or fusion polypeptides), as well as nucleic acids, plasmids, vectors (including expression vectors), and cells that express the polypeptides, pharmaceutical compositions, and vaccines.
  • coronavirus disease 2019 (COVID-19) pandemic demonstrated that infectious diseases pose a serious threat to human health and the global economy.
  • safe and effective vaccines against the causative agent severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)
  • SARS-CoV-2 severe acute respiratory syndrome coronavirus 2
  • SARS-CoV-2 was the third highly pathogenic beta-coronavirus to emerge in humans in the first two decades of this century, novel beta- coronavirus outbreaks are likely in the future and thus, there is an urgent need for a proactive vaccine strategy that prevents another beta-coronavirus pandemic.
  • S spike glycoprotein on the SARS-CoV-2 virion surface
  • beta-coronaviruses use different host receptors for cell entry and their S proteins consequently exhibit variability in the receptor binding domain (RBD).
  • RBD receptor binding domain
  • S2 domains are relatively more conserved across beta- coronaviruses, cross-reactive antibodies that target S2 are only weakly neutralizing.
  • CD4+ and CD8+ effector T cell epitopes that can be used for protection against or treat ment of infection by beta-coronavirues generally and for their use in the development of effective pharmaceuticals and vaccines.
  • T cells play a critical role in protection against coronavirus disease.
  • the polypeptides, nucleic acids and other compounds and vaccines as provided herein leverage cellular immunity to reduce morbidity and mortality with induction of long-lasting memory cytolytic T lymphocytes and helper T cells that recognize sequences conserved across beta-coronaviruses and stand ready to kill virus-infected cells and support both cellular and humoral immunity upon exposure to a novel beta-coronavirus.
  • T cell targeting vaccine composed of conserved epitopes may provide effective, long-term coronavirus immunity.
  • a study of correlates of protection against SARS-CoV-2 infection in rhesus macaques showed that natural immune protection following recovery from infection was significantly reduced by depletion of CD8+ T cells in animals rechallenged with SARS-CoV-2.
  • This finding shows that cellular immunity may play a role in protection in the absence of protective antibody titers and suggests that vaccination does not need to generate a neutralizing antibody response to protect.
  • T cells protect against severe infection and re-infection in animal coronavirus models.
  • virus-specific CD8+ T cells protect against acute respiratory distress syndrome (ARDS).
  • ARDS acute respiratory distress syndrome
  • this disclosure provides a safe and effective pan beta- coronavirus T cell-directed peptides, nucleaic acids and other compunds that may be used as a treatment or prevention vaccine that stimulates robust, broad, and durable CD4+ and CD8+ T cells that recognize T cell epitopes shared widely by beta-coronaviruses.
  • novel, therapeutic pan beta-coronavirus T cell epitope compounds and compositions including one or more of e.g., peptides or polypeptides as disclosed herein, including polypeptides having an amino acid sequence comprising, consisting of, or consisting essentially of one or more of SEQ ID NOS: 4-1676, 1713-2595, 2605-2638, 2647-2718, and 2735-86921305, 1616, 1317, 1628, 1333, 1644, 1311, 1622, 1309, 1620, 1330, 1641, 1254, 1565, 1314, 1625, 1272, 1583, 1303, 1614, 1110, 1421, 1099, 1410,
  • MERS MERS, SARS, and/or SARS-CoV-2 infection and related diseases caused by MERS, SARS, and/or SARS-CoV-2
  • methods of treating and/or preventing against MERS, SARS, and/or SARS-CoV-2 infection and related diseases caused by MERS, SARS, and/or SARS-CoV-2 in a subject including MERS, SARS, and/or SARS-CoV-2 infection and related diseases caused by MERS, SARS, and/or SARS-CoV-2, and methods of treating and/or preventing against MERS, SARS, and/or SARS-CoV-2 infection and related diseases caused by MERS, SARS, and/or SARS-CoV-2 in a subject.
  • a T-cell epitope compound or composition of the present disclosure includes one or more peptides or polypeptides as disclosed herein.
  • the present disclosure is directed to a peptide or polypeptide having an amino acid sequence comprising, consisting of, or consisting essentially of one or more ofSEQ ID NOS: 1305, 1616, 1317, 1628, 1333, 1644, 1311, 1622, 1309, 1620, 1330, 1641, 1254, 1565, 1314, 1625, 1272, 1583, 1303, 1614, 1110,
  • phrases “consisting essentially of’ is intended to mean that a peptide or polypeptide according to the present disclosure, in addition to the sequence according to any of SEQ ID NOS: 1305, 1616, 1317, 1628, 1333, 1644, 1311, 1622, 1309, 1620, 1330, 1641, 1254, 1565, 1314, 1625, 1272, 1583, 1303, 1614, 1110, 1421, 1099, 1410, 1063, 1374, 1079,
  • polypeptides of the present disclosure may be isolated, synthetic, and/or recombinant, and may comprise post-transcriptional modifications such as glycosylation, added chemical groups, etc.
  • the peptides or polypeptides can be either in neutral (uncharged) or salt forms, and may be either free of or include modifications such as glycosylation, side chain oxidation, or phosphorylation.
  • the peptides or polypeptides of the instant disclosure can be capped with an N-terminal acetyl and/or C- terminal amino group.
  • peptides or polypeptides of the instant disclosure having SEQ ID NOS: 1305, 1616, 1317, 1628, 1333, 1644, 1311, 1622, 1309, 1620, 1330, 1641, 1254, 1565, 1314, 1625, 1272, 1583, 1303, 1614, 1110, 1421, 1099, 1410, 1063, 1374, 1079, 1390,
  • the instant disclosure is directed to a peptide or polypeptide comprising, consisting, or consisting essentially of an amino acid sequence of SEQ ID NOS: 1305, 1616,
  • the instant disclosure is directed to a peptide or polypeptide having a core amino acid sequence comprising, consisting of, or consisting essentially of one or more peptides or polypeptides having an amino acid sequence of SEQ ID NOS: 1305, 1616, 1317, 1628, 1333, 1644, 1311, 1622, 1309, 1620, 1330, 1641, 1254, 1565,
  • flanking amino acids can be distributed in any ratio to the C-terminus and the N-terminus (for example, all flanking amino acids can be added to one terminus, or
  • the instant disclosure is directed to a peptide or polypeptide having a core sequence comprising, consisting of, or consisting essentially of one or more peptides or polypeptides having an amino acid sequence of SEQ ID NOS: 1305, 1616, 1317, 1628, 1333, 1644, 1311, 1622, 1309, 1620, 1330, 1641, 1254, 1565, 1314, 1625, 1272, 1583, 1303, 1614, 1110, 1421, 1099, 1410, 1063, 1374, 1079, 1390, 1338, 1649, 1264, 1575, 1296, 1607, 1246, 1557, 1247, 1558, 1334, 1645, 1949, 2587, 1102, 1413, 1248, 1559, 1103, 1414, 1065, 1376, 1185, 1496, 1184, 1495, 1091, 1402, 1161, 1471, 1337, 1648, 1191, 1502, 1293, 1604, 1203, 1514, 1217, 1528,
  • said polypeptide with the flanking amino acids is still able to bind to a same HLA molecule (i.e., retain MHC binding propensity) and/or retain the same TCR specificity as said polypeptide core sequence without said flanking amino acids.
  • said polypeptide with the flanking amino acids is still able to bind to a same HLA molecule (i.e., retain MHC binding propensity) and/or retain the same TCR specificity, and/or retain anti-viral activity, including anti-beta-coronavirus activity, as said polypeptide core sequence without said flanking amino acids.
  • flanking amino acid sequences are those that also flank the peptides or polypeptides included therein in the naturally occurring protein from which the peptide or polypeptide is found.
  • a peptide or polypeptide having a core sequence comprising, consisting of, or consisting essentially of one or more peptides or polypeptides having an amino acid sequence of SEQ ID NOS: 1305, 1616, 1317, 1628, 1333, 1644, 1311, 1622, 1309, 1620, 1330, 1641, 1254, 1565, 1314, 1625, 1272, 1583, 1303, 1614,
  • the extensions of 1 to 12 amino acids are those found flanking the amino acid sequence of SEQ ID NOS: 1305, 1616, 1317, 1628, 1333, 1644, 1311, 1622, 1309, 1620, 1330, 1641, 1254, 1565, 1314, 1625, 1272, 1583, 1303, 1614, 1110, 1421, 1099,
  • flanking amino acid sequences as described herein may serve as a MHC stabilizing region.
  • the use of a longer peptide may allow endogenous processing by patient cells and may lead to more effective antigen presentation and induction of T cell responses.
  • the extension(s) may serve to improve the biochemical properties of the peptides or polypeptides (e.g., but not limited to, solubility or stability) or to improve the likelihood for efficient proteasomal processing of the peptide.
  • the polypeptides of the present disclosure may be islated, synthetic, and/or recombinant, and may comprise post-transcriptional modifications such as glycosylation, added chemical groups, etc.
  • the peptides or polypeptides can be either in neutral (uncharged) or salt forms, and may be either free of or include modifications such as glycosylation, side chain oxidation, or phosphorylation.
  • the peptides or polypeptides of the instant disclosure can be capped with an N- terminal acetyl and/or C-terminal amino group.
  • the present disclosure is directed to a concatemeric polypeptide or peptide that comprises two or more of the instantly-disclosed polypeptides or peptides (e.g., but not limited to, a peptide or polypeptide comprising, consisting, or consisting essentially of an amino acid sequence of SEQ ID NOS: 1305, 1616, 1317, 1628, 1333, 1644, 1311, 1622, 1309,
  • Such additional peptide or polypeptide may be one or more of the instantly instantly-disclosed polypeptides or peptides (optionally a peptide or polypeptide having the sequence of any one of SEQ ID NOS: 4-1676, 1713-2595, 2605-2638, 2647-2718, and 2735- 8692), or may be an additional peptide or polypeptide of interest.
  • a concatemeric peptide is composed of 3 or more, 4 or more, 5 or more, 6 or more, 7 or more, 8 or more, or 9 or more, of the instantly-disclosed peptides or polypeptides.
  • the concatemeric peptides or polypeptides include 1000 or more, 1000 or less, 900 or less, 500 or less, 100 or less, 75 or less, 50 or less, 40 or less, 30 or less, 20 or less or 10 or less peptide epitopes.
  • a concatemeric peptide has 3-100, 5-100, 10-100, 15-100, 20-100, 25-100, 30- 100, 35-100, 40-100, 45-100, 50-100, 55-100, 60-100, 65-100, 70-100, 75-100, 80-100, 90- 100, 5-50, 10-50, 15-50, 20-50, 25-50, 30-50, 35-50, 40-50, 45-50, 100-150, 100-200, 100- 300, 100-400, 100-500, 50-500, 50-800, 50-1000, or 100-1000 of the instantly-disclosed peptides or polypeptides linked, fused, or joined together.
  • Each peptide or polypeptide of the concatemeric polypeptide may optionally have one or more linkers, which may optionally be or include cleavage sensitive sites, adjacent to their N- and/or C-terminal end.
  • linkers may optionally be or include cleavage sensitive sites, adjacent to their N- and/or C-terminal end.
  • two or more of the peptide epitopes may have a cleavage sensitive site between them.
  • two or more of the peptide epitopes may be connected directly to one another or through a linker that is not a cleavage sensitive site.
  • a concatemeric polypeptide of the present disclosure comprises, consists of, or consists essentially of one or more of SEQ ID NOS: 1677-1692, 2593-2604, 2639-2646, and 2719-2734 and/or fragments and variants thereof.
  • the instantly-disclosed concatermeric polypepide or peptide sequences do not correspond to a naturally occurring sequence, i.e., each of the one or more of the instantly-disclosed polypeptides or peptides (e.g., but not limited to, a peptide or polypeptide comprising, consisting, or consisting essentially of an amino acid sequence of SEQ ID NOS: 1305, 1616, 1317, 1628, 1333, 1644, 1311, 1622, 1309, 1620, 1330, 1641, 1254, 1565, 1314, 1625, 1272, 1583, 1303, 1614, 1110, 1421, 1099, 1410, 1063, 1374, 1079, 1390,
  • 1415, 1239,1550, 1250, 1561, 1231, 1542, 1100, 1411, 1118, 1429, 1062, 1373, 1086, 1397, 1177, 1487) are linked, fused, or joined together (e.g., fused in-frame, chemically-linked, or otherwise bound) to an additional peptide or polypeptide (which may be one or more of the instantly-disclosed peptides) in such a fashion such that the overall concatermic polypeptide does not correspond to a naturally occurring coronavirus sequence.
  • the concatemeric polypeptides of the present disclosure may be isolated, synthetic, and/or recombinant, and may comprise post-transcriptional modifications such as glycosylation, added chemical groups, etc.
  • the concatemeric polypeptides can be either in neutral (uncharged) or salt forms, and may be either free of or include modifications such as glycosylation, side chain oxidation, or phosphorylation.
  • the concatemeric polypeptides of the instant disclosure can be capped with an N-terminal acetyl and/or C- terminal amino group.
  • one or more peptides or polypeptides or concatemeric polypeptides of the instant disclosure e.g., but not limited to, a peptide or polypeptide comprising, consisting, or consisting essentially of an amino acid sequence of SEQ ID NOS: SEQ ID NOS: 1305, 1616, 1317, 1628, 1333, 1644, 1311, 1622, 1309, 1620, 1330, 1641, 1254, 1565, 1314, 1625, 1272,
  • the one or more peptides or polypeptides or concatemeric polypeptides of the instant disclosure may be joined to, linked to (e.g., fused in- frame, chemically-linked, or otherwise bound), and/or inserted into a heterologous polypeptide as a whole, although it may be made up from a joined to, linked to (e.g., fused in-frame, chemically-linked, or otherwise bound), and/or inserted amino acid sequence, together with flanking amino acids of the heterologous polypeptide.
  • the present disclosure is directed to a chimeric or fusion polypeptide composition (which in aspects may be isolated, synthetic, or recombinant) comprising one or more peptides, polypeptides, or concatemeric peptides of the present disclosure.
  • a chimeric or fusion polypeptide composition of the present disclosure comprises one or more peptides, polypeptides, and/or concatemeric peptides of the present disclosure (e.g., a peptide or polypeptide comprising, consisting of, or consisting essentially of one or more peptides or polypeptides having an amino acid sequence of SEQ ID NOS: 1305, 1616, 1317, 1628, 1333, 1644, 1311, 1622, 1309, 1620, 1330, 1641, 1254, 1565, 1314, 1625, 1272, 1583, 1303, 1614,
  • the one or more peptides, polypeptides, and/or concatemeric peptides of the present disclosure may be inserted into the heterologous polypeptide, may be added to the C-terminus (with or without the use of linkers, as is known in the art), and/or added to the N-terminus (with or without the use of linkers, as is known in the art) of the heterologous polypeptide.
  • the one or more peptide, polypeptides, or concatemeric peptides may be joined to, linked to (e.g., fused in-frame, chemically-linked, or otherwise bound), and/or inserted into a heterologous polypeptide as a whole, although it may be made up from joined to, linked to (e.g., fused in-frame, chemically-linked, or otherwise bound), and/or inserted amino acid sequence, together with flanking amino acids of the heterologous polypeptide.
  • a chimeric or fusion polypeptide composition of the present disclosure comprises a peptide, polypeptide, and/or concatemeric peptide of the instant disclosure, said peptide, polypeptide, and/or concatemeric peptide having a sequence that is not naturally included in the heterologous polypeptide and/or is not located at its natural position in the heterologous polypeptide.
  • the one or more peptides, polypeptides, and/or concatemeric peptides of the present disclosure may be inserted into a beta-coronavirus sequence in which the beta-coronavirus sequence does not include the one or more peptides, polypeptides, and/or concatemeric peptides of the present disclosure (e.g., the SARS-CoV-2 sequence is mutated to not include the one or more peptides, polypeptides, and/or concatemeric peptides of the present disclosure) or the one or more peptides, polypeptides, and/or concatemeric peptides of the present disclosure is inserted into a beta-coronavirus sequence but not at its natural position.
  • the beta-coronavirus sequence does not include the one or more peptides, polypeptides, and/or concatemeric peptides of the present disclosure (e.g., the SARS-CoV-2 sequence is mutated to not include the one or more peptides, poly
  • the one or more of peptide, polypeptide, and/or concatemeric peptide of the present disclosure can be joined, linked to (e.g., fused in- frame, chemically-linked, or otherwise bound), and/or inserted into the heterologous polypeptide.
  • the chimeric or fusion polypeptides may be isolated, synthetic, or recombinant.
  • the instant disclosure is directed to a nucleic acid (e.g., DNA or RNA, including mRNA) encoding one or more peptides, polypeptides, concatemeric peptides, and/or chimeric or fusion polypeptides as described herein.
  • a nucleic acid e.g., DNA or RNA, including mRNA
  • the instant disclosure is directed to a nucleic acid encoding a peptide or polypeptide comprising, consisting of, or consisting essentially of one or more peptides or polypeptides having an amino acid sequence ofSEQ IDNOS: 1305, 1616, 1317, 1628, 1333, 1644, 1311, 1622, 1309, 1620, 1330, 1641, 1254, 1565, 1314, 1625, 1272, 1583, 1303, 1614, 1110, 1421, 1099, 1410, 1063, 1374,
  • the present disclosure is directed to a vector, such as an expression vector, comprising such a nucleic acid as described.
  • the present disclosure is directed to expression cassettes, plasmids, expression vectors, recombinant viruses, or cells comprising a nucleic acid as described herein.
  • the present disclosure is directed to a cell or vaccine comprising such a vector as described.
  • the present disclosure is directed to a cell comprising a vector of the present disclosure.
  • the instant disclosure is directed to a pharmaceutical composition, the pharmaceutical composition comprising a T-cell epitope compound or composition of the instant disclosure (e.g., one or more of: polypeptides as disclosed herein; concatemeric peptides as disclosed herein; chimeric or fusion polypeptide compositions as disclosed herein; nucleic acids as disclosed herein, including nucleic acids encoding such peptides, polypeptides, concatemeric peptides, or chimeric of fusion polypeptide compositions as disclosed herein; expression cassettes; plasmids; expression vectors; recombinant viruses; or cells as disclosed herein) and a pharmaceutically acceptable carrier, excipient, and/or adjuvant.
  • a T-cell epitope compound or composition of the instant disclosure e.g., one or more of: polypeptides as disclosed herein; concatemeric peptides as disclosed herein; chimeric or fusion polypeptide compositions as disclosed herein; nucleic acids as disclosed herein, including
  • the one or more nucleic acids encoding said peptides or polypeptides are DNA, RNA, or mRNA.
  • the composition comprises at least 2, at least 3, at least 4, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 25, at least 30, at least 40, at least 50, at least 100, at least 200, at least 300, at least 400, at least 500, at least 600, at least 700, at least 800, at least 900, or at least 1000, peptides, polypeptides, and/or concatemeric peptides, as disclosed herein, including every value or range therebetween.
  • the instant disclosure is directed to a vaccine comprising a T-cell epitope compound or composition of the instant disclosure (e.g., one or more of: polypeptides as disclosed herein; concatemeric peptides as disclosed herein; chimeric or fusion polypeptide compositions as disclosed herein; nucleic acids as disclosed herein, including nucleic acids encoding such peptides, polypeptides, concatemeric peptides, or chimeric of fusion polypeptide compositions as disclosed herein; expression cassettes; plasmids; expression vectors; recombinant viruses; or cells as disclosed herein; pharmaceutical compositions as disclosed herein; or vaccines as described herein) and, optionally, a carrier, excipient, and/or an adjuvant.
  • a T-cell epitope compound or composition of the instant disclosure e.g., one or more of: polypeptides as disclosed herein; concatemeric peptides as disclosed herein; chimeric or fusion polypeptide compositions as disclosed
  • the present disclosure also relates to methods of immunizing or inducing an immune response, optionally a T cell response, in a subject, said method comprising administering to said subject one more peptides, polypeptides, concatemeric peptides, chimeric or fusion polypeptides, nucleic acids, expression cassettes, plasmids, expression vectors, recombinant viruses, cells pharmaceutical compositions, or vaccines as described herein.
  • the subject is a human.
  • the present disclosure is directed to to methods of immunizing or inducing an immune response in a subject, comprising administering to said subject a T-cell epitope compound or composition of the instant disclosure (e.g., one or more of: polypeptides as disclosed herein; concatemeric peptides as disclosed herein; chimeric or fusion polypeptide compositions as disclosed herein; nucleic acids as disclosed herein, including nucleic acids encoding such peptides, polypeptides, concatemeric peptides, or chimeric or fusion polypeptide compositions as disclosed herein; expression cassettes, plasmids; expression vectors; recombinant viruses; cells as disclosed herein; pharmaceutical compositions as disclosed herein; or vaccines as described herein).
  • a T-cell epitope compound or composition of the instant disclosure e.g., one or more of: polypeptides as disclosed herein; concatemeric peptides as disclosed herein; chimeric or fusion polypeptide compositions as disclosed herein
  • the subject is a human.
  • the present disclosure is directed to a method of stimulating, inducing, and/or expanding an immune response to a beta-coronavirus infection and/or related diseases caused by a beta- coronavirus (e.g. MERS, SARS, COVID-19) in a subject, comprising administering to said subject a T-cell epitope compound or composition of the instant disclosure.
  • a beta-coronavirus infection and/or related diseases caused by a beta- coronavirus e.g. MERS, SARS, COVID-19
  • the present disclosure also relates to methods of treating and/or preventing a beta- coronavirus infection and/or related diseases caused by a beta-coronavirus, including MERS, SARS, and COVID-19, in a subject, such as a human, comprising administering to said subject a T-cell epitope compound or composition of the instant disclosure (e.g., one or more of: polypeptides as disclosed herein; concatemeric peptides as disclosed herein; chimeric or fusion polypeptide compositions as disclosed herein; nucleic acids as disclosed herein, including nucleic acids encoding such peptides, polypeptides, concatemeric peptides, or chimeric or fusion polypeptide compositions as disclosed herein; expression cassettes; plasmids; expression vectors; recombinant viruses; cells as disclosed herein; pharmaceutical compositions as disclosed herein; or vaccines as described herein).
  • T cell epitope compounds or compositions of the instant disclosure as described herein may be used to induce an immune response and/or to vaccinate a subject. It is particularly useful to vaccinate against any beta-coronavirus infection and/or related diseases caused by any beta-coronavirus.
  • the T cell epitope compounds or compositions of the instant disclosure serve as a pan beta-coronavirus preventative, treatment, or composition for the amelioration of any symptoms of infection by any beta-coronavirus. While it is recognized that much of the present disclosure is directed to SARS-CoV-2 as an exemplary beta-coronavirus, the T cell epitope compounds or compositions are useful against all beta-coronaviruses, optionally as a pan beta-coronavirus vaccine or treatment.
  • FIG. 1 is an overview of MHC class II cluster selection from the envelope (SEQ ID NO: 1) of SARS-CoV-2.
  • the cluster address gives the location of the peptide within the sequences that were provided for analysis.
  • the core peptide (middle amino acids in bold, SEQ ID NO: in parentheses) defines the actual cluster that was identified during the analysis.
  • the stabilizing flanks (N-terminal and C-terminal, not bold) are included for use with the core sequence.
  • the number of hits is the number of EpiMatrix Z-scores above 1.64 or top 5% found within the sequence.
  • the EpiMatrix Cluster Score is derived from the number of hits normalized for the length of the cluster. Cluster Score is thus the excess or shortfall in predicted aggregate immunogenicity relative to a random peptide standard. Hydrophobicity scores of 2 and above are predictive of difficulty synthesizing peptides.
  • FIG. 2 is an EpiMatrix Cluster detail report for identified MHC class II clusters of the envelope (SEQ ID NO: 1) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”.
  • FIG. 3 is an EpiMatrix Cluster detail report for identified MHC class II clusters of the envelope (SEQ ID NO: 1) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”.
  • FIG. 4 is an EpiMatrix Cluster detail report for identified MHC class II clusters of the envelope (SEQ ID NO: 1) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”.
  • FIG. 5 is an overview of MHC class II cluster selection from the membrane (SEQ ID NO: 2) of SARS-CoV-2.
  • the cluster address gives the location of the peptide within the sequences that were provided for analysis.
  • the core peptide (middle amino acids having the sequence of the SEQ ID NO: in parentheses) defines the actual cluster that was identified during the analysis.
  • the stabilizing flanks (N-terminal and C-terminal) are included for use with the core sequence.
  • the number of hits is the number of EpiMatrix Z-scores above 1.64 or top 5% found within the sequence.
  • the EpiMatrix Cluster Score is derived from the number of hits normalized for the length of the cluster. Cluster Score is thus the excess or shortfall in predicted aggregate immunogenicity relative to a random peptide standard. Hydrophobicity scores of 2 and above are predictive of difficulty synthesizing peptides.
  • FIG. 6 is EpiMatrix Cluster detail reports for identified MHC class II clusters of the membrane (SEQ ID NO: 2) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”.
  • FIG. 7 is EpiMatrix Cluster detail reports for identified MHC class II clusters of the membrane (SEQ ID NO: 2) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”.
  • FIG. 8 is EpiMatrix Cluster detail reports for identified MHC class II clusters of the membrane (SEQ ID NO: 2) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”.
  • FIG. 9 is EpiMatrix Cluster detail reports for identified MHC class II clusters of the membrane (SEQ ID NO: 2) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”.
  • FIG. 10 is EpiMatrix Cluster detail reports for identified MHC class II clusters of the membrane (SEQ ID NO: 2) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”.
  • FIG. 11 is EpiMatrix Cluster detail reports for identified MHC class II clusters of the membrane (SEQ ID NO: 2) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”.
  • FIG. 12 is EpiMatrix Cluster detail reports for identified MHC class II clusters of the membrane (SEQ ID NO: 2) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”.
  • FIG. 13 is EpiMatrix Cluster detail reports for identified MHC class II clusters of the membrane (SEQ ID NO: 2) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”.
  • FIG. 14 is EpiMatrix Cluster detail reports for identified MHC class II clusters of the membrane (SEQ ID NO: 2) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”.
  • FIG. 15 is EpiMatrix Cluster detail reports for identified MHC class II clusters of the membrane (SEQ ID NO: 2) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”.
  • FIG. 16 is an overview of MHC class II cluster selection from the spike (SEQ ID NO: 3) of SARS-CoV-2.
  • the cluster address gives the location of the peptide within the sequences that were provided for analysis.
  • the core peptide (middle amino acids having the sequence of the SEQ ID NO: in parentheses) defines the actual cluster that was identified during the analysis.
  • the stabilizing flanks (N-terminal and C-terminal) are included for use with the core sequence.
  • the number of hits is the number of EpiMatrix Z-scores above 1.64 or top 5% found within the sequence.
  • the EpiMatrix Cluster Score is derived from the number of hits normalized for the length of the cluster. Cluster Score is thus the excess or shortfall in predicted aggregate immunogenicity relative to a random peptide standard. Hydrophobicity scores of 2 and above are predictive of difficulty synthesizing peptides
  • FIG. 17 is EpiMatrix Cluster detail reports for identified MHC class II clusters of the spike (SEQ ID NO: 3) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. *Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 18 is EpiMatrix Cluster detail reports for identified MHC class II clusters of the spike (SEQ ID NO: 3) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. *Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 19 is EpiMatrix Cluster detail reports for identified MHC class II clusters of the spike (SEQ ID NO: 3) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. *Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined. FIG.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. *Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 21 is EpiMatrix Cluster detail reports for identified MHC class II clusters of the spike (SEQ ID NO: 3) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. *Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 22 is EpiMatrix Cluster detail reports for identified MHC class II clusters of the spike (SEQ ID NO: 3) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. *Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 23 is EpiMatrix Cluster detail reports for identified MHC class II clusters of the spike (SEQ ID NO: 3) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. *Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 24 is EpiMatrix Cluster detail reports for identified MHC class II clusters of the spike (SEQ ID NO: 3) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. *Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 25 is EpiMatrix Cluster detail reports for identified MHC class II clusters of the spike (SEQ ID NO: 3) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. *Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 26 is EpiMatrix Cluster detail reports for identified MHC class II clusters of the spike (SEQ ID NO: 3) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. *Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 27 is EpiMatrix Cluster detail reports for identified MHC class II clusters of the spike (SEQ ID NO: 3) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. *Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 28 is EpiMatrix Cluster detail reports for identified MHC class II clusters of the spike (SEQ ID NO: 3) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. *Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 29 is EpiMatrix Cluster detail reports for identified MHC class II clusters of the spike (SEQ ID NO: 3) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. *Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 30 is EpiMatrix Cluster detail reports for identified MHC class II clusters of the spike (SEQ ID NO: 3) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. *Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 31 is EpiMatrix Cluster detail reports for identified MHC class II clusters of the spike (SEQ ID NO: 3) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. *Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 32 is EpiMatrix Cluster detail reports for identified MHC class II clusters of the spike (SEQ ID NO: 3) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. *Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 33 is EpiMatrix Cluster detail reports for identified MHC class II clusters of the spike (SEQ ID NO: 3) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. *Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined. FIG.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. *Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 35 is EpiMatrix Cluster detail reports for identified MHC class II clusters of the spike (SEQ ID NO: 3) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. *Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 36 is EpiMatrix Cluster detail reports for identified MHC class II clusters of the spike (SEQ ID NO: 3) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. *Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 37 is EpiMatrix Cluster detail reports for identified MHC class II clusters of the spike (SEQ ID NO: 3) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. *Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 38 is EpiMatrix Cluster detail reports for identified MHC class II clusters of the spike (SEQ ID NO: 3) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. *Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 39 is EpiMatrix Cluster detail reports for identified MHC class II clusters of the spike (SEQ ID NO: 3) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. *Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 40 is EpiMatrix Cluster detail reports for identified MHC class II clusters of the spike (SEQ ID NO: 3) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. *Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 41 is EpiMatrix Cluster detail reports for identified MHC class II clusters of the spike (SEQ ID NO: 3) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. *Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 42 is EpiMatrix Cluster detail reports for identified MHC class II clusters of the spike (SEQ ID NO: 3) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. *Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 43 is EpiMatrix Cluster detail reports for identified MHC class II clusters of the spike (SEQ ID NO: 3) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. *Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 44 is EpiMatrix Cluster detail reports for identified MHC class II clusters of the spike (SEQ ID NO: 3) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. *Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 45 is EpiMatrix Cluster detail reports for identified MHC class II clusters of the spike (SEQ ID NO: 3) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. *Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 46 is EpiMatrix Cluster detail reports for identified MHC class II clusters of the spike (SEQ ID NO: 3) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. *Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 47 is EpiMatrix Cluster detail reports for identified MHC class II clusters of the spike (SEQ ID NO: 3) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. *Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined. FIG.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. *Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 49 is EpiMatrix Cluster detail reports for identified MHC class II clusters of the spike (SEQ ID NO: 3) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. *Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 50 is EpiMatrix Cluster detail reports for identified MHC class II clusters of the spike (SEQ ID NO: 3) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. *Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 51 is EpiMatrix Cluster detail reports for identified MHC class II clusters of the spike (SEQ ID NO: 3) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. *Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 52 is EpiMatrix Cluster detail reports for identified MHC class II clusters of the spike (SEQ ID NO: 3) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. *Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 53 is EpiMatrix Cluster detail reports for identified MHC class II clusters of the spike (SEQ ID NO: 3) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. *Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 54 is EpiMatrix Cluster detail reports for identified MHC class II clusters of the spike (SEQ ID NO: 3) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. *Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 55 is EpiMatrix Cluster detail reports for identified MHC class II clusters of the spike (SEQ ID NO: 3) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. *Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 56 is the JanusMatrix reports for identified MHC class II clusters of the envelope (SEQ ID NO: 1) of SARS-CoV-2.
  • the number of HUMAN matches is the count of HUMAN JanusMatrix matches found in the search database.
  • a Janus Matrix match is a 9-mer derived from the search database (e.g., the human genome) which is predicted to bind to the same allele as the EpiMatrix Hit and shares TCR facing contacts with the EpiMatrix Hit.
  • the Janus Homology Score represents the average depth of coverage in the search database for each EpiMatrix hit in the input sequence. For example, an input peptide with eight EpiMatrix hits, all of which have one match in the search database, has a Janus Homology Score of 1. An input peptide with four EpiMatrix Hits, all of which have two matches in the search database, has a Janus Homology Score of 2. The JanusMatrix Homology Score considers all constituent 9-mers in any given peptide, including flanks.
  • FIG. 57 is the JanusMatrix reports for identified MHC class II clusters of the membrane (SEQ ID NO: 2) of SARS-CoV-2.
  • the number of HUMAN matches is the count of HUMAN JanusMatrix matches found in the search database.
  • a Janus Matrix match is a 9-mer derived from the search database (e.g., the human genome) which is predicted to bind to the same allele as the EpiMatrix Hit and shares TCR facing contacts with the EpiMatrix Hit.
  • the Janus Homology Score represents the average depth of coverage in the search database for each EpiMatrix hit in the input sequence. For example, an input peptide with eight EpiMatrix hits, all of which have one match in the search database, has a Janus Homology Score of 1. An input peptide with four EpiMatrix Hits, all of which have two matches in the search database, has a Janus Homology Score of 2. The JanusMatrix Homology Score considers all constituent 9-mers in any given peptide, including flanks.
  • FIG. 58 is the JanusMatrix reports for identified MHC class II clusters of the spike (SEQ ID NO: 3) of SARS-CoV-2.
  • the number of HUMAN matches is the count of HUMAN JanusMatrix matches found in the search database.
  • a Janus Matrix match is a 9-mer derived from the search database (e.g., the human genome) which is predicted to bind to the same allele as the EpiMatrix Hit and shares TCR facing contacts with the EpiMatrix Hit.
  • the Janus Homology Score represents the average depth of coverage in the search database for each EpiMatrix hit in the input sequence. For example, an input peptide with eight EpiMatrix hits, all of which have one match in the search database, has a Janus Homology Score of 1. An input peptide with four EpiMatrix Hits, all of which have two matches in the search database, has a Janus Homology Score of 2. The JanusMatrix Homology Score considers all constituent 9-mers in any given peptide, including flanks.
  • FIG. 59 is an EpiMatrix staircase report for identified MHC class I clusters of the envelope (SEQ ID NO: 1) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer or 10- mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figure. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”.
  • FIG. 60 is an EpiMatrix staircase report for identified MHC class I clusters of the membrane (SEQ ID NO: 2) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer or 10- mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figure. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”.
  • FIG. 61 is an EpiMatrix staircase report for identified MHC class I clusters of the spike (SEQ ID NO: 3) of SARS-CoV-2.
  • Z-score indicates the potential of a 9-mer or 10-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figure. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”.
  • FIG. 62 shows the sequences of the envelope (SEQ ID NO: 1) of SARS-CoV-2, the membrane (SEQ ID NO: 2) of SARS-CoV-2, and the spike (SEQ ID NO: 3) of SARS-CoV-2.
  • FIG. 63A is ex vivo immune recall responses differentiate SARS-CoV-2 naive individuals.
  • FIG. 63B is ex vivo immune recall responses differentiate SARS-CoV-2 experienced individuals and exhibit different COVID-19 immunotypes.
  • FIG. 64A and FIG. 64B shows that strong ex vivo immune recall responses are found or may be found in SARS-CoV-2 experienced individuals using polypeptides of the instant disclosure.
  • FIG. 64B shows that strong ex vivo immune recall responses are found or may be found in SARS-CoV-2 experienced individuals using polypeptides of the instant disclosure.
  • FIG. 65 shows polypeptides of the instant disclosure stimulate ex vivo immune recall response in natural SARS-CoV-2 infection.
  • FIG. 66A show polypeptides of the instant disclosure stimulate or may stimulate higher IFN-g responses in naive donors following expansion in culture.
  • FIG. 66B show polypeptides of the instant disclosure stimulate or may stimulate higher IFN-g responses in COVID-19 convalescent donors following expansion in culture.
  • FIG. 67A shows polypeptides of the instant disclosure stimulate or may stimulate low frequency epitope-specific T cells following expansion in culture in naive donors.
  • FIG. 67B shows polypeptides of the instant disclosure stimulate or may stimulate low frequency epitope-specific T cells following expansion in culture in COVID-19 convalescent donors.
  • FIG. 68 shows polypeptides of the instant disclosure stimulate low frequency epitope- specific T cells following expansion in culture in naive and COVID-19 convalescent donors.
  • FIGS. 69 shows the sequences of the nucleocapsid (SEQ ID NO: 1693), ORF3a (SEQ ID NO: 1694), ORF6 (SEQ ID NO: 1695), ORF7a (SEQ ID NO: 1696), ORF8 (SEQ ID NO: 1697), ORFIO (SEQ ID NO: 1698), ORFlab non-structural protein 2 (NSP2) (SEQ ID NO: 1699), ORFlab non-structural protein 3 (NSP3) (SEQ ID NO: 1700), ORFlab non-structural protein 4 (NSP4) (SEQ ID NO: 1701), ORFlab 3C-like proteinase (SEQ ID NO: 1702), ORFlab non-structural protein 6 (NSP6) (SEQ ID NO: 1703), ORFlab non-structural protein 7 (NSP7) (SEQ ID NO: 1704), ORFlab non-structural protein 8 (NSP8) (SEQ ID NO: 1705), ORFlab non-structural protein 9 (NSP9)
  • FIG. 70 is an overview of MHC class II cluster selection from the various proteins of SARS-CoV-2 with the corresponding SEQ ID NO: in parentheses.
  • the cluster address given the location of the peptide within the sequences that were provided for analysis.
  • the core peptide (SEQ ID NO: in parentheses) defines the actual cluster that was identified during the analysis.
  • the stabilizing flanks (N-terminal and C-terminal) are included for use with the core sequence, and are labeled by the SEQ ID NO: not listed in parentheses.
  • the number of hits is the number of EpiMatrix Z-scores above 1.64 or top 5% found within the sequence.
  • the EpiMatrix Cluster Score is derived from the number of hits normalized for the length of the cluster. Cluster Score is thus the excess or shortfall in predicted aggregate immunogenicity relative to a random peptide standard. Hydrophobicity scores of 2 and above are predictive of difficulty synthesizing peptides.
  • FIG. 71 are EpiMatrix Cluster detail reports for identified MHC class II clusters of exemplary SARS-CoV-2 peptides of FIG. 70.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 72 are EpiMatrix Cluster detail reports for identified MHC class II clusters of exemplary SARS-CoV-2 peptides of FIG. 70.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 73 are EpiMatrix Cluster detail reports for identified MHC class II clusters of exemplary SARS-CoV-2 peptides of FIG. 70.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 74 are EpiMatrix Cluster detail reports for identified MHC class II clusters of exemplary SARS-CoV-2 peptides of FIG. 70.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 75 are EpiMatrix Cluster detail reports for identified MHC class II clusters of exemplary SARS-CoV-2 peptides of FIG. 70.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 76 are EpiMatrix Cluster detail reports for identified MHC class II clusters of exemplary SARS-CoV-2 peptides of FIG. 70.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 77 are EpiMatrix Cluster detail reports for identified MHC class II clusters of exemplary SARS-CoV-2 peptides of FIG. 70.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 78 are EpiMatrix Cluster detail reports for identified MHC class II clusters of exemplary SARS-CoV-2 peptides of FIG. 70.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 79 are EpiMatrix Cluster detail reports for identified MHC class II clusters of exemplary SARS-CoV-2 peptides of FIG. 70.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 80 are EpiMatrix Cluster detail reports for identified MHC class II clusters of exemplary SARS-CoV-2 peptides of FIG. 70.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 81 are EpiMatrix Cluster detail reports for identified MHC class II clusters of exemplary SARS-CoV-2 peptides of FIG. 70.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined. FIG.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HFA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 83 are EpiMatrix Cluster detail reports for identified MHC class II clusters of exemplary SARS-CoV-2 peptides of FIG. 70.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HFA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HFA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 84 are EpiMatrix Cluster detail reports for identified MHC class II clusters of exemplary SARS-CoV-2 peptides of FIG. 70.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HFA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HFA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 85 are EpiMatrix Cluster detail reports for identified MHC class II clusters of exemplary SARS-CoV-2 peptides of FIG. 70.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HFA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HFA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 86 are EpiMatrix Cluster detail reports for identified MHC class II clusters of exemplary SARS-CoV-2 peptides of FIG. 70.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HFA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 87 are EpiMatrix Cluster detail reports for identified MHC class II clusters of exemplary SARS-CoV-2 peptides of FIG. 70.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 88 are EpiMatrix Cluster detail reports for identified MHC class II clusters of exemplary SARS-CoV-2 peptides of FIG. 70.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 89 are EpiMatrix Cluster detail reports for identified MHC class II clusters of exemplary SARS-CoV-2 peptides of FIG. 70.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 90 are EpiMatrix Cluster detail reports for identified MHC class II clusters of exemplary SARS-CoV-2 peptides of FIG. 70.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 91 are EpiMatrix Cluster detail reports for identified MHC class II clusters of exemplary SARS-CoV-2 peptides of FIG. 70.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 92 are EpiMatrix Cluster detail reports for identified MHC class II clusters of exemplary SARS-CoV-2 peptides of FIG. 70.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figures. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”. Scores in the top 10% are considered elevated. Frames containing 4 or more alleles scoring above 1.64 are referred to as EpiBars. These frames have an increased likelihood of binding to HLA. Flanking amino acids, added to stabilize the cluster during in vitro testing, are underlined.
  • FIG. 93 are EpiMatrix staircase reports for identified MHC class I clusters of exemplary SARS-CoV-2 peptides of FIG. 70.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figure. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”.
  • FIG. 94 are EpiMatrix staircase reports for identified MHC class I clusters of exemplary SARS-CoV-2 peptides of FIG. 70.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figure. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”.
  • FIG. 95 are EpiMatrix staircase reports for identified MHC class I clusters of exemplary SARS-CoV-2 peptides of FIG. 70.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figure. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”.
  • FIG. 96 are EpiMatrix staircase reports for identified MHC class I clusters of exemplary SARS-CoV-2 peptides of FIG. 70.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figure. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”.
  • FIG. 97 are EpiMatrix staircase reports for identified MHC class I clusters of exemplary SARS-CoV-2 peptides of FIG. 70.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figure. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”.
  • FIG. 98 are EpiMatrix staircase reports for identified MHC class I clusters of exemplary SARS-CoV-2 peptides of FIG. 70.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figure. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”.
  • FIG. 99 are EpiMatrix staircase reports for identified MHC class I clusters of exemplary SARS-CoV-2 peptides of FIG. 70.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figure. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”.
  • FIG. 100 are EpiMatrix staircase reports for identified MHC class I clusters of exemplary SARS-CoV-2 peptides of FIG. 70.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figure. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”.
  • FIG. 101 are EpiMatrix staircase reports for identified MHC class I clusters of exemplary SARS-CoV-2 peptides of FIG. 70.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figure. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”.
  • FIG. 102 are EpiMatrix staircase reports for identified MHC class I clusters of exemplary SARS-CoV-2 peptides of FIG. 70.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figure. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”.
  • FIG. 103 are EpiMatrix staircase reports for identified MHC class I clusters of exemplary SARS-CoV-2 peptides of FIG. 70.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figure. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”.
  • FIG. 104 are EpiMatrix staircase reports for identified MHC class I clusters of exemplary SARS-CoV-2 peptides of FIG. 70.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figure. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”.
  • FIG. 105 are EpiMatrix staircase reports for identified MHC class I clusters of exemplary SARS-CoV-2 peptides of FIG. 70.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figure. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”.
  • FIG. 106 are EpiMatrix staircase reports for identified MHC class I clusters of exemplary SARS-CoV-2 peptides of FIG. 70.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figure. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”.
  • FIG. 107 are EpiMatrix staircase reports for identified MHC class I clusters of exemplary SARS-CoV-2 peptides of FIG. 70.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figure. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”.
  • FIG. 108 are EpiMatrix staircase reports for identified MHC class I clusters of exemplary SARS-CoV-2 peptides of FIG. 70.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figure. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”.
  • FIG. 109 are EpiMatrix staircase reports for identified MHC class I clusters of exemplary SARS-CoV-2 peptides of FIG. 70.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figure. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”.
  • FIG. 110 are EpiMatrix staircase reports for identified MHC class I clusters of exemplary SARS-CoV-2 peptides of FIG. 70.
  • Z-score indicates the potential of a 9-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figure. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”.
  • FIG. Ill are EpiMatrix staircase reports for identified MHC class I clusters of exmplary SARS-CoV-2 peptides of FIG. 70.
  • Z-score indicates the potential of a 10-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figure. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”.
  • FIG. 112 are EpiMatrix staircase reports for identified MHC class I clusters of exmplary SARS-CoV-2 peptides of FIG. 70.
  • Z-score indicates the potential of a 10-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figure. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”.
  • FIG. 113 are EpiMatrix staircase reports for identified MHC class I clusters of exmplary SARS-CoV-2 peptides of FIG. 70.
  • Z-score indicates the potential of a 10-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figure. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”.
  • FIG. 114 are EpiMatrix staircase reports for identified MHC class I clusters of exmplary SARS-CoV-2 peptides of FIG. 70.
  • Z-score indicates the potential of a 10-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figure. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”.
  • FIG. 115 are EpiMatrix staircase reports for identified MHC class I clusters of exmplary SARS-CoV-2 peptides of FIG. 70.
  • Z-score indicates the potential of a 10-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figure. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”.
  • FIG. 116 are EpiMatrix staircase reports for identified MHC class I clusters of exmplary SARS-CoV-2 peptides of FIG. 70.
  • Z-score indicates the potential of a 10-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figure. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”.
  • FIG. 117 are EpiMatrix staircase reports for identified MHC class I clusters of exmplary SARS-CoV-2 peptides of FIG. 70.
  • Z-score indicates the potential of a 10-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figure. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”.
  • FIG. 118 are EpiMatrix staircase reports for identified MHC class I clusters of exmplary SARS-CoV-2 peptides of FIG. 70.
  • Z-score indicates the potential of a 10-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figure. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”.
  • FIG. 119 are EpiMatrix staircase reports for identified MHC class I clusters of exmplary SARS-CoV-2 peptides of FIG. 70.
  • Z-score indicates the potential of a 10-mer frame to bind to a given HLA allele; the strength of the score is indicated by the hashing as noted in the respective Figure. All scores in the Top 5% (Z-Score > 1.64) are considered “Hits”.
  • FIG. 120 shows the sequences of some of the concatemers disclosed herein along with their corresponding sequence identifiers.
  • FIG. 121C shows predicted SARS-CoV-2 T cell epitopes are antigenic ex vivo in COVID-19 convalescent donors but not healthy donors whererin the frequency of responses to unique peptides within the cohort (vertical lines denote 20% of each cohort).
  • FIG. 122B SARS-CoV-2 experienced individuals exhibit variable immune recall responses ex vivo wherein the correlation of cumulative T cell responses and age according to gender.
  • FIG. 123B shows antigen-specific T cell expansion increases responses in COVID-19 convalescents and uncovers pre-existing SARS-CoV-2 immunity in healthy donors wherin IFNy responses to individual peptides were also assessed, identifying the breadth of response in individual donors.
  • FIG. 123C shows antigen-specific T cell expansion increases responses in COVID-19 convalescents and uncovers pre-existing SARS-CoV-2 immunity in healthy donors wherein the frequency of responses to unique peptides within the cohort (vertical lines denote 20% of each cohort).
  • FIG. 124A shows EPV-CoV-19 immunization stimulates strong type 1 -skewed T cell responses in HLA-DR3 transgenic mice. Eight days post-boost, murine splenocytes were isolated and assayed for epitope-specific recall responses. Cells were plated in dual IFNy/IL-4 fluorospot plates, and restimulated with peptide pools for 48h wherein representative images and spot counts are shown for both.
  • FIG. 124B shows EPV-CoV-19 immunization stimulates strong type 1 -skewed T cell responses in HLA-DR3 transgenic mice. Eight days post-boost, murine splenocytes were isolated and assayed for epitope-specific recall responses. Cells were plated in dual IFNy/IL-4 fluorospot plates, and restimulated with peptide pools for 48h wherein IFNy SFC counts were normalized to lxlO 6 cells and adjusted by background subtraction.
  • FIG. 124C shows EPV-CoV-19 immunization stimulates strong type 1 -skewed T cell responses in HFA-DR3 transgenic mice. Eight days post-boost, murine splenocytes were isolated and assayed for epitope-specific recall responses. Cells were plated in dual IFNy/IE-4 fluorospot plates, and restimulated with peptide pools for 48h wherein IFNy SI index was determined by calculating the fold change of individual restimulation replicates over background.
  • FIG. 124D shows EPV-CoV-19 immunization stimulates strong type 1 -skewed T cell responses in HFA-DR3 transgenic mice. Eight days post-boost, murine splenocytes were isolated and assayed for epitope-specific recall responses. Cells were plated in dual IFNy/IE-4 fluorospot plates, and restimulated with peptide pools for 48h wherein IFNy SI index was determined by calculating the fold change of individual restimulation replicates over background.
  • FIG. 124E illustrates exemplary assays.
  • FIG. 124F shows that from the reported IFNy and IF-4 stimulation indexes, we calculated the IFNy:IF-4 ratio of each restimulation replicate to model the overall skewing of the immune response, identifying a sharply type 1 skewed phenotype in all vaccinated animals.
  • FIG. 125A shows EPV-CoV-19 immunization stimulates type 1 -skewed Memory CD4 and CD8 T cells in HFA-DR3 transgenic mice.
  • Splenocytes were restimulated with a vaccine- matched peptide pool 6 hours in the presence of brefeldin A and monensin. Following incubation, cells were stained for surface markers, fixed and permeabilized, stained for intracellular markers, and expression of markers was recorded by flow cytometry.
  • Memory CD4 + T cells and CD8 + T cells were assessed for IFNy, IL-4, or IL-5-production (both frequency in parent T cell population and mean fluorescence intensity (MFI) of cytokines) wherein representative images of type 1 and type 2 skewed, epitope-specific memory T cell populations are shown.
  • FIG. 125B shows EPV-CoV-19 immunization stimulates type 1 -skewed Memory CD4 and CD8 T cells in HLA-DR3 transgenic mice.
  • Splenocytes were restimulated with a vaccine- matched peptide pool 6 hours in the presence of brefeldin A and monensin. Following incubation, cells were stained for surface markers, fixed and permeabilized, stained for intracellular markers, and expression of markers was recorded by flow cytometry.
  • Memory CD4 + T cells and CD8 + T cells were assessed for IFNy production (both frequency in parent T cell population and mean fluorescence intensity (MFI) of cytokines) wherein the fold increase of epitope-specific responses (over CD28 stimulated controls) identify vaccine-specific induction of IFNy.
  • MFI mean fluorescence intensity
  • FIG. 125C shows EPV-CoV-19 immunization stimulates type 1 -skewed Memory CD4 and CD8 T cells in HLA-DR3 transgenic mice.
  • Splenocytes were restimulated with a vaccine- matched peptide pool 6 hours in the presence of brefeldin A and monensin. Following incubation, cells were stained for surface markers, fixed and permeabilized, stained for intracellular markers, and expression of markers was recorded by flow cytometry.
  • FIG. 125D shows EPV-CoV-19 immunization stimulates type 1 -skewed Memory CD4 and CD8 T cells in HLA-DR3 transgenic mice.
  • Splenocytes were restimulated with a vaccine- matched peptide pool 6 hours in the presence of brefeldin A and monensin. Following incubation, cells were stained for surface markers, fixed and permeabilized, stained for intracellular markers, and expression of markers was recorded by flow cytometry.
  • the present disclosure generally relates to T-cell epitope-based compounds and compositions, including pan beta-coronavirus vaccines, for use against any beta-coronavirus infection and related diseases caused by a beta-coronavirus, including MERS, SARS, and COVID-19.
  • the disclosure relates to immunogenic peptides, polypeptides, concatemeric peptides, and chimeric or fusion polypeptides and the uses thereof, particularly in pharmaceutical and vaccine compositions.
  • the present disclosure also relates to nucleic acids, vectors (including expression vectors), and cells which express the peptides, polypeptides, concatemeric peptides, and chimeric or fusion polypeptides and the uses thereof.
  • the peptides, polypeptides, concatemeric peptides, and chimeric or fusion polypeptides of the present disclosure more specifically comprise an agretope predicted to be a ligand of HLA class I and/or HLA class II MHC molecules, as well as an epitope that is predicted to be recognized by T-cells (including CD8+ and/or CD4+ T-cells) in the context of MHC class I and/or class II molecules.
  • the instant disclosure is particularly suited to produce vaccines for humans, particularly for vaccinating against beta-coronavirus infection, including MERS-CoV, SARS- CoV, and/or SARS-CoV-2 infection and related diseases caused by MERS-CoV, SARS-CoV, and/or SARS-CoV-2, including MERS, SARS, and COVID-19.
  • beta-coronavirus infection including MERS-CoV, SARS- CoV, and/or SARS-CoV-2 infection and related diseases caused by MERS-CoV, SARS-CoV, and/or SARS-CoV-2, including MERS, SARS, and COVID-19.
  • T-cells It is possible to exploit epitope-specific T-cells to induce an immune response against specific antigens. This discovery has implications for the design of therapeutic regimens and antigen-specific therapies against particular pathogens and infections.
  • the instant disclosure and data relates to identified beta-coronvirus T cell epitopes that are recognized in natural infection and stimulate pre-existing immunity to all beta-coronaviruses. These epitopes are excellent candidates for T cell-directed vaccine development.
  • a T cell targeting vaccine composed of conserved epitopes may provide rapid, effective and long-term immunity at sites of infection with production of tissue resident memory CD8 + T cells, as well as memory CD4 + T cells that support antibody responses.
  • T-cell epitopes including a T-cell epitope compound or composition of the present disclosure (including two or more of peptides or polypeptides having a sequence comprising, consisting of, or consisting essentially of one or more of SEQ ID NOS: 1305, 1616, 1317, 1628, 1333,
  • concatemeric peptides as disclosed herein including a concatemeric polypeptide of the present disclosure that comprises, consists of, or consists essentially of one or more of SEQ ID NOS: 1305, 1616, 1317, 1628, 1333, 1644, 1311, 1622, 1309, 1620, 1330, 1641, 1254, 1565, 1314, 1625, 1272, 1583, 1303, 1614, 1110, 1421, 1099,
  • the T-cell epitope compounds and compositions of the present disclosure are useful in the selective engagement and activation of immunogenic T-cells. It is demonstrated herein that certain naturally occurring T-cells (in aspects, including CD4+ and CD8+ T-cells), can be engaged, activated, and/or applied to induce immunity or induce an immune response against pathogens such as MERS CoV, SARS CoV, or SARS-CoV-2 and related diseases caused by MERS CoV, SARS CoV, or SARS-CoV-2, including MERS, SARS, and COVID-19.
  • MERS CoV MERS CoV
  • SARS CoV SARS CoV
  • SARS-CoV-2 SARS-CoV-2 and related diseases caused by MERS CoV, SARS CoV, or SARS-CoV-2, including MERS, SARS, and COVID-19.
  • T-cell epitope compounds and compositions of the present disclosure can be used to stimulate, induce, and/or expand an immune response to a beta-coronavirus, including MERS CoV, SARS CoV, or SARS-CoV-2 in a subject, and thus can be used in methods of treating and/or preventing beta-coronavirus infection and related diseases caused by beta-coronavirus in a subject.
  • a beta-coronavirus including MERS CoV, SARS CoV, or SARS-CoV-2 in a subject
  • Ranges provided herein are understood to be shorthand for all of the values within the range.
  • a range of 1 to 25 is understood to include any number, combination of numbers, or sub-range from the group consisting of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, or 25, as well as all intervening decimal values between the aforementioned integers such as, for example, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, and 1.9.
  • sub-ranges “nested sub-ranges” that extend from either end point of the range are specifically contemplated.
  • a nested sub-range of an exemplary range of 1 to 25 may comprise 1 to 5, 1 to 10, 1 to 15, and 1 to 20 in one direction, or 25 to 20, 25 to 15, 25 to 10, and 25 to 5 in the other direction.
  • biological sample refers to any sample of tissue, cells, or secretions from an organism.
  • medical condition includes, but is not limited to, any condition or disease manifested as one or more physical and/or psychological symptoms for which treatment and/or prevention is desirable, and includes previously and newly identified diseases and other disorders.
  • the term “immune response” refers to the concerted action of lymphocytes, antigen presenting cells, phagocytic cells, granulocytes, and soluble macromolecules produced by the above cells or the liver (including antibodies, cytokines, and complement) that results in selective damage to, destruction of, or elimination from the human body of cancerous cells, metastatic tumor cells, malignant melanoma, invading pathogens (including a virus), cells or tissues infected with pathogens, or, in cases of autoimmunity or pathological inflammation, normal human cells or tissues.
  • an immune response includes a measurable cytotoxic T lymphocyte (CTL) response (e.g., against a virus expressing an immunogenic polypeptide) or a measurable B cell response, such as the production of antibodies, (e.g., against an immunogenic polypeptide).
  • CTL cytotoxic T lymphocyte
  • B cell response such as the production of antibodies, (e.g., against an immunogenic polypeptide).
  • B lymphocyte and T lymphocyte assays are well known, such as ELISAs, EliSpot assays, cytotoxic T lymphocyte CTL assays, such as chromium release assays, proliferation assays using peripheral blood lymphocytes (PBL), tetramer assays, and other cytokine production assays.
  • PBL peripheral blood lymphocytes
  • the term “effective amount”, “therapeutically effective amount”, or the like of a composition, including a T cell epitope compound or composition of the present disclosure including one or more of peptides or polypeptides having a sequence comprising, consisting of, or consisting essentially of one or more ofSEQ IDNOS: 1305, 1616, 1317, 1628, 1333, 1644, 1311, 1622, 1309, 1620, 1330, 1641, 1254, 1565, 1314, 1625, 1272, 1583, 1303,
  • concatemeric peptides as disclosed herein including concatemeric polypeptides comprising, consisting of, or consisting essentially of one or more of SEQ ID NOS: 1305, 1616, 1317, 1628, 1333, 1644, 1311, 1622, 1309, 1620, 1330, 1641, 1254, 1565, 1314, 1625, 1272, 1583, 1303, 1614, 1110, 1421, 1099, 1410, 1063, 1374, 1079,
  • compositions of the present disclosure administered to the subject will depend on the type and severity of the disease and on the characteristics of the individual, such as general health, age, sex, body weight and tolerance to drugs. It will also depend on the degree, severity and type of disease. The skilled artisan will be able to determine appropriate dosages depending on these and other factors.
  • the T cell epitope compounds and compositions of the present disclosure can also be administered in combination with each other or with one or more additional therapeutic compounds.
  • anti-beta-coronavirus activity As used herein, “anti-beta-coronavirus activity”, “anti-beta-coronavirus polypeptides”, “anti-beta-coronavirus compounds and compositions”, and the like are intended to mean that the T cell epitope compounds and compositions of the of the present diclsoure (including polypeptides, concatemeric polypeptides, chimeric or fusion proteins, nucleic acids, plasmids, vectors, pharmaceutical compositions, vaccines, and other compositions of the instant disclosure) have anti-beta-coronavirus activity and thus are capable of suppressing, controlling, and/or killing an invading beta-coronavirues broadly.
  • anti-betacoronavirus activity means that the instantly-disclosed therapeutic T-cell epitope comopounds and compositions are, in aspects: capable of stimulating, inducing, and/or expanding an immune response to any beta-coronavirus including MERS CoV, SARS CoV, or SARS-CoV-2 (e.g., a cellular (CD4+ and/or CD8+ T-cell response) or humoral immune response to such beta- coronaviruses) and/or associated diseases in a subject; capable of stimulating, inducing, and/or expanding a beta-coronavirus-specific IFNy response (e.g., by lymphocytes such as PMBC, or effector CD4+ and/or CD8+ T-cells), capable of inhibiting beta-coronavirus viral replication or infectivity, and/or capable of inducing immunity against any beta-coronavirus, optionally MERS CoV, SARS CoV, and/or SARS-CoV-2.
  • MERS CoV e.
  • a T-cell epitope compound or composition of the present disclosure having anti-beta-coronavirus activity will reduce the disease symptoms resulting from beta-coronavirus challenge by at least about 5% to about 50%, at least about 10% to about 60%, at least about 30% to about 70%, at least about 40% to about 80%, or at least about 50% to about 90% or greater, including any value or range therebetween.
  • Anti-beta-coronavirus activity can be determined by various experiments and assays as known to those of skill in the art, including methods such as by antibody titrations of sera, e.g., by ELISA and/or seroneutralization assay analysis and/or by vaccination challenge evaluation, including use of experiments and assays as disclosed in the Examples herein.
  • T cell epitope means an MHC ligand or protein determinant, 7 to 30 amino acids in length, and capable of specific binding to human leukocyte antigen (HLA) molecules and interacting with specific T cell receptors (TCRs).
  • HLA human leukocyte antigen
  • TCRs T cell receptors
  • the terms “engage”, “engagement” or the like means that when bound to a MHC molecule (e.g. human leukocyte antigen (HLA) molecules), the T cell epitope is capable of interacting with the TCR of the T cell and activating the T cell.
  • MHC molecule e.g. human leukocyte antigen (HLA) molecules
  • T cell epitopes are linear and do not express specific three-dimensional characteristics.
  • T-cell epitopes are not affected by the presence of denaturing solvents.
  • the ability to interact with T cell epitopes can be predicted by in silico methods (De Groot AS et ah, (1997), AIDS Res Hum Retroviruses, 13(7):539-41; Schafer JR et al., (1998), Vaccine, 16(19): 1880-4; De Groot AS et al., (2001), Vaccine, 19(31):4385-95; De Groot AR et al, (2003), Vaccine, 21(27-30):4486-504, all of which are herein incorporated by reference in their entirety.
  • T cell epitope cluster refers to polypeptide that contains between about 4 to about 40 MHC binding motifs.
  • the T-cell epitope cluster contains between about 5 to about 35 MHC binding motifs, between about 8 and about 30 MHC binding motifs; and between about 10 and 20 MHC binding motifs.
  • immune-stimulating T-cell epitope polypeptide refers to a molecule capable of inducing an immune response, e.g., a humoral, T cell-based, or innate immune response.
  • the term “regulatory T cell”, “Treg” or the like means a subpopulation of T cells that suppress immune effector function, including the suppression or down regulation of CD4+ and/or CD8+ effector T cell (Teff) induction, proliferation, and/or cytokine production, through a variety of different mechanisms including cell-cell contact and suppressive cytokine production.
  • CD4+ Tregs are characterized by the presence of certain cell surface markers including but not limited to CD4, CD25, and FoxP3.
  • CD4+ regulatory T cells secrete immune suppressive cytokines and chemokines including but not limited to IL-10 and/or TGFp.
  • CD4+ Tregs may also exert immune suppressive effects through direct killing of target cells, characterized by the expression upon activation of effector molecules including but not limited to granzyme B and perforin.
  • CD8+ Tregs are characterized by the presence of certain cell surface markers including but not limited to CD8, CD25, and, upon activation, FoxP3.
  • regulatory CD8+ T cells secrete immune suppressive cytokines and chemokines including but not limited to IFNy, IL-10, and/or TGFp.
  • CD8 + Tregs may also exert immune suppressive effects through direct killing of target cells, characterized by the expression upon activation of effector molecules including but not limited to granzyme B and/or perforin.
  • regulatory T cell epitope refers to a “T cell epitope” that causes a tolerogenic response (Weber CA et ah, (2009), Adv Drug Deliv, 61(11):965-76) and is capable of binding to MHC molecules and engaging (i.e. interacting with and activating) circulating naturally occurring Tregs (in aspects, including natural Tregs and/or adaptive Tregs).
  • CD4+ regulatory T cells secrete immune suppressive cytokines and chemokines including but not limited to IL-10 and/or TGFfi.
  • CD4+ Tregs may also exert immune suppressive effects through direct killing of target cells, characetized by the expression upon activation of effector molecules including but not limited to granzyme B and perforin, leads to the expression of the immune suppressive cytokines including, but not limited to, IL-10 and TGF-b and TNF-a.
  • regulatory CD8+ T cells secrete immune suppressive cytokines and chemokines including but not limited to IFNy, IL-10, and/or TGFp.
  • CD8 + Tregs may also exert immune suppressive effects through direct killing of target cells, characetized by the expression upon activation of effector molecules including but not limited to granzyme B and/or perforin.
  • B-cell epitope means a protein determinant capable of specific binding to an antibody.
  • B-cell epitopes usually consist of chemically active surface groupings of molecules such as amino acids or sugar side chains and usually have specific three-dimensional structural characteristics, as well as specific charge characteristics. Conformational and non-conformational epitopes are distinguished in that the binding to the former but not the latter is lost in the presence of denaturing solvents.
  • subject refers to any living organism in which an immune response is elicited.
  • subject includes, but is not limited to, humans, nonhuman primates such as chimpanzees and other apes and monkey species; farm animals such as cattle, sheep, pigs, goats and horses; domestic mammals such as dogs and cats; laboratory animals including rodents such as mice, rats and guinea pigs, and the like.
  • the term does not denote a particular age or sex. Thus, adult, child, and newborn subjects, as well as fetuses, whether male or female, are intended to be covered.
  • the terms “the major histocompatibility complex (MHC)”, “MHC molecules”, “MHC proteins” or “HLA proteins” are to be understood as meaning, in particular, proteins capable of binding peptides resulting from the proteolytic cleavage of protein antigens and representing potential T-cell epitopes, transporting them to the cell surface and presenting them there to specific cells, in particular cytotoxic T-lymphocytes or T-helper cells.
  • the major histocompatibility complex in the genome comprises the genetic region whose gene products expressed on the cell surface are important for binding and presenting endogenous and/or foreign antigens and thus for regulating immunological processes.
  • the major histocompatibility complex is classified into two gene groups coding for different proteins, namely molecules of MHC class I and molecules of MHC class II.
  • the molecules of the two MHC classes are specialized for different antigen sources.
  • the molecules of MHC class I present endogenously synthesized antigens, for example viral proteins and tumor antigens.
  • the molecules of MHC class II present protein antigens originating from exogenous sources, for example bacterial products.
  • the cellular biology and the expression patterns of the two MHC classes are adapted to these different roles.
  • MHC molecules of class I consist of a heavy chain and a light chain and are capable of binding a peptide of about 8 to 11 amino acids, but usually 9 or 10 amino acids, if this peptide has suitable binding motifs, and presenting it to cytotoxic T-lymphocytes.
  • the peptide bound by the MHC molecules of class I originates from an endogenous protein antigen.
  • the heavy chain of the MHC molecules of class I is preferably an HLA-A, HLA-B or HLA-C monomer, and the light chain is b-2-microglobulin.
  • MHC molecules of class II consist of an a-chain and a b-chain and are capable of binding a peptide of about 12 to 25 amino acids if this peptide has suitable binding motifs, and presenting it to T-helper cells.
  • the peptide bound by the MHC molecules of class II usually originates from an extracellular of exogenous protein antigen.
  • the a-chain and the b-chain are in particular HLA- DR, HLA-DQ and HLA-DP monomers.
  • MHC complex refers to a protein complex capable of binding with a specific repertoire of polypeptides known as HLA ligands and transporting said ligands to the cell surface.
  • MHC Ligand means a polypeptide capable of binding to one or more specific MHC alleles.
  • HLA ligand is interchangeable with the term “MHC Ligand”.
  • Cells expressing MHC/Ligand complexes on their surface are referred to as “Antigen Presenting Cells” (APCs).
  • MHC binding peptide relates to a peptide which binds to an MHC class I and/or an MHC class II molecule. In the case of MHC class I/peptide complexes, the binding peptides are typically 8-10 amino acids long although longer or shorter peptides may be effective.
  • T Cell Receptor refers to a protein complex expressed by T cells that is capable of engaging a specific repertoire of MHC/Ligand complexes as presented on the surface of cells, such as antigen presenting cells (APCs).
  • MHC Binding Motif refers to a pattern of amino acids in a protein sequence that predicts binding to a particular MHC allele.
  • AAA cleavage motif refers to the short amino acid motif consisting of the sequence “alanine-alanine -tyrosine” capable of promoting proteasome- mediated cleavage of a peptide or protein, promoting the binding of the transporter associated with antigen processing to a peptide or protein, and/or increasing proteasome degradation at specific sites within a peptide or protein.
  • immune synapse means the protein complex formed by the simultaneous engagement of a given T cell epitope to both a cell surface MHC complex and TCR.
  • polypeptide refers to a polymer of amino acids, and not to a specific length; thus, peptides, oligopeptides and proteins are included within the definition of a polypeptide.
  • a polypeptide is said to be “isolated” or “purified” when it is substantially free of cellular material when it is isolated from recombinant and non-recombinant cells, or free of chemical precursors or other chemicals when it is chemically synthesized.
  • a peptide or polypeptide e.g., a polypeptide comprising, consisting of, or consisting essentially of one or more of SEQ ID NOS: 4-1676, 1713-2595, 2605-2638, 2647-2718, and 2735-8692 or variants and fragments thereof, optionally SEQ ID NOs: 1305, 1616, 1317, 1628, 1333, 1644, 1311, 1622, 1309, 1620, 1330, 1641, 1254, 1565, 1314, 1625, 1272, 1583, 1303, 1614, 1110, 1421,
  • one or more T-cell epitopes of the present disclosure can be joined to, linked to, or inserted into another polypeptide wherein said one or more T-cell epitopes of the present disclosure is not naturally included in the polypeptide and/or said one or more T cell epitopes of the present disclosure is not located at its natural position in the polypeptide.
  • concatemeric peptide or polypeptide refers to a series of at least two peptides or polypeptides linked together. Such linkages may form of string-of-beads design.
  • concatemeric polypeptides of the instant disclosure include concatemeric polypeptides comprising, consisting of, or consisting essentially of one or more of SEQ ID NOs: 1305, 1616, 1317, 1628, 1333, 1644, 1311, 1622, 1309, 1620, 1330, 1641, 1254, 1565,
  • linker refers to a peptide added between two peptide domains such as epitopes or vaccine sequences to connect said peptide domains.
  • a linker sequence is used to reduce steric hindrance between each one or more identified peptides of the instant disclosure, is well translated, and supports or allows processing of the each one or more identified polypeptides of the instant disclosure.
  • the linker should have little or no immunogenic sequence elements.
  • each peptide or polypeptide of the concatemeric polypeptide may optionally have one or more linkers, which may optionally be cleavage sensitive sites, adjacent to their N and/or C terminal end. In such a concatemeric peptide, two or more of the peptides may have a cleavage sensitive site between them. Alternatively two or more of the peptides may be connected directly to one another or through a linker that is not a cleavage sensitive site.
  • the term “pharmaceutically acceptable” refers to approved or approvable by a regulatory agency of the Federal or a state government or listed in the U.S. Pharmacopeia or other generally recognized pharmacopeia for use in animals, including humans.
  • the term “pharmaceutically acceptable excipient, carrier, or diluent” or the like refer to an excipient, carrier, or diluent that can be administered to a subject, together with an agent, and which does not destroy the pharmacological activity thereof and is nontoxic when administered in doses sufficient to deliver a therapeutic amount of the agent.
  • the term “purpose built computer program” refers to a computer program designed to fulfill a specific purpose; typically to analyze a specific set of raw data and answer a specific scientific question.
  • z-score indicates how many standard deviations an element is from the mean.
  • the singular forms “a,” “an,” and “the” are intended to include the plural forms, including “at least one,” unless the content clearly indicates otherwise.
  • “Or” means “and/or.”
  • the term “and/or” and “one or more” includes any and all combinations of the associated listed items.
  • the term “one or more” with respect to the “one or more of SEQ ID NOS: 4-1676, 1713-2595, 2605-2638, 2647-2718, and 2735- 8692 of the present disclosure” includes any and all combinations of SEQ ID NOS: 4-1676, 1713-2595, 2605-2638, 2647-2718, and 2735-8692.
  • the term “or a combination thereof’ means a combination including at least one of the foregoing elements.
  • a “variant” peptide or polypeptide can differ in amino acid sequence by one or more substitutions, deletions, insertions, inversions, fusions, and truncations or a combination of any of these.
  • a variant peptide or polypeptide can differ in amino acid sequence by one or more substitutions, deletions, insertions, inversions, fusions, and truncations or a combination of any of these provided said variants retain MHC binding propensity and/or TCR specificity, and/or beta-coronavirus activity.
  • the present disclosure also includes fragments of the peptide or polypeptides of the invention.
  • the disclosure also encompasses fragments of the variants of the T-cell epitopes described herein, provided said fragments and/or variants at least in part retain MHC binding propensity and/or TCR specificity, and/or retain anti-beta-coronavirus activity.
  • the present disclosure also provides chimeric or fusion polypeptides (which in aspects may be isolated, synthetic, or recombinant) wherein one or more of the instantly-disclosed peptides, polypeptides, or concatemeric peptides is a part thereof.
  • a chimeric or fusion polypeptide composition comprises one or more peptides, polypeptides, or concatemeric peptides of the instant disclosure linked to a heterologous polypeptide.
  • heterologous polypeptide is intended to mean that the one or more T-cell epitopes (e.g., one or more of SEQ ID NOS: 1305, 1616, 1317, 1628, 1333, 1644, 1311, 1622, 1309, 1620, 1330, 1641, 1254, 1565, 1314, 1625, 1272, 1583, 1303, 1614, 1110, 1421, 1099, 1410,
  • the one or more peptides, polypeptides, or concatemeric peptides of the instant disclosure may be inserted into the heterologous polypeptide (e.g., through mutagenesis or other known means in the art), may be added to the C-terminus (with or without the use of linkers, as is known in the art), and/or added to the N- terminus (with or without the use of linkers, as is known in the art) of the heterologous polypeptide.
  • protein engineering by mutagenesis can be performed using site- directed mutagenesis techniques, or other mutagenesis techniques known in the art (see e.g., James A. Brannigan and Anthony J. Wilkinson., 2002, Protein engineering 20 years on. Nature Reviews Molecular Cell Biology 3, 964-970; Turanli-Yildiz B. et ah, 2012, Protein Engineering Methods and Applications, intechopen.com, which are herein incorporated by reference in their entirety).
  • chimeric or fusion polypeptides comprise one or more peptides, polypeptides, or concatemeric peptides of the instant disclosure operatively linked to a heterologous polypeptide. “Operatively linked” indicates that the polypeptide (e.g., the one or more T-cell epitope polypeptides of the present disclosure) and the heterologous protein are fused in-frame or chemically-linked or otherwise bound.
  • the instantly-disclosed chimeric or fusion polypeptides may be isolated, synthetic, or recombinant.
  • an “isolated” peptide, polypeptide, concatemeric peptide e.g., an isolated T-cell activating T-cell epitope or T-cell epitope polypeptide
  • chimeric or fusion polypeptide can be purified from cells that naturally express it, purified from cells that have been altered to express it (recombinant), or synthesized using known protein synthesis methods.
  • a peptide, polypeptide, or concatemeric peptide is produced by recombinant DNA or RNA techniques.
  • a nucleic acid molecule encoding the peptide, polypeptide, concatemeric peptide, or chimeric or fusion polypeptide is cloned into an expression vector, the expression vector introduced into a host cell and the peptide, polypeptide, concatemeric peptide, or chimeric or fusion polypeptide is expressed in the host cell.
  • the peptide, polypeptide, concatemeric peptide, or chimeric or fusion polypeptide can then be isolated from the cells by an appropriate purification scheme using standard protein purification techniques.
  • peptides, polypeptides, concatemeric peptides, or chimeric or fusion polypeptides of the instant disclosure can include, for example, modified forms of naturally occurring amino acids such as D-stereoisomers, non-naturally occurring amino acids; amino acid analogs; and mimetics.
  • peptides, polypeptides, concatemeric peptides, or chimeric or fusion polypeptides of the instant disclosure can include retro-inverso peptides of the instantly disclosed peptides, polypeptides, concatemeric peptides, or chimeric or fusion polypeptides of the instant disclosure, provided said peptides, polypeptides, concatemeric peptides, or chimeric or fusion polypeptides of the instant disclosure at least in part retain MHC binding propensity and/or TCR specificity, and/or retain anti-beta-coronavirus activity.
  • the present disclosure provides a novel class of T-cell epitopes (which may be isolated, synthetic, or recombinant), which comprise a peptide or polypeptide chain derived from beta-coronavirus proteins (e.g., encoded proteins from a beta-coronavirus genome), including the envelope, membrane, spike, nucleocapsid, ORF3a, ORF6, ORF7a, ORF8, ORFIO, ORFlab non- structural protein 2 (NSP2), ORFlab non-structural protein 3 (NSP3), ORFlab non-structural protein 4 (NSP4), ORFlab 3C-like proteinase, ORFlab non-structural protein 6 (NSP6), ORFlab non-structural protein 7 (NSP7), ORFlab non-structural protein 8 (NSP8), ORFlab non-structural protein 9 (NSP9), ORFlab non-structural protein 10 (NSP10), ORFlab RNA-dependent RNA polymerase, ORFlab helicase,
  • T-cell epitopes of the present disclosure are highly conserved among known variants of their source proteins, and SARS- CoV-2 (taxid: 2697049), SARS-CoV-1 (taxid: 694009), MERS-CoV (taxid: 1335626), and human CoV (taxids: 11137, 443239, 277944 and 31631) antigen sequences isolated from human hosts were obtained from GenBank at the National Center for Biotechnology Information. SARS-CoV-2 epitopes were compared across sequences obtained from isolates with fully sequenced genomes isolated from December 2019 to December 2020 for T cell epitope mapping. SARS-CoV-2 Wuhan-Hu-1 (GenBank id: MN908947) was selected as the reference strain.
  • T-cell epitopes of the present disclosure comprise at least one putative T cell epitope as identified by EpiMatrixTM analysis.
  • EpiMatrixTM is a proprietary computer algorithm developed by EpiVax (Providence, Rhode Island), which is used to screen protein sequences for the presence of putative T cell epitopes.
  • the algorithm uses matrices for prediction of 9- and 10-mer peptides binding to MHC molecules. Each matrix is based on position-specific coefficients related to amino acid binding affinities that are elucidated by a method similar to, but not identical to, the pocket profile method (Sturniolo, T. et al. , Nat. BiotechnoL, 17:555-561, 1999).
  • Input sequences are, for example, parsed into overlapping 9-mer frames or 10-mer where each frame overlaps the last by 8 or 9 amino acids, respectively.
  • Each of the resulting frames form the mutated peptide and the non-mutated peptide are then scored for predicted binding affinity with respect to MHC class I alleles (e.g., but not limited to, HLA-A and HLA-B alleles) and MHC class II alleles (e.g., but not limited to HLA-DRB1 alleles).
  • Raw scores are normalized against the scores of a large sample of randomly generated peptides.
  • the resulting “Z” scores are normally distributed and directly comparable across alleles.
  • the resulting “Z” score is reported.
  • any 9-mer or 10- mer peptide with an allele-specific EpiMatrixTM Z-score in excess of 1.64, theoretically the top 5% of any given sample is considered a putative T cell epitope.
  • peptides containing clusters of putative T cell epitopes are more likely to test positive in validating in vitro and in vivo assays.
  • the results of the initial EpiMatrixTM analysis are further screened for the presence of putative T cell epitope “clusters” using a second proprietary algorithm known as ClustimerTM algorithm.
  • the ClustimerTM algorithm identifies sub-regions contained within any given amino acid sequence that contains a statistically unusually high number of putative T cell epitopes.
  • Typical T-cell epitope “clusters” range from about 9 to roughly 30 amino acids in length and, considering their affinity to multiple alleles and across multiple 9-mer frames, can contain anywhere from about 4 to about 40 putative T cell epitopes.
  • FIG. 1 is an overview of MHC class II cluster selection from the envelope (SEQ ID NO: 1) of SARS-CoV-2.
  • FIG. 5 is an overview of MHC class II cluster selection from the membrane (SEQ ID NO: 2) of SARS- CoV-2.
  • FIG. 16 is an overview of MHC class II cluster selection from the spike (SEQ ID NO: 3) of SARS-CoV-2.
  • Each epitope cluster identified an aggregate EpiMatrixTM score is calculated by summing the scores of the putative T cell epitopes and subtracting a correcting factor based on the length of the candidate epitope cluster and the expected score of a randomly generated cluster of the same length. EpiMatrixTM cluster scores in excess of +10 are considered significant.
  • the T-cell epitopes of the instant disclosure contain several putative T-cell epitopes forming a pattern known as a T-cell epitope cluster.
  • FIGS. 2-4 are EpiMatrix Cluster detail reports for identified MHC class II clusters of the envelope (SEQ ID NO: 1) of SARS-CoV-2 and relate to SEQ ID NOS: 4-68 and 1003- 1005.
  • FIGS. 6-15 are EpiMatrix Cluster detail reports for identified MHC class II clusters of the membrane (SEQ ID NO: 2) of SARS-CoV-2 and relate to SEQ ID NOS: 69-209, 1006- 1015, 2255, 2561, and 8691, and 8692.
  • FIGS. 17-55 are EpiMatrix Cluster detail reports for identified MHC class II clusters of the spike (SEQ ID NO: 3) of SARS-CoV-2 and relate to SEQ ID NOS: 210-707 and 1016-1054.
  • FIG. 59 is an EpiMatrix staircase report for identified MHC class I clusters of the envelope (SEQ ID NO: 1) of SARS-CoV-2 and relates to SEQ ID NOS: 708-739.
  • FIG. 60 is an EpiMatrix staircase report for identified MHC class I clusters of the membrane (SEQ ID NO: 2) of SARS-CoV-2 and relates to SEQ ID NOS: 740-851.
  • FIG. 61 is an EpiMatrix staircase report for identified MHC class I clusters of the spike (SEQ ID NO: 3) of SARS-CoV-2 and relates to SEQ ID NOS: 852-1002.
  • FIGS. 71-92 are EpiMatrix Cluster detail reports for identified MHC class II clusters of the SARS-CoV-2 peptides listed in FIG. 70 and relate to SEQ ID NOS: 1713-2010.
  • FIGS. 93-101 are EpiMatrix staircase reports for identified MHC class I clusters of the SARS-CoV-2 peptides identified in FIG. 70 and relate to SEQ ID NOS: 1713-2158.
  • FIGS. 102-110 are EpiMatrix staircase reports for identified MHC class II clusters of the SARS-CoV-2 peptides identified in FIG. 70 and relate to SEQ ID NOS: 1713-2158.
  • FIGS. 111-119 are EpiMatrix staircase reports for identified MHC class I lOmer clusters of the SARS-CoV-2 peptides identified in FIG. 70 and relate to SEQ ID NOS: 2159-2569.
  • Putative T-cell epitopes were also screened for cross-conservation with the human proteome using JanusMatrix, as further described in more detail in the Examples.
  • the JanusMatrix system (EpiVax, Buffalo, Rhode Island) useful for screening peptide sequences for cross-conservation with a host proteome.
  • JanusMatrix is an algorithm that predicts the potential for cross-reactivity between peptide clusters and the host genome or proteome, based on conservation of TCR-facing residues in their putative MHC ligands.
  • the JanusMatrix algorithm first considers all the predicted epitopes contained within a given protein sequence and divides each predicted epitope into its constituent agretope and epitope.
  • cross-conservation between human epitopes and the antigenic epitopes may indicate that such a candidate utilizes immune camouflage, thereby evading the immune response and making for an ineffective vaccine.
  • the host is, for example, a human
  • the peptide clusters are screened against human genomes and proteomes, based on conservation of TCR-facing residues in their putative HLA ligands. The peptides are then scored using the JanusMatrix Homology Score.
  • peptides with a JanusMatrix Homology Score below 2.5 or below 3.0 indicate low tolerogenicity potential and may be useful for pharmaceutical formulations and vaccines for the treatment/prevention of beta-coronavirus infection and related diseases caused by beta- coronavirues, and in aspects may be included from the T cell epitope compositions and methods of the present disclosure.
  • peptides with a JanusMatrix Homology Score above 3.0 indicate high tolerogenicity potential and may not be useful for pharmaceutical formulations and vaccines for the treatment/prevention of beta-coronavirus infection and related diseases caused by beta-coronaviruses, and in aspects may be excluded from the T cell epitope compositions and methods of the present disclosure.
  • one or more of a peptide or polypeptide having an amino acid sequence comprising, consisting of, or consisting essentially of one or more of SEQ ID NOS: 1056-1057, 1059, 1367-1368, 1370, 1063, 1068, 101, 1374, 1379, 1382, 1073, 1075, 1083-1084, 1094, 1101, 1103, 1108, 1111, 1112, 1117, 1384, 1386, 1394-1395, 1405, 1412, 1414, 1419, 1422, 1423, 1428, 1125, 1436, 1131, 1133, 1134, 1442, 1444, 1445, 1141, 1452, 1142, 1145, 1453, 1456, 1152, 1157, 1159, 1164, 1463,
  • FIG. 56 is the JanusMatrix reports for identified MHC class II clusters of the envelope (SEQ ID NO: 1) of SARS-CoV-2.
  • FIG. 57 is the JanusMatrix reports for identified MHC class II clusters of the membrane (SEQ ID NO: 2) of SARS-CoV- 2.
  • FIG. 58 is the JanusMatrix reports for identified MHC class II clusters of the spike (SEQ ID NO: 3) of SARS-CoV-2.
  • T-cell epitopes of the present disclosure are highly conserved among related coronaviruses including those that infect humans such as highly pathogenic SARS-CoV and MERS-CoV and low pathogenicity common cold coronaviruses (CCCs) OC43, HKU1, NL63, and 229E.
  • CCCs common cold coronaviruses
  • Prior exposure to these viruses may have established T cell memory that can be recalled upon beta-coronavirus infection and vaccination.
  • beta-coronavirus infection and vaccination establishes T cell memory that can influence responses in future infections to these viruses or yet-to-emerge coronaviruses.
  • TCR-face of select beta-coronavirus epitopes for homology with all coronaviruses, using the JanusMatrix algorithm (e.g., Table 58A and Table 58B).
  • the JanusMatrix algorithm e.g., Table 58A and Table 58B.
  • all of the selected membrane and envelope sequences shared identical TCR-face patterns with SARS-CoV.
  • Half the selected spike clusters were unique to SARS-CoV-2 and the other half are conserved with SARS-CoV. Only three selected clusters were cross-conserved outside SARS viruses. Given reports of preexisting T cell immunity in people with no SARS-CoV-2 experience, we relaxed the requirement for 100% identity at every TCR-face position.
  • JanusMatrix predicted an expanded cross-conservation landscape for selected SARS-CoV-2 spike and membrane clusters. Most selected spike clusters were conserved, by these criteria, in the subset of coronaviruses that infect humans. The remainder of the selected sequences with crossreactivity potential are cross-conserved among highly pathogenic beta-coronaviruses or among high and low pathogenicity beta-coronaviruses. Only three clusters were unique to SARS-CoV- 2 by the cristeria described above, and none are solely conserved with SARS-CoV.
  • the single membrane selected clusters were cross-conserved in the highly pathogenic beta-coronaviruses and coronaviruses that infect humans subsets. As the vast majority of people were not exposed to SARS-CoV and MERS-CoV, we also explored cross-conservation between SARS-CoV-2 and CCCs only.
  • T-cell epitopes of the present disclosure bind to at least one and preferably two or more common HLA class I and/or class II alleles with at least a moderate affinity (e.g ., in aspects, ⁇ 1000 mM ICso, ⁇ 500 mM ICso , ⁇ 400 mM ICso, ⁇ 300 mM ICso, or ⁇ 200 mM ICso in HLA binding assays based on soluble HLA molecules).
  • T-cell epitopes of the present disclosure are capable of being presented at the cell surface by cells in the context of at least one and, in other aspects, two or more alleles of the HLA.
  • the epitope-HLA complex can be recognized by CD4+ and/or CD8+ T-cells having TCRs that are specific for the epitope-HLA complex and circulating in subjects.
  • the recognition of the epitope-HLA complex can cause the matching T-cell to be activated and to secrete activating cytokines (e.g., effector cytokines such as IFNy) and chemokines.
  • a T-cell epitope compounds or compositions of the present disclosure includes one or more peptides or polypeptides a disclosed herein.
  • the present disclosure is directed to a peptide or polypeptide having an amino acid sequence comprising, consisting of, or consisting essentially of one or more ofSEQ IDNOS: 1305, 1616, 1317, 1628, 1333, 1644, 1311, 1622, 1309, 1620, 1330, 1641, 1254, 1565, 1314, 1625, 1272, 1583, 1303,
  • the peptides or polypeptides of the instant disclosure can be either in neutral (uncharged) or salt forms, and may be either free of or include modifications such as glycosylation, side chain oxidation, or phosphorylation.
  • the peptides or polypeptides of the instant disclosure can be capped with an n-terminal acetyl and/or c-terminal amino group.
  • peptides or polypeptides of the instant disclosure having SEQ ID NOS: 1305, 1616, 1317, 1628, 1333, 1644, 1311, 1622, 1309, 1620, 1330, 1641, 1254, 1565, 1314, 1625, 1272, 1583, 1303, 1614, 1110, 1421, 1099, 1410, 1063, 1374,
  • the instant disclosure is directed to a peptide or polypeptide comprising, consisting, or consisting essentially of an amino acid sequence of SEQ ID NOS: 1305, 1616, 1317, 1628, 1333, 1644, 1311, 1622, 1309, 1620, 1330, 1641, 1254, 1565, 1314, 1625, 1272,
  • a peptide or polypeptide have a core amino acid sequence comprising, consisting of, or consisting essentially of one or more peptides or polypeptides having an amino acid sequence of SEQ ID NOS: 1305, 1616, 1317, 1628, 1333, 1644, 1311, 1622, 1309, 1620, 1330, 1641, 1254, 1565,
  • flanking amino acids can be distributed in any ratio to the C-terminus and the N-terminus (for example all flanking amino acids can be added to one terminus,
  • the instant disclosure is directed to a peptide or polypeptide have a core sequence comprising, consisting of, or consisting essentially of one or more peptides or polypeptides having an amino acid sequence of SEQ ID NOS: 1305, 1616, 1317, 1628, 1333, 1644, 1311, 1622, 1309, 1620, 1330, 1641, 1254, 1565, 1314, 1625, 1272, 1583,
  • flanking amino acids 1 to 12, 1 to 3, 2 to 4, 3 to 6, 1 to 10, 1 to 8, 1 to 6, 2 to 12, 2 to 10, 2 to 8, 2 to 6, 3 to 12, 3 to 10, 3 to 8, 3 to 6, 4 to 12, 4 to 10, 4 to 8, 4 to 6, 5 to 12, 5 to 10, 5 to 8, 5 to 6, 6 to 12, 6 to 10, 6 to 8, 7 to 12, 7 to 10, 7 to 8, 8 to 12, 8 to 10, 9 to 12, 9 to 10, or 10 to 12, wherein the flanking amino acids can be distributed in any ratio to the C-termin
  • said polypeptide with the flanking amino acids is still able to bind to the same HLA molecule (i.e., retain MHC binding propensity) and/or retain the same TCR specificity, and/or retain anti-beta-coronavirus activity, as said polypeptide core sequence without said flanking amino acids.
  • the extension(s) may serve and be designed to improve the biochemical properties of the peptides or polypeptides (e.g., but not limited to, solubility or stability) or to improve the likelihood for efficient proteasomal processing of the peptide.
  • the polypeptides of the present disclosure may be islated, synthetic, and/or recombinant, and may comprise post-transcriptional modifications such as glycosylation, added chemical groups, etc.
  • said flanking amino acid sequences are those that also flank the peptides or polypeptides included therein in the naturally occurring protein (for example, as found in SARS-CoV-2 Wuhan-Hu-1 (GenBank id: MN908947), which was selected as the reference strain).
  • said flanking amino acid sequences are those that also flank the peptides or polypeptides included therein in the naturally occurring protein.
  • flanking amino acid sequences as described herein may serve as a MHC stabilizing region.
  • the use of a longer peptide may allow endogenous processing by patient cells and may lead to more effective antigen presentation and induction of T cell responses.
  • the peptides or polypeptides of the instant disclosure can be isolated, recombinant, and/or synthetic.
  • the peptides or polypeptides can be either in neutral (uncharged) or salt forms, and may be either free of or include modifications such as glycosylation, side chain oxidation, or phosphorylation.
  • the peptides or polypeptides of the instant disclosure can be capped with an n-terminal acetyl and/or c-terminal amino group.
  • the instant disclosure is directed to one or more Class II polypeptides (“clusters”) of Table 1, (and/or fragments or variants thereof), and optionally 1 to 12 additional amino acids distributed in any ratio on the N terminus and/or C-terminus of a polypeptide of Table 1.
  • the instant disclosure is directed to one or more T cell epitope polypeptides (“clusters”) comprising, consisting, or consisting essentially of an amino acid sequence of SEQ ID NOS: 1305, 1616, 1317, 1628, 1333, 1644, 1311, 1622, 1309, 1620, 1330, 1641, 1254, 1565, 1314, 1625, 1272, 1583, 1303, 1614, 1110, 1421, 1099, 1410, 1063, 1374, 1079, 1390,
  • polypeptides may be used/delivered via a micro needle patch, as are known in the art.
  • the instant disclosure is directed to a polypeptide comprising an amino acid sequence having at least 75%, 80%, 85%, 90%, or 95% homology to any one of SEQ ID NOS: 1305, 1317, 1616, 1628, 1333, 1644, 1311, 1622, 1309, 1620, 1330, 1641, 1254, 1565, 1314,
  • the present disclosure is directed to a concatemeric polypeptide or peptide that comprises at one or more of the instantly-disclosed polypeptides or peptides (e.g., but not limited to, a peptide or polypeptide comprising, consisting, or consisting essentially of an amino acid sequence of SEQ ID NOS: 1305, 1317, 1616, 1628, 1333, 1644, 1311, 1622, 1309, 1620, 1330, 1641, 1254, 1565, 1314, 1625, 1272, 1583, 1303, 1614, 1110, 1421, 1099, 1410,
  • a concatemeric polypeptide or peptide that comprises at one or more of the instantly-disclosed polypeptides or peptides (e.g., but not limited to, a peptide or polypeptide comprising, consisting, or consisting essentially of an amino acid sequence of SEQ ID NOS: 1305, 1317, 1616, 1628, 1333, 1644, 1311, 1622
  • additional peptide or polypeptide may be one or more of the instantly instantly-disclosed polypeptides or peptides, or may be an additional peptide or polypeptide of interest.
  • a concatemeric peptide is composed of 3 or more, 4 or more, 5 or more 6 or more 7 or more, 8 or more, 9 or more of the instantly-disclosed peptides or polypeptides.
  • the concatemeric peptides or polypeptides include 1000 or more, 1000 or less, 900 or less, 500 or less, 100 or less, 75 or less, 50 or less, 40 or less, 30 or less, 20 or less or 100 or less peptide epitopes.
  • a concatemeric peptide has 3-100, 5-100, 10-100, 15-100, 20-100, 25-100, 30-100, 35-100, 40-100, 45-100, 50-100, 55-100, 60-100, 65-100, 70-100, 75-100, 80-100, 90-100, 5-50, 10-50, 15-50, 20-50, 25-50, 30-50, 35-50, 40-50, 45-50, 100-150, 100-200, 100-300, 100-400, 100-500, 50-500, 50- 800, 50-1,000, or 100-1,000 of the instantly-disclosed peptides or polypeptides linked, fused, or joined together.
  • Each peptide or polypeptide of the concatemeric polypeptide may optionally have one or more linkers, which may optionally be cleavage sensitive sites, adjacent to their N and/or C terminal end.
  • linkers and cleavage sensitive sites including AAY cleavage motifs or a poly GS linker which may be include on the N terminus of the C-terminal element, are known in the art.
  • two or more of the peptide epitopes may have a linker, which may act as a cleavage sensitive site, between them.
  • two or more of the peptide epitopes may be connected directly to one another or through a linker that is not a cleavage sensitive site.
  • linker is antigenically neutral, and the liker is preferably less than the length of a peptidyl backbone of 9 amino acids linearly arranged.
  • linker length is the length of a peptidyl backbone of between 2 and 8 amino acids, linearly arranged.
  • the spacer is unable to hydrogen bond in any spatially distinct manner to other distinct elements of the enhancing hybrid peptide.
  • a spacer may be composed of alternating units, for example of hydrophobic, lipophilic, aliphatic and aryl-aliphatic sequences, optionally interrupted by heteroatoms such as O, N, or S.
  • Such components of a spacer are preferably chosen from the following classes of compounds: sterols, alkyl alcohols, polyglycerides with varying alkyl functions, alkyl-phenols, alkyl-amines, amides, hydroxyphobic polyoxyalkylenes, and the like.
  • Other examples are hydrophobic polyanhydrides, polyorthoesters, polyphosphazenes, polyhydroxy acids, polycaprolactones, polylactic, polyglycolic polyhydroxy-butyric acids.
  • a linker may also contain repeating short aliphatic chains, such as polypropylene, isopropylene, butylene, isobutylene, pentamethlyene, and the like, separated by oxygen atoms.
  • a linker has a chemical group incorporated within which is subject to cleavage.
  • a chemical group may be designed for cleavage catalyzed by a protease, by a chemical group, or by a catalytic monoclonal antibody.
  • tryptic targets two amino acids with cationic side chains
  • chymotryptic targets with a hydrophobic side chain
  • cathepsin sensitivity B, D or S
  • tryptic target is used herein to describe sequences of amino acids which are recognized by trypsin and trypsin-like enzymes.
  • chymotryptic target is used herein to describe sequences of amino acids which are recognized by chymotrypsin and chymotrypsin-like enzymes.
  • chemical targets of catalytic monoclonal antibodies, and other chemically cleaved groups are well known to persons skilled in the art of peptide synthesis, enzymatic catalysis, and organic chemistry in general, and can be designed into the hybrid structure and synthesized, using routine experimental methods.
  • a concatemeric polypeptide of the instant disclosure is produced using the EpiAssembler System (EpiVax).
  • the EpiAssembler system is useful for assembling overlapping epitopes to Immunogenic Consensus Sequences (ICS).
  • ICS Immunogenic Consensus Sequences
  • EpiAssembler is an algorithm that optimizes the balance between pathogen and population coverage.
  • EpiAssembler uses the information from the sequences produced by conserveatrix and EpiMatrix to form highly immunogenic consensus sequences.
  • the concatemeric peptides of the instant disclosure include those of including one or more of SEQ ID NOS: 1305, 1317, 1616, 1628, 1333, 1644, 1311, 1622, 1309, 1620, 1330, 1641, 1254, 1565, 1314, 1625,
  • RNA mRNA, DNA, cDNA nucleic acids encoding such concatemeric peptides.
  • the present disclosure provides a concatemeric polypeptide with at least 60%, 70%, 80%, 90%, or 95% homology to each of SEQ ID NOS: 1305, 1317, 1616, 1628, 1333, 1644, 1311, 1622, 1309, 1620, 1330, 1641, 1254, 1565, 1314, 1625, 1272,
  • the present disclosure provides a concatemeric polypeptide having anti- beta-coronavirus activity, said polyeptide having at least 60%, 70%, 80%, 90%, or 95% homology to each of SEQ ID NOS: 1305, 1317, 1616, 1628, 1333, 1644, 1311, 1622, 1309, 1620, 1330, 1641, 1254, 1565, 1314, 1625, 1272, 1583, 1303, 1614, 1110, 1421, 1099, 1410,
  • anti-beta- coronavirus activity means that the instantly-disclosed therapeutic T-cell epitope compounds and compositions are, in aspects: capable of stimulating, inducing, and/or expanding an immune response to a beta-coronavirus (e.g., a cellular (CD4+ and/or CD8+ T-cell response) or humoral immune response to SARS-CoV-2 or other beta-coronaviruses) and/or associated diseases in a subject; capable of stimulating, inducing, and/or expanding a beta-coronavirus- specific IFNy response (e.g., by lymphocytes such as PMBC, or effector CD4+ and/or CD8+ T-cells), capable of inhibiting beta-coronavirus viral replication or infectivity, and/or capable of inducing immunity against beta-coronavirus.
  • a beta-coronavirus e.g., a cellular (CD4+ and/or CD8+ T-cell response) or humoral immune response to SARS-Co
  • a T-cell epitope compound or composition of the present disclosure having anti-SARS-CoV-2 activity will reduce the disease symptoms resulting from beta-coronavirus challenge by at least about 5% to about 50%, at least about 10% to about 60%, at least about 30% to about 70%, at least about 40% to about 80%, or at least about 50% to about 90% or greater, including any value or range therebetween.
  • anti- beta-coronavirus activity can be determined by various experiments and assays as known to those of skill in the art, including methods such as by antibody titrations of sera, e.g., by ELISA and/or seroneutralization assay analysis and/or by vaccination challenge evaluation, including use of experiments and assays as disclosed in the Examples herein.
  • the concatemeric polypeptides of the instant disclosure can be isolated, recombinant, and/or synthetic.
  • the concatemeric peptides or polypeptides can be either in neutral (uncharged) or salt forms, and may be either free of or include modifications such as glycosylation, side chain oxidation, or phosphorylation.
  • the concatemeric peptides or polypeptides of the instant disclosure can be capped with an n-terminal acetyl and/or c-terminal amino group.
  • one or more peptides or polypeptides of the instant disclosure e.g., but not limited to, a peptide or polypeptide comprising, consisting, or consisting essentially of an amino acid sequence of SEQ ID NOS: 1305, 1317, 1616, 1628, 1333, 1644, 1311, 1622, 1309, 1620, 1330, 1641, 1254, 1565, 1314, 1625, 1272, 1583, 1303, 1614, 1110, 1421, 1099, 1410,
  • heterologous polypeptide is intended to mean that the one or more T-cell epitopes of the instant disclosure are heterologous to, or not included naturally, in the heterologous polypeptide.
  • a heterologous polypeptide may include, but are not limited to, e.g. monoclonal antibody, polyclonal antibody, mouse antibody, human antibody, humanized antibody, mono specific antibody, bispecific antibody, glycosylated antibody, Fc-modified antibody, or antibody-drug conjugates; an antibody of different class or subclass (e.g., IgG (e.g., IgGl, IgG2, IgG3, IgG4), IgM, IgA, IgD or IgE molecules) or antigen-specific antibody fragments thereof (including, but not limited to, a Fab, F(ab')2, Fv, disulfide linked Fv, scFv, single domain antibody, closed conformation multispecific antibody, disulfide-linked scFv, diabody)).
  • an antibody of different class or subclass e.g., IgG (e.g., IgGl, IgG2, IgG3, IgG4), IgM, I
  • one or more of the instantly-disclosed polypeptides may be inserted into the heterologous polypeptide (e.g., through recombinant techniques, mutagenesis, or other known means in the art), may be added to the C-terminus (with or without the use of linkers, as is known in the art), and/or added to the N-terminus (with or without the use of linkers, as is known in the art) of the heterologous polypeptide.
  • one or more of the instantly-disclosed polypeptides may be inserted into or replace amino acids in a Fc domain as disclosed in U.S. Patent No. 7,442,778, U.S. PatentNo. 7,645,861, U.S. PatentNo.
  • protein engineering by mutagenesis can be performed using site-directed mutagenesis techniques, or other mutagenesis techniques known in the art (see e.g., James A. Brannigan and Anthony J. Wilkinson., 2002, Protein engineering 20 years on. Nature Reviews Molecular Cell Biology 3, 964-970; Turanli-Yildiz B. et ah, 2012, Protein Engineering Methods and Applications, intechopen.com, which are herein incorporated by reference in their entirety).
  • chimeric or fusion polypeptides comprise one or more of the instantly-disclosed polypeptides of the present disclosure operatively linked to a heterologous polypeptide.
  • “Operatively linked” indicates that the one or more of the instantly-disclosed polypeptides and the heterologous protein are fused in-frame or chemically linked or otherwise bound.
  • the one or more of the instantly-disclosed polypeptides may be covalently bound to one or more internal conjugation site(s) in an Fc domain as disclosed in U.S. Patent No. 8,008,453, U.S. Patent No. 9,114,175, and/or U.S. Patent No. 10,188,740 (each of which are herein incorporated by reference in their entirety).
  • the one or more peptides or polypeptides of the instant disclosure e.g., but not limited to, a peptide or polypeptide comprising, consisting, or consisting essentially of an amino acid sequence of SEQ ID NOS: 1305, 1317, 1616, 1628, 1333, 1644, 1311, 1622, 1309, 1620, 1330, 1641, 1254, 1565, 1314,
  • 1116, 1427, 1104, 1415, 1239,1550, 1250, 1561, 1231, 1542, 1100, 1411, 1118, 1429, 1062, 1373, 1086, 1397, 1177, or 1487) may be joined to, linked to (e.g., fused in-frame, chemically- linked, or otherwise bound), and/or inserted into a heterologous polypeptide as a whole, although it may be made up from a joined to, linked to (e.g., fused in-frame, chemically-linked, or otherwise bound), and/or inserted amino acid sequence, together with flanking amino acids of the heterologous polypeptide.
  • polypeptide which, in aspects, may be an isolated, synthetic, or recombinant
  • polypeptide having a sequence comprising one or more of SEQ ID NOS: 1305, 1317, 1616, 1628, 1333, 1644, 1311, 1622, 1309, 1620, 1330, 1641, 1254, 1565, 1314, 1625, 1272, 1583, 1303, 1614, 1110, 1421, 1099,
  • 1090, 1401, 1116, 1427, 1104, 1415, 1239,1550, 1250, 1561, 1231, 1542, 1100, 1411, 1118, 1429, 1062, 1373, 1086, 1397, 1177, or 1487 is not naturally included in the polypeptide and/or said one or more of SEQ ID NOS: 1305, 1317, 1616, 1628, 1333, 1644, 1311, 1622, 1309, 1620, 1330, 1641, 1254, 1565, 1314, 1625, 1272, 1583, 1303, 1614, 1110, 1421, 1099, 1410,
  • the one or more peptides, polypeptides, and/or concatemeric peptides of the present disclosure may be inserted into a SARS-CoV-2 sequence in which the SARS-CoV-2 sequence does not include the one or more peptides, polypeptides, and/or concatemeric peptides of the present disclosure (e.g., the SARS-CoV-2 sequence does not include, or is mutated to not include, the one or more peptides, polypeptides, and/or concatemeric peptides of the present disclosure) or the one or more peptides, polypeptides, and/or concatemeric peptides of the present disclosure is inserted into a SARS-CoV-2 sequence but not at its natural position.
  • the one or more peptides or polypeptides of the instant disclosure can be joined or linked to (e.g., fused in-frame, chemically-linked, or otherwise bound) to a small molecule (e.g., albumin or other known carriers and proteins), drug, or drag fragment, for example but not limited to, a drug or drug fragment that is binds with high affinity to defined HLAs.
  • a small molecule e.g., albumin or other known carriers and proteins
  • drug, or drag fragment for example but not limited to, a drug or drug fragment that is binds with high affinity to defined HLAs.
  • two polypeptides are substantially homologous or identical when the amino acid sequences are at least about 45-55%, typically at least about 70-75%, more typically at least about 80-85%, more typically greater than about 90%, and more typically greater than 95% or more homologous or identical.
  • the sequences are aligned for optimal comparison purposes (e.g. , gaps can be introduced in the sequence of one polypeptide or nucleic acid molecule for optimal alignment with the other polypeptide or nucleic acid molecule).
  • the amino acid residues or nucleotides at corresponding amino acid positions or nucleotide positions are then compared.
  • the percent identity between the two sequences is a function of the number of identical positions shared by the sequences, taking into account the number of gaps, and the length of each gap, which need to be introduced for optimal alignment of the two sequences. Sequence homology for polypeptides is typically measured using sequence analysis software. As used herein, amino acid or nucleic acid "homology” is equivalent to amino acid or nucleic acid “identity”. In aspects, the percent homology between the two sequences is a function of the number of identical positions shared by the sequences ( e.g ., percent homology equals the number of identical positions/total number of positions x 100).
  • the present disclosure also encompasses polypeptides having a lower degree of identity but having sufficient similarity so as to perform one or more of the same functions performed by a polypeptide of the instant disclosure (e.g., a polypeptide having a sequence comprising, consisting of, or consisting essentially of one or more of SEQ ID NOS: 4-1676, 1713-2595, 2605-2638, 2647-2718, and 2735-86921305, 1317, 1616, 1628, 1333, 1644, 1311, 1622, 1309, 1620, 1330, 1641, 1254, 1565, 1314, 1625, 1272, 1583, 1303, 1614, 1110, 1421,
  • conservative substitutions are the replacements, one for another, among the aliphatic amino acids Ala, Val, Leu, Met, and lie; interchange of the hydroxyl residues Ser and Thr, exchange of the acidic residues Asp and Glu, substitution between the amide residues Asn and Gin, exchange of the basic residues His, Lys and Arg and replacements among the aromatic residues Trp, Phe and Tyr.
  • Guidance concerning which amino acid changes are likely to be phenotypically silent are found (Bowie JU et al., (1990), Science, 247(4948):130610, which is herein incorporated by reference in its entirety).
  • a variant polypeptide can differ in amino acid sequence by one or more substitutions, deletions, insertions, inversions, fusions, and truncations or a combination of any of these.
  • Variant polypeptides can be fully functional (e.g., retain MHC binding propensity and/or TCR specificity, and/or retain anti-beta-coronavirus activity) or can lack function in one or more activities.
  • Fully functional variants typically contain only conservative variation or variation in non-critical residues or in non-critical regions; in this case, typically MHC contact residues provided MHC binding is preserved.
  • Functional variants can also contain substitution of similar amino acids that result in no change or an insignificant change in function (e.g., retain MHC binding propensity and/or TCR specificity, and/or retain anti-beta-coronavirus activity). Alternatively, such substitutions can positively or negatively affect function to some degree.
  • Non-functional variants typically contain one or more non-conservative amino acid substitutions, deletions, insertions, inversions, or truncation or a substitution, insertion, inversion, or deletion in a critical residue or critical region; in this case, typically TCR contact residues.
  • a variant and/or a homologous polypeptide retains the desired anti- beta- coronavirus activity of the instant disclsoure (e.g.: capable of stimulating, inducing, and/or expanding an immune response to beta-coronavirus (e.g., a cellular (CD4+ and/or CD8+ T-cell response) or humoral immune response to beta-coronavirus) and/or associated diseases in a subject; capable of stimulating, inducing, and/or expanding a beta-coronavirus-specific IFNy response (e.g., by lymphocytes such as PMBC, or effector CD4+ and/or CD8+ T-cells); and/or capable of inhibiting beta-coronavirus viral replication or infectivity, and/or capable of inducing immunity against beta-coronavirus, optionally all beta-coronaviruses).
  • an immune response to beta-coronavirus e.g., a cellular (CD4+ and/or CD8+ T-cell response
  • Non- functional variants typically contain one or more non-conservative amino acid substitutions, deletions, insertions, inversions, or truncation or a substitution, insertion, inversion, or deletion in a critical residue or critical region; in this case, typically TCR contact residues.
  • functional variants of a polypeptide having a sequence (or a core sequence) comprising, consisting of, or consisting essentially of one or more ofSEQ IDNOS: 1305, 1317, 1616, 1628, 1333, 1644, 1311, 1622, 1309, 1620, 1330, 1641, 1254, 1565, 1314, 1625, 1272, 1583, 1303,
  • 1250, 1561, 1231, 1542, 1100, 1411, 1118, 1429, 1062, 1373, 1086, 1397, 1177, or 1487 as disclosed herein may contain one or more conservative substitutions, and in aspects one or more non-conservative substitutions, at amino acid residues that are not believed to be essential for functioning (with amino acid residues considered being essential for functioning, including, e.g., retain MHC binding propensity and/or TCR specificity, and/or retain anti-beta- coronavirus activity) of the instantly-disclosed polypeptides.
  • a variant polypeptide having a sequence (or a core sequence) comprising, consisting of, or consisting essentially of one or more of SEQ ID NOS: 1305, 1317, 1616, 1628, 1333, 1644, 1311, 1622, 1309, 1620, 1330, 1641, 1254, 1565, 1314, 1625, 1272, 1583, 1303, 1614, 1110, 1421, 1099,
  • MHC binding assays are well known in the art.
  • such assays may include the testing of binding affinity with respect to MHC class I and class II alleles in in vitro binding assays, with such binding assays as are known in the art.
  • Exampels include, e.g., the soluble binding assays as disclosed in U.S.
  • a fully functional variant polypeptide having a sequence (or a core sequence) comprising, consisting of, or consisting essentially of one or more of SEQ ID NOS: 41305, 1317, 1616, 1628, 1333, 1644, 1311, 1622, 1309, 1620, 1330, 1641, 1254, 1565, 1314, 1625, 1272, 1583, 1303, 1614, 1110, 1421, 1099, 1410, 1063,
  • the TCR-binding epitope (which can be referred to as TCR binding residues, TCR facing epitope, TCR facing residues, or TCR contacts) for a 9-mer identified epitope (which may be a 9-mer fragment of one or more of SEQ ID NOS: 1305, 1317, 1616, 1628,
  • the TCR binding epitope for a 9-mer identified epitope (which may be a 9- mer fragment of one or more of SEQ ID NOS1305, 1317, 1616, 1628, 1333, 1644, 1311, 1622, 1309, 1620, 1330, 1641, 1254, 1565, 1314, 1625, 1272, 1583, 1303, 1614, 1110, 1421, 1099,
  • the TCR binding epitope for a 10-mer identified epitope that bind to a MHC class I molecule are at position 4, 5, 6, 7, 8, and 9 of the identified epitope (which may be a 10- mer fragment of one or more of SEQ ID NOS: 1305, 1317, 1616, 1628, 1333, 1644, 1311, 1622, 1309, 1620, 1330, 1641, 1254, 1565, 1314, 1625, 1272, 1583, 1303, 1614, 1110, 1421,
  • MHC binding agretope for a 10-mer identified epitope which may be a 10-mer fragment of one or more of SEQ ID NOS: 1305, 1317, 1616, 1628, 1333, 1644, 1311, 1622,
  • the TCR-binding epitope for a 9-mer identified epitope (which may be a 9- mer fragment of one or more of SEQ ID NOS: 1305, 1317, 1616, 1628, 1333, 1644, 1311, 1622, 1309, 1620, 1330, 1641, 1254, 1565, 1314, 1625, 1272, 1583, 1303, 1614, 1110, 1421,
  • the TCR binding epitope for 9-mer identified epitope (which may be a 9- mer fragment of one or more of SEQ ID NOS: 1305, 1317, 1616, 1628, 1333, 1644, 1311, 1622, 1309, 1620, 1330, 1641, 1254, 1565, 1314, 1625, 1272, 1583, 1303, 1614, 1110, 1421,
  • the TCR-binding epitope for a 10-mer identified epitope (which may be a 10-mer fragment of one or more of SEQ ID NOS: 1305, 1317, 1616, 1628, 1333, 1644, 1311, 1622, 1309, 1620, 1330, 1641, 1254, 1565, 1314, 1625, 1272, 1583, 1303, 1614, 1110, 1421,
  • any given amino acid may be, with respect to a given 9-mer epitope or 10-mer epitope, MHC facing and, with respect to another 9-mer epitope, TCR facing.
  • the present disclosure also includes fragments of the instantly-disclosed polypeptides and concatemeric polypeptides. In aspects, the present disclosure also encompasses fragments of the variants of the instantly-disclosed polypeptides and concatemeric polypeptides as described herein. In aspects, as used herein, a fragment comprises at least about nine contiguous amino acids. In aspects, the present disclosure also encompasses fragments of the variants of the T-cell epitopes described herein. Useful fragments (and fragments of the variants of the polypeptides and concatemeric polypeptides described herein) include those that retain one or more of the biological activities, particularly: MHC binding propensity and/or TCR specificity, and/or anti-SARS-CoV-2 activity.
  • Biologically active fragments are, for example, about 9, 10, 11, 12, 1, 14, 15, 16, 20, 25, 30, 40, 50, 60, 70, 80, 90, 100 or more amino acids in length, including any value or range therebetween. Fragments can be discrete (not fused to other amino acids or polypeptides) or can be within a larger polypeptide. Several fragments can be comprised within a single larger polypeptide. In aspects, a fragment designed for expression in a host can have heterologous pre- and pro-polypeptide regions fused to the amino terminus of the polypeptide fragment and an additional region fused to the carboxyl terminus of the fragment.
  • the instantly disclosed polypeptides and concatemeric polypeptides of the present disclosure can include allelic or sequence variants (“mutants”) or analogs thereof, or can include chemical modifications (e.g., pegylation, glycosylation).
  • a mutant retains the same function, particularly MHC binding propensity and/or TCR specificity, and/or anti-beta-coronavirus activity.
  • a mutant can provide for enhanced binding to MHC molecules.
  • a mutant can lead to enhanced binding to TCRs.
  • a mutant can lead to a decrease in binding to MHC molecules and/or TCRs.
  • polypeptides of the present disclosure will vary widely, depending upon the nature of the various elements comprising the molecule.
  • an isolated polypeptide can be purified from cells that naturally express it, purified from cells that have been altered to express it (recombinant), or synthesized using known protein synthesis methods.
  • the synthetic procedures may be selected so as to be simple, provide for high yields, and allow for a highly purified stable product.
  • polypeptides of the instant disclosure can be produced either from a nucleic acid disclosed herein, or by the use of standard molecular biology techniques, such as recombinant techniques, mutagenesis, or other known means in the art.
  • An isolated polypeptide can be purified from cells that naturally express it, purified from cells that have been altered to express it (recombinant), or synthesized using known protein synthesis techniques.
  • a polypeptide of the instant disclosure is produced by recombinant DNA or RNA techniques.
  • a polypeptide of the instant disclosure can be produced by expression of a recombinant nucleic acid of the instant disclosure in an appropriate host cell. For example, a nucleic acid molecule encoding the polypeptide is cloned into an expression cassette or expression vector, the expression cassette or expression vector introduced into a host cell and the polypeptide expressed in the host cell. The polypeptide can then be isolated from the cells by an appropriate purification scheme using standard protein purification techniques.
  • polypeptide can be produced by a combination of ex vivo procedures, such as protease digestion and purification.
  • polypeptides of the instant disclosure can be produced using site-directed mutagenesis techniques, or other mutagenesis techniques known in the art (see e.g., James A. Brannigan and Anthony J. Wilkinson., 2002, Protein engineering 20 years on. Nature Reviews Molecular Cell Biology 3, 964-970; Turanli-Yildiz B. et al., 2012, Protein Engineering Methods and Applications, intechopen.com, which are herein incorporated by reference in their entirety).
  • the present disclosure also provides chimeric or fusion polypeptide compositions.
  • the present disclosure is directed to a chimeric or fusion polypeptide composition (which in aspects may be isolated, synthetic, or recombinant) comprising one or more peptides, polypeptides, or concatemeric peptides of the present disclosure (e.g., one or more peptides or polypeptides of the present disclosure have a sequence, e.g. but not limited to, comprising, consisting, or consisting essentially of an amino acid sequence of SEQ ID NOS: 1305, 1317, 1616, 1628, 1333, 1644, 1311, 1622, 1309, 1620, 1330, 1641, 1254, 1565, 1314,
  • a chimeric or fusion polypeptide composition of the present disclosure comprises one or more peptides, polypeptides, and/or concatemeric peptides of the present disclosure joined to, linked to (e.g., fused in-frame, chemically-linked, or otherwise bound), and/or inserted into a heterologous polypeptide, such as an unrelated protein.
  • heterologous polypeptide is intended to mean that the one or more T- cell epitopes of the instant disclosure are heterologous to, or not included naturally, in the heterologous polypeptide.
  • a heterologous polypeptide may include, but are not limited to, e.g.
  • monoclonal antibody polyclonal antibody, mouse antibody, human antibody, humanized antibody, mono specific antibody, bispecific antibody, glycosylated antibody, Fc- modified antibody, or antibody-drug conjugates; an antibody of different class or subclass (e.g., IgG (e.g., IgGl, IgG2, IgG3, IgG4), IgM, IgA, IgD or IgE molecules) or antigen-specific antibody fragments thereof (including, but not limited to, a Fab, F(ab')2, Fv, disulfide linked Fv, scFv, single domain antibody, closed conformation multispecific antibody, disulfide-linked scFv, diabody)).
  • IgG e.g., IgGl, IgG2, IgG3, IgG4
  • IgM IgA, IgD or IgE molecules
  • antigen-specific antibody fragments thereof including, but not limited to, a Fab, F(ab'
  • one or more of the instantly-disclosed peptides, polypeptides, or concatemeric peptides may be inserted into the heterologous polypeptide (e.g., through recombinant techniques, mutagenesis, or other known means in the art), may be added to the C-terminus (with or without the use of linkers, as is known in the art), and/or added to the N- terminus (with or without the use of linkers, as is known in the art) of the heterologous polypeptide.
  • one or more of the instantly-disclosed polypeptides may be inserted into or replace amino acids in a Fc domain as disclosed in U.S. Patent No. 7,442,778, U.S. PatentNo.
  • chimeric or fusion polypeptides comprise one or more of the instantly-disclosed peptides, polypeptides, or concatemeric peptides operatively linked to a heterologous polypeptide.
  • "Operatively linked” indicates that the one or more of the instantly-disclosed peptides, polypeptides, or concatemeric peptides and the heterologous polypeptide are fused in-frame or chemically-linked or otherwise bound.
  • the one or more of the instantly-disclosed polypeptides may be covalently bound to one or more internal conjugation site(s) in an Fc domain as disclosed in U.S. Patent No. 8,008,453, U.S. Patent No.
  • the one or more peptides, polypeptides, or concatemeric peptides of the instant disclosure may be joined to, linked to (e.g., fused in- frame, chemically- linked, or otherwise bound), and/or inserted into a heterologous polypeptide as a whole, although it may be made up from a joined to, linked to (e.g., fused in-frame, chemically-linked, or otherwise bound), and/or inserted amino acid sequence, together with flanking amino acids of the heterologous polypeptide.
  • a chimeric or fusion polypeptide composition comprises a peptide, polypeptide, or concatemeric peptide of the instant disclosure wherein said one or more of peptides, polypeptides, or concatemeric peptides is not naturally included in the heterologous polypeptide and/or said one or more of peptides, polypeptides, or concatemeric peptides is not located at its natural position in the heterologous polypeptide.
  • the one or more of peptide or polypeptides of the present disclosure can be joined, linked to (e.g., fused in-frame, chemically-linked, or otherwise bound), and/or inserted into the heterologous polypeptide.
  • chimeric or fusion polypeptide compositions comprise one or more of the instantly-disclosed T-cell epitopes (e.g., peptides, polypeptides, or concatemeric peptides of the instant disclosure) operatively linked to a heterologous polypeptide having an amino acid sequence not substantially homologous to the T-cell epitope.
  • the chimeric or fusion polypeptide does not affect function of the T-cell epitope per se.
  • the fusion polypeptide can be a GST-fusion polypeptide in which the T-cell epitope sequences are fused to the C-terminus of the GST sequences.
  • fusion polypeptides include, but are not limited to, enzymatic fusion polypeptides, for example beta- galactosidase fusions, yeast two-hybrid GAL fusions, poly-His fusions and Ig fusions.
  • fusion polypeptides particularly poly-His fusions or affinity tag fusions, can facilitate the purification of recombinant polypeptide.
  • expression and/or secretion of a polypeptide can be increased by using a heterologous signal sequence. Therefore, in aspects, the chimeric or fusion polypeptide contains a heterologous signal sequence at its N-terminus.
  • the heterologous polypeptide or polypeptide comprises a biologically active molecule.
  • the biologically active molecule is selected from the group consisting of an immunogenic molecule, a T cell epitope, a viral protein, and a bacterial protein.
  • the one or more peptides, polypeptides, or concatemeric peptides of the instant disclosure can be joined or linked to (e.g., fused in-frame, chemically-linked, or otherwise bound) to a small molecule, drug, or drug fragment.
  • the one or more peptides, polypeptides, or concatemeric peptides of the instant disclosure can be joined or linked to (e.g., fused in-frame, chemically-linked, or otherwise bound) to an unrelated peptide or protein, a small molecule (e.g., albumin or other known carriers and proteins), drug, or drag fragment, for example but not limited to, a drug or drug fragment that is binds with high affinity to defined HLAs.
  • the chimeric or fusion polypeptide compositions can be recombinant, isolated, and/or synthetic.
  • a chimeric or fusion polypeptide composition can be produced by standard recombinant DNA or RNA techniques as are known in the art. For example, DNA or RNA fragments coding for the different polypeptide sequences may be ligated together in-frame in accordance with conventional techniques. In another embodiment, the fusion gene can be synthesized by conventional techniques including automated DNA synthesizers.
  • PCR polymerase chain reaction
  • anchor primers which give rise to complementary overhangs between two consecutive nucleic acid fragments which can subsequently be annealed and re-amplified to generate a chimeric nucleic acid sequence
  • one or more peptides, polypeptides or concatemeric of the instant disclosure e.g., but not limited to, a peptide or polypeptide comprising, consisting, or consisting essentially of an amino acid sequence of SEQ ID NOS: 1305, 1317, 1616, 1628, 1333, 1644, 1311, 1622, 1309, 1620, 1330, 1641, 1254, 1565, 1314,
  • 1116, 1427, 1104, 1415, 1239,1550, 1250, 1561, 1231, 1542, 1100, 1411, 1118, 1429, 1062, 1373, 1086, 1397, 1177, or 1487) can be inserted into a heterologous polypeptide or inserted into a non-naturally occurring position of a polypeptide through recombinant techniques, synthetic polymerization techniques, mutagenesis, or other standard techniques known in the art.
  • protein engineering by mutagenesis can be performed using site-directed mutagenesis techniques, or other mutagenesis techniques known in the art (see e.g., James A. Brannigan and Anthony J. Wilkinson., 2002, Protein engineering 20 years on. Nature Reviews Molecular Cell Biology 3, 964-970; Turanli-Yildiz B. et ah, 2012, Protein Engineering Methods and Applications, intechopen.com, which are herein incorporated by reference in their entirety).
  • polypeptides, concatemeric polypeptides, and chimeric or fusion polypeptides can be purified to homogeneity or partially purified. It is understood, however, that preparations in which the T-cell epitope compounds and compositions are not purified to homogeneity are useful. The critical feature is that the preparation allows for the desired function of the composition, even in the presence of considerable amounts of other components. Thus, the present disclosure encompasses various degrees of purity.
  • the language “substantially free of cellular material” includes preparations of the polypeptides, concatemeric polypeptides, and chimeric or fusion polypeptides having less than about 30% (by dry weight) other proteins (e.g ., contaminating protein), less than about 20% other proteins, less than about 10% other proteins, less than about 5% other proteins, less than about 4% other proteins, less than about 3% other proteins, less than about 2% other proteins, less than about 1 % other proteins, or any value or range therebetween.
  • other proteins e.g ., contaminating protein
  • the composition can also be substantially free of culture medium, for example, culture medium represents less than about 20%, less than about 10%, or less than about 5% of the volume of the the polypeptides, concatemeric polypeptides, and chimeric or fusion polypeptides preparation.
  • culture medium represents less than about 20%, less than about 10%, or less than about 5% of the volume of the the polypeptides, concatemeric polypeptides, and chimeric or fusion polypeptides preparation.
  • substantially free of chemical precursors or other chemicals includes preparations of the the polypeptides, concatemeric polypeptides, and chimeric or fusion polypeptides in which it is separated from chemical precursors or other chemicals that are involved in the T-cell epitope’s synthesis.
  • substantially free of chemical precursors or other chemicals can include, for example, preparations of the the polypeptides, concatemeric polypeptides, and chimeric or fusion polypeptides having less than about 30% (by dry weight) chemical precursors or other chemicals, less than about 20% chemical precursors or other chemicals, less than about 10% chemical precursors or other chemicals, less than about 5% chemical precursors or other chemicals, less than about 4% chemical precursors or other chemicals, less than about 3% chemical precursors or other chemicals, less than about 2% chemical precursors or other chemicals, or less than about 1% chemical precursors or other chemicals.
  • the present disclosure also includes pharmaceutically acceptable salts of the T-cell epitope compounds and compositions (including one or more of e.g., peptides or polypeptides as disclosed herein; concatemeric peptides as disclosed herein; chimeric or fusion polypeptide compositions as disclosed herein (which in aspects may be isolated, synthetic, and/or recombinant).
  • “Pharmaceutically acceptable salt” means a salt that is pharmaceutically acceptable and that possesses the desired pharmacological activity of the parent peptide or polypeptide (e.g., peptides, polypeptides, concatemeric peptides, and/or chimeric or fusion polypeptides as disclosed herein).
  • pharmaceutically acceptable salt refers to derivative of the instantly-disclosed polypeptides, concatemeric polypeptides, and/or chimeric or fusion polypeptides, wherein such compounds are modified by making acid or base salts thereof.
  • pharmaceutically acceptable salts include, but are not limited to, mineral or organic acid salts of basic residues such as amines, alkali or organic salts of acidic residues such as carboxylic acids, and the like.
  • the pharmaceutically acceptable salts include the conventional non-toxic salts or the quaternary ammonium salts of the parent compound formed, for example, from non-toxic inorganic or organic acids.
  • such conventional nontoxic salts include, but are not limited to, those derived from inorganic and organic acids selected from 2-acetoxybenzoic, 2-hydroxyethane sulfonic, acetic, ascorbic, benzene sulfonic, benzoic, bicarbonic, carbonic, citric, edetic, ethane disulfonic, 1,2-ethane sulfonic, fumaric, glucoheptonic, gluconic, glutamic, glycolic, glycollyarsanilic, hexylresorcinic, hydrabamic, hydrobromic, hydrochloric, hydroiodic, hydroxymaleic, hydroxynaphthoic, isethionic, lactic, lactobionic, lauryl sulfonic, maleic, malic, mandelic, methane sulfonic, napsylic, nitric, oxalic, pamoic, pantothenic, phenylacetic, phosphoric
  • the present disclosure also provides for nucleic acids (e.g., DNAs (including cDNA, RNAs (such as, but limited to mRNA), vectors, viruses, or hybrids thereof, all of which may be isolated, synthetic, or recombinant) that encode in whole or in part one or more one or more peptides, polypeptides, concatemeric peptides, and/or chimeric or fusion polypeptides of the present disclosure as described herein.
  • DNAs including cDNA, RNAs (such as, but limited to mRNA), vectors, viruses, or hybrids thereof, all of which may be isolated, synthetic, or recombinant
  • the nucleic acid further comprises, or is contained within, an expression cassette, a plasmid, and expression vector, or recombinant virus, wherein optionally the nucleic acid, or the expression cassette, plasmid, expression vector, or recombinant virus is contained within a cell, optionally a human cell or a non-human cell, and optionally the cell is transformed with the nucleic acid, or the expression cassette, plasmid, expression vector, or recombinant virus.
  • cells are transduced, transfected, or otherwise engineered to contain within one or more of e.g., polypeptides of the present disclosure; isolated, synthetic, or recombinant nucleic acids, expression cassettes, plasmids, expression vectors, or recombinant viruses as disclosed herein; and/or isolated, synthetic, or recombinant chimeric or fusion polypeptide compositions as disclosed herein.
  • the cell can be a mammalian cell, bacterial cell, insect cell, or yeast cell.
  • the nucleic acid molecules of the present disclosure can be inserted into vectors and used, for example, as expression vectors or gene therapy vectors.
  • Gene therapy vectors can be delivered to a subject by, e.g., intravenous injection, local administration (U.S. Pat. No. 5,328,470) or by stereotactic injection (Chen SH et al., (1994), Proc Natl Acad Sci USA, 91(8):3054-7, which are herein incorporated by reference in their entirety).
  • the nucleic acid molecules of the present disclosure can be inserted into plasmids.
  • the pharmaceutical preparation of the gene therapy vector can include the gene therapy vector in an acceptable diluent, or can comprise a slow release matrix in which the gene delivery vehicle is imbedded.
  • the pharmaceutical preparation can include one or more cells that produce the gene delivery system.
  • Such pharmaceutical compositions can be included in a container, pack, or dispenser together with instructions for administration.
  • nucleic acids e.g., DNAs, RNAs, vectors, viruses, or hybrids thereof
  • the nucleic acids encode one or more peptides or polypeptides of the instant disclosure as described above (e.g., but not limited to, a peptide or polypeptide comprising, consisting, or consisting essentially of an amino acid sequence of SEQ ID NOS: 1305, 1317, 1616, 1628, 1333, 1644, 1311, 1622, 1309, 1620, 1330, 1641, 1254, 1565, 1314,
  • the present disclosure is directed to a vector comprising a nucleic acid of the present disclosure encoding one or more polypeptides of the present disclosure or chimeric or fusion polypeptide composition of the present disclosure.
  • the present disclosure is directed to a cell comprising a vector of the present disclosure.
  • the cell can be a mammalian cell, bacterial cell, insect cell, or yeast cell.
  • the nucleic acid of the instant disclosure may be DNAs (including but not limited to cDNA) or RNAs (including but not limited to mRNA), single- or double-stranded.
  • the nucleic acid is typically DNA or RNA (including mRNA).
  • the nucleic acid may be produced by techniques well known in the art, such as synthesis, or cloning, or amplification of the sequence encoding the immunogenic polypeptide; synthesis, or cloning, or amplification of the sequence encoding the cell membrane addressing sequence; ligation of the sequences and their cloning/amplification in appropriate vectors and cells.
  • nucleic acids provided herein that encode in whole or in part one or more peptides, polypeptides, concatemeric peptides, and/or chimeric or fusion polypeptides as described herein can be isolated from a variety of sources, genetically engineered, amplified, synthetically produced, and/or expressed/generated recombinantly. Recombinant polypeptides generated from these nucleic acids can be individually isolated or cloned and tested for a desired activity. Any recombinant expression system can be used, including e.g. in vitro, bacterial, fungal, mammalian, yeast, insect or plant cell expression systems.
  • nucleic acids provided herein are synthesized in vitro by well-known chemical synthesis techniques (as described in, e.g., Adams (1983) J. Am. Chem. Soc. 105:661; Belousov (1997) Nucleic Acids Res. 25:3440-3444; Frenkel (1995) Free Radic. Biol. Med. 19:373-380; Blommers (1994) Biochemistry 33:7886-7896; Narang (1979) Meth. Enzymol. 68:90; Brown (1979) Meth. Enzymol. 68:109; Beaucage (1981) Tetra. Lett. 22:1859; U.S. Pat. No. 4,458,066, all of which are herein incorporated by reference in their entirety).
  • nucleic acids provided herein, such as, e.g., subcloning, labeling probes (e.g., random-primer labeling using Klenow polymerase, nick translation, amplification), sequencing, hybridization and the like are well described in the scientific and patent literature (see, e.g., Sambrook, ed., MOLECULAR CLONING: A LABORATORY MANUAL (2ND ED.), Vols. 1-3, Cold Spring Harbor Laboratory, (1989); CURRENT PROTOCOLS IN MOLECULAR BIOLOGY, Ausubel, ed.
  • a further object of the invention relates to a nucleic acid molecule encoding one or more peptides, polypeptides, concatemeric peptides, and/or chimeric or fusion polypeptides as described herein.
  • the nucleic acid may be used to produce the one or more peptides, polypeptides, concatemeric peptides, and/or chimeric or fusion polypeptides as described herein in vitro or in vivo, or to produce cells expressing the polypeptide on their surface, or to produce vaccines wherein the active agent is the nucleic acid or a vector containing the nucleic acid.
  • the nucleic acid may be, e.g., DNA, cDNA, PNA, CNA, RNA, either single- and/or double-stranded, or native or stabilized forms of polynucleotides as are known in the art.
  • nucleic acid molecules according to the present disclosure may be provided in the form of a nucleic acid molecule per se such as naked nucleic acid molecules; a plasmid, a vector; virus or host cell, etc., either from prokaryotic or eukaryotic origin.
  • Vectors include expression vectors that contain a nucleic acid molecule of the invention.
  • An expression vector capable of expressing a polypeptide can be prepared. Expression vectors for different cell types are well known in the art and can be selected without undue experimentation.
  • the (e.g., cDNA, or RNA, including mRNA) is inserted into an expression vector, such as a plasmid, in proper orientation and correct reading frame for expression.
  • the DNA may be linked to the appropriate transcriptional and translational regulatory control nucleotide sequences recognized by the desired host (e.g., bacteria), although such controls are generally available in the expression vector.
  • the vector is then introduced into the host bacteria for cloning using standard techniques.
  • the vectors of the present invention may, for example, comprise a transcriptional promoter, and/or a transcriptional terminator, wherein the promoter is operably linked with the nucleic acid molecule, and wherein the nucleic acid molecule is operably linked with the transcription terminator.
  • One or more peptides or polypeptides of the present disclosure may be encoded by a single expression vector.
  • Such nucleic acid molecules may act as vehicles for delivering peptides/polypeptides to the subject in need thereof, in vivo, in the form of, e.g., DNA/RNA vaccines.
  • the vector may be a viral vector comprising a nucleic acid as defined above.
  • the viral vector may be derived from different types of viruses, such as, Swinepox, Fowlpox, Pseudorabies, Aujezky's virus, salmonella, vaccinia virus, BHV (Bovine Herpes Virus), HVT (Herpes Virus of Turkey), adenovirus, TGEV (Transmissible Gastroenteritidis Coronavirus), Erythrovirus, and SIV (Simian Immunodeficiency Virus).
  • Other expression systems and vectors may be used as well, such as plasmids that replicate and/or integrate in yeast cells.
  • the instant disclosure also relates to a method for preparing a peptide, polypeptide, concatemeric peptide, and/or chimeric or fusion polypeptide of the instant disclosure, the method comprising culturing a host cell containing a nucleic acid or vector as defined above under conditions suitable for expression of the nucleic acid and recovering the polypeptide.
  • the proteins and peptides may be purified according to techniques known per se in the art.
  • the T cell epitope compositions of the present disclosure may be comprised in a pharmaceutical composition or formulation.
  • the instantly-disclosed pharmaceutical compositions or formulations generally comprise a T-cell epitope composition of the present disclosure and a pharmaceutically- acceptable carrier and/or excipient.
  • a pharmaceutical composition or formulation comprises an adjuvant.
  • said pharmaceutical compositions are suitable for administration.
  • Pharmaceutically-acceptable carriers and/or excipients are determined in part by the particular composition being administered, as well as by the particular method used to administer the composition. Accordingly, there is a wide variety of suitable formulations of pharmaceutical compositions for administering the instantly-disclosed T-cell epitope compositions (see, e.g., Remington’s Pharmaceutical Sciences. (18 TH Ed, 1990), Mack Publishing Co., Easton, PA Publ)).
  • the pharmaceutical compositions are generally formulated as sterile, substantially isotonic, and in full compliance with all Good Manufacturing Practice (GMP) regulations of the U.S. Food and Drug Administration.
  • Pharmaceutical compositions as disclosed herein are able for use in stimulating, inducing, and/or expanding an immune response to a SARS-CoV-2 infection and/or related diseases caused by SARS-CoV-2, including COVID-19, in a subject, and can be used in methods of treating and/or preventing SARS-CoV-2 infection and/or related diseases caused by SARS- CoV-2, including COVID-19, in a subject, such as a human.
  • compositions, carriers, excipients, and reagents are used interchangeably and represent that the materials are capable of administration to or upon a subject without the production of undesirable physiological effects to a degree that would prohibit administration of the composition.
  • pharmaceutically-acceptable excipient means, for example, an excipient that is useful in preparing a pharmaceutical composition that is generally safe, non-toxic, and desirable, and includes excipients that are acceptable for veterinary use as well as for human pharmaceutical use.
  • excipients can be solid, liquid, semisolid, or, in the case of an aerosol composition, gaseous.
  • a person of ordinary skill in the art would be able to determine the appropriate timing, sequence and dosages of administration for particular T-cell epitope compositions of the present disclosure.
  • preferred examples of such carriers or diluents include, but are not limited to, water, saline, Ringer's solutions, dextrose solution, and 5% human serum albumin.
  • Liposomes and non-aqueous vehicles such as fixed oils can also be used.
  • the use of such media and compounds for pharmaceutically active substances is well known in the art. Except insofar as any conventional media or compound is incompatible with the T-cell epitope compounds and compositions of the present disclosure and as previously described above, use thereof in the compositions is contemplated. Supplementary active compounds can also be incorporated into the compositions.
  • T-cell epitope compounds and compositions of the present disclosure are formulated to be compatible with its intended route of administration.
  • the T-cell epitope compounds and compositions of the present disclosure can be administered by parenteral, topical, intravenous, oral, subcutaneous, intraarterial, intradermal, transdermal, rectal, intracranial, intrathecal, intraperitoneal, intranasal; vaginally; intramuscular route or as inhalants.
  • T-cell epitope compounds and compositions of the present disclosure can be injected directly into a particular tissue where deposits have accumulated, e.g., intracranial injection.
  • intramuscular injection or intravenous infusion may be used for administration of T-cell epitope compounds and compositions of the present disclosure.
  • T-cell epitope compounds and compositions of the present disclosure are administered as a sustained release composition or device, such as but not limited to a MedipadTM device.
  • T-cell epitope compounds and compositions of the present disclosure are administered intradermally, e.g., by using a commercial needle-free high- pressure device such as Pulse NeedleFree technology (Pulse 50TM Micro Dose Injection System, Pulse NeedleFree Systems; Lenexa, KS, USA).
  • Pulse NeedleFree technology Pulse 50TM Micro Dose Injection System, Pulse NeedleFree Systems; Lenexa, KS, USA.
  • said commercial needle- free high-pressure device confers one or more of the following benefits: non-invasive, reduces tissue trauma, reduces pain, requires a smaller opening in the dermal layer to deposit the composition in the subject (e.g., only requires a micro skin opening), instant dispersion of the composition, better absorption of the composition, greater dermal exposure to the composition, and/or reduced risk of sharps injury.
  • T cell epitope compounds and compositions of the present disclosure can optionally be administered in combination with other agents that are at least partly effective in treating various medical conditions as described herein.
  • solutions or suspensions used for parenteral, intradermal, or subcutaneous application can include, but are not limited to, the following components: a sterile diluent such as water for injection, saline solution, fixed oils, polyethylene glycols, glycerine, propylene glycol or other synthetic solvents; antibacterial compounds such as benzyl alcohol or methyl parabens; antioxidants such as ascorbic acid or sodium bisulfite; chelating compounds such as ethylenediaminetetraacetic acid (EDTA); buffers such as acetates, citrates or phosphates, and compounds for the adjustment of tonicity such as sodium chloride or dextrose.
  • a sterile diluent such as water for injection, saline solution, fixed oils, polyethylene glycols, glycerine, propylene glycol or other synthetic solvents
  • antibacterial compounds such as benzyl alcohol or methyl parabens
  • antioxidants such as ascorbic acid or sodium bisulfit
  • the pH can be adjusted with acids or bases, such as hydrochloric acid or sodium hydroxide.
  • excipients can include starch, glucose, lactose, sucrose, gelatin, malt, rice, flour, chalk, silica gel, water, ethanol, DMSO, glycol, propylene, dried skim milk, and the like.
  • the composition can also contain pH buffering reagents, and wetting or emulsifying agents.
  • compositions or formulations suitable for injectable use include sterile aqueous solutions (where water soluble) or dispersions and sterile powders for the extemporaneous preparation of sterile injectable solutions or dispersion.
  • suitable carriers include physiological saline, bacteriostatic water, Cremophor ELTM (BASF, Parsippany, NJ) or phosphate buffered saline (PBS).
  • the composition is sterile and should be fluid to the extent that easy syringeability exists. It is stable under the conditions of manufacture and storage and is preserved against the contaminating action of microorganisms such as bacteria and fungi.
  • formulations including a T-cell epitope compound or composition of the present disclosure may include aggregates, fragments, breakdown products and post-translational modifications, to the extent these impurities bind HLA and present the same TCR face to cognate T cells they are expected to function in a similar fashion to pure T-cell epitopes.
  • the carrier can be a solvent or dispersion medium containing, e.g., water, ethanol, polyol (e.g., glycerol, propylene glycol, and liquid polyethylene glycol, and the like), and suitable mixtures thereof.
  • the proper fluidity can be maintained, e.g., by the use of a coating such as lecithin, by the maintenance of the required particle size in the case of dispersion and by the use of surfactants.
  • Prevention of the action of microorganisms can be achieved by various antibacterial and antifungal compounds, e.g., parabens, chlorobutanol, phenol, ascorbic acid, thimerosal, and the like.
  • isotonic compounds e.g., sugars, polyalcohols such as manitol, sorbitol, and sodium chloride in the composition.
  • Prolonged absorption of the injectable compositions can be brought about by including in the composition a compound that delays absorption, e.g., aluminum monostearate and gelatin.
  • sterile injectable solutions e.g., sterile solutions suitable for injectable and/or intradermal needle-free high-pressure device
  • sterile injectable solutions can be prepared by incorporating the T- cell epitope compounds and compositions of the present disclosure in the required amount in an appropriate solvent with one or a combination of ingredients enumerated above, as required, followed by filtered sterilization.
  • dispersions are prepared by incorporating the binding agent into a sterile vehicle that contains a basic dispersion medium and the required other ingredients from those enumerated above.
  • T-cell epitope compounds and compositions of the present disclosure can be administered in the form of a depot injection or implant preparation that can be formulated in such a manner as to permit a sustained or pulsatile release of the active ingredient.
  • oral compositions generally include an inert diluent or an edible carrier and can be enclosed in gelatin capsules or compressed into tablets.
  • the binding agent can be incorporated with excipients and used in the form of tablets, troches, or capsules.
  • Oral compositions can also be prepared using a fluid carrier for use as a mouthwash, wherein the compound in the fluid carrier is applied orally and swished and expectorated or swallowed.
  • Pharmaceutically compatible binding compounds, and/or adjuvant materials can be included as part of the composition.
  • the tablets, pills, capsules, troches and the like can contain any of the following ingredients, or compounds of a similar nature: a binder such as microcrystalline cellulose, gum tragacanth or gelatin; an excipient such as starch or lactose, a disintegrating compound such as alginic acid, PRIMOGEL or corn starch; a lubricant such as magnesium stearate or sterotes; a glidant such as colloidal silicon dioxide; a sweetening compound such as sucrose or saccharin; or a flavoring compound such as peppermint, methyl salicylate or orange flavoring.
  • a binder such as microcrystalline cellulose, gum tragacanth or gelatin
  • an excipient such as starch or lactose, a disintegrating compound such as alginic acid, PRIMOGEL or corn starch
  • a lubricant such as magnesium stearate or sterotes
  • a glidant such as colloidal silicon dioxide
  • T-cell epitope compounds and compositions of the present disclosure can be delivered in the form of an aerosol spray from pressured container or dispenser that contains a suitable propellant, e.g., a gas such as carbon dioxide, or a nebulizer.
  • a suitable propellant e.g., a gas such as carbon dioxide, or a nebulizer.
  • systemic administration of the T-cell epitope compounds and compositions of the present disclosure can also be by transmucosal or transdermal means.
  • penetrants appropriate to the barrier to be permeated are used in the formulation.
  • penetrants are generally known in the art, and include, e.g., for transmucosal administration, detergents, bile salts, and fusidic acid derivatives.
  • Transmucosal administration can be accomplished through the use of nasal sprays or suppositories.
  • the T-cell epitope compounds and compositions may be formulated into ointments, salves, gels, or creams and applied either topically or through transdermal patch technology as generally known in the art.
  • the T-cell epitope compounds and compositions of the present disclosure can also be prepared in the form of suppositories (e.g., with conventional suppository bases such as cocoa butter and other glycerides) or retention enemas for rectal delivery.
  • suppositories e.g., with conventional suppository bases such as cocoa butter and other glycerides
  • retention enemas for rectal delivery.
  • the T-cell epitope compounds and compositions of the present are prepared with carriers that protect the T-cell epitope compounds and compositions against rapid elimination from the body, such as a controlled-release formulation, including implants and microencapsulated delivery systems.
  • a controlled-release formulation including implants and microencapsulated delivery systems.
  • Biodegradable, biocompatible polymers can be used, such as, for example, ethylene vinyl acetate, polyanhydrides, polyglycolic acid, collagen, polyorthoesters, and polylactic acid. Methods for preparation of such formulations will be apparent to those skilled in the art.
  • the materials can also be obtained commercially, e.g., from Alza Corporation and Nova Pharmaceuticals, Inc.
  • Liposomal suspensions can also be used as pharmaceutically-acceptable carriers. These can be prepared according to methods known to those skilled in the art (U.S. Pat. No. 4,522,811, which is herein incorporated by reference in its entirety).
  • the T-cell epitope compounds and compositions of the present disclosure can be implanted within or linked to a biopolymer solid support that allows for the slow release of the T-cell epitope compounds and compositions to the desired site.
  • Dosage unit form refers to physically discrete units suited as unitary dosages for the subject to be treated; each unit containing a predetermined quantity of binding agent calculated to produce the desired therapeutic effect in association with the required pharmaceutical carrier.
  • the specification for the dosage unit forms of the instant disclosure are dictated by and directly dependent on the unique characteristics of the binding agent and the particular therapeutic effect to be achieved, and the limitations inherent in the art of compounding such T-cell epitope compounds and compositions for the treatment of a subject.
  • the composition may comprise at least 2, at least 3, at least 4, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, or at least 20 of the instantly-disclosed peptides or polypetides (including concatemeric polypeptides) or nucleic acids encoding such peptides or polypeptides (including concatemeric polypeptides).
  • a pharmaceutical composition can comprise at least 2, at least 3, at least 4, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, or at least 20 peptides or polypeptides (including up to 50 peptides or polypetides), including any value or range therebetween, comprising, consisting of, or consisting essentially of one or more peptides or polypeptides having an amino acid sequence comprising, consisting of, or consisting essentially of one or more of SEQ ID NOS: 1305, 1317, 1616, 1628, 1333, 1644, 1311, 1622, 1309, 1620, 1330, 1641, 1254, 1565, 1314, 1625, 1272,
  • vaccine as used herein includes an agent which may be used to cause, stimulate or amplify the immune system of animals (e.g., humans) against a pathogen.
  • Vaccines of the invention are able to cause or stimulate or amplify an immune response against a wide variety of beta-coronavirues and serve as a pan-beta-coronavirus vaccine agains infection by two or more beta-coronaviruses such as but not limited to SARS-CoV-2 infection (or a closely related virus such as Severe Acute Respiratory Syndrome coronavirus (SARS-CoV) or Middle East respiratory syndrome coronavirus (MERS-CoV)) and/or related diseases caused by MERS CoV, SARS CoV, or SARS-CoV-2, including MERS, SARS, and COVID-19.
  • SARS-CoV-2 infection or a closely related virus such as Severe Acute Respiratory Syndrome coronavirus (SARS-CoV) or Middle East respiratory syndrome coronavirus (MER
  • Immunization includes the process of delivering an immunogen to a subject. Immunization may, for example, enable a continuing high level of antibody and/or cellular response in which T-lymphocytes can kill or suppress the pathogen in the immunized animal, such as a human, which is directed against a pathogen or antigen to which the animal has been previously exposed.
  • Vaccines of the instant disclosure comprise an immunologically effective amount of a T cell epitope compound or composition of the instant disclosure as described above, and in aspects in a pharmaceutically acceptable vehicle and optionally with additional excipients and/or an adjuvant.
  • animals, and in aspects humans become at least partially or completely immune to any beta- coronavirus infection and/or related diseases caused by any beta-coronavirus, or resistant to developing moderate or severe beta-coronavirus infection and/or related diseases caused by any beta-coronavirus.
  • the instantly disclosed vaccines may be used to elicit a humoral and/or a cellular response, including CD4+ and CD8+ T effector cell responses.
  • an animal subject such as a human, is protected to an extent to which one to all of the adverse physiological symptoms or effects of beta-coronavirus infection and/or related diseases caused by beta-coronavirus are significantly reduced, ameliorated or totally prevented.
  • an immunologically effective dose may vary from subject to subject depending on factors such as the age and general condition of the subject, the nature of the formulation and the mode of administration.
  • An appropriate “effective amount” may be determined by one of ordinary skill in the art using only routine experimentation. For instance, methods are known in the art for determining or titrating suitable dosages of a vaccine to find minimal effective dosages based on the weight of the animal subject, including human subject, concentration of the vaccine and other typical factors.
  • the dosage of the vaccines of the present disclosure will depend on the species, breed, age, size, vaccination history, and health status of the animal (e.g., swine/pig) to be vaccinated, as well as the route of administration, e.g., subcutaneous, intradermal, oral intramuscular or intravenous administration.
  • the vaccines of the instant disclosure can be administered as single doses or in repeated doses.
  • the vaccines of the instant disclosure can be administered alone, or can be administered simultaneously or sequentially administered with one or more further compositions, such as other porcine immunogenic or vaccine compositions. Where the compositions are administered at different times, the administrations may be separate from one another or overlapping in time.
  • the vaccine comprises a unitary dose of between 0.1-3000 ⁇ g, including any value or range therebetween of polypeptide and/or nucleic acid of the instant disclosure.
  • the dosage of the vaccine, concentration of components therein and timing of administering the vaccine, which elicit a suitable immune response can be determined by methods such as by antibody titrations of sera, e.g., by ELISA and/or seroneutralization assay analysis and/or by vaccination challenge evaluation.
  • the vaccine comprises a novel, therapeutic T cell epitope compounds or compositions as disclosed herein) in purified form, optionally in combination with any suitable excipient, carrier, adjuvant, and/or additional protein antigen.
  • the vaccine comprises a nucleic acid as defined above, optionally in combination with any suitable excipient, carrier, adjuvant, and/or additional protein antigen.
  • the vaccine comprises a viral vector containing a nucleic acid as defined above.
  • the vaccine comprises one or more plasmid vectors.
  • Vaccine constructs including a T-cell epitope compound or composition of the present disclosure upon administration to a subject may initiate a strong T-cell mediated immune response, but may not induce a humoral immune response. Therefore, aspects of a vaccine against beta-coronavirus infection and/or related diseases caused by beta-coronavirus contains a combination of the putative T-cell epitopes together with either live attenuated virus (LAV, for example live attenuated MERS CoV, SARS CoV, SARS-CoV-2) or inactivated virus (for example inactivated MERS CoV, SARS, CoV, or SARS-CoV-2).
  • LAV live attenuated virus
  • MERS CoV live attenuated MERS CoV
  • SARS CoV SARS CoV
  • SARS-CoV-2 live attenuated virus
  • inactivated virus for example inactivated MERS CoV, SARS, CoV, or SARS-CoV-2
  • This vaccine composition upon administration to a subject may induce both cellular and humoral immune responses, thereby conferring comprehensive immunity against any beta-coronavirus infection and/or related diseases caused by any beta-coronavirus, in animals, including humans.
  • Vaccines may include other ingredients, known per se by one of ordinary skill in the art, such as pharmaceutically acceptable carriers, excipients, diluents, adjuvants, freeze drying stabilizers, wetting or emulsifying agents, pH buffering agents, gelling or viscosity enhancing additives, and preservatives, depending on the route of administration.
  • pharmaceutically acceptable carriers such as pharmaceutically acceptable carriers, excipients, diluents, adjuvants, freeze drying stabilizers, wetting or emulsifying agents, pH buffering agents, gelling or viscosity enhancing additives, and preservatives, depending on the route of administration.
  • Examples of pharmaceutically acceptable carriers, excipients or diluents include, but are not limited to demineralised or distilled water; saline solution; vegetable based oils such as peanut oil, arachis oil, safflower oil, olive oil, cottonseed oil, maize oil, sesame oil, or coconut oil; silicone oils, including polysiloxanes, such as methyl polysiloxane, phenyl polysiloxane and methylphenyl polysolpoxane; volatile silicones; mineral oils such as light liquid paraffin oil, or heavy liquid paraffin oil; squalene; cellulose derivatives such as methylcellulose, ethylcellulose, carboxymethylcellulose, carboxymethylcellulose sodium salt, or hydroxypropyl methylcellulose; lower alkanols, for example ethanol or isopropanol; lower aralkanols; lower polyalkylene glycols or lower alkylene glycols, for example polyethylene glycol, polypropylene glycol,
  • the carrier or carriers will form from 10% to 99.9% by weight of the vaccine composition and may be buffered by conventional methods using reagents known in the art, such as sodium hydrogen phosphate, sodium dihydrogen phosphate, potassium hydrogen phosphate, potassium dihydrogen phosphate, a mixture thereof, and the like.
  • adjuvants include, but are not limited to, oil in water emulsions, aluminum hydroxide (alum), immunostimulating complexes, non-ionic block polymers or copolymers, cytokines (like IL-1, IL-2, IL-7, IFN-a, IFN-b, IFN-g, etc.), saponins, monophosphoryl lipid A (MLA), muramyl dipeptides (MDP), MCA, and the like.
  • alum aluminum hydroxide
  • immunostimulating complexes include, but are not limited to, oil in water emulsions, aluminum hydroxide (alum), immunostimulating complexes, non-ionic block polymers or copolymers, cytokines (like IL-1, IL-2, IL-7, IFN-a, IFN-b, IFN-g, etc.), saponins, monophosphoryl lipid A (MLA), muramyl dipeptides (MDP), MCA, and the like.
  • Suitable adjuvants include, for example, aluminum potassium sulfate, heat-labile or heat-stable enterotoxin(s) isolated from Escherichia coli, cholera toxin or the B subunit thereof, diphtheria toxin, tetanus toxin, pertussis toxin, Freund's incomplete or complete adjuvant, etc.
  • Toxin-based adjuvants such as diphtheria toxin, tetanus toxin and pertussis toxin may be inactivated prior to use, for example, by treatment with formaldehyde.
  • Further adjuvants may include, but are not limited to, poly- ICLC, 1018 ISS, aluminum salts, Amplivax, AS 15, BCG, CP-870,893, CpG7909, CyaA, dSLIM, GM-CSF, IC30, IC31, Imiquimod, ImuFact IMP321, IS Patch, ISS, ISCOMATRTX, Juvlmmune, LipoVac, MF59, monophosphoryl lipid A, Montanide IMS 1312, Montanide ISA 206, Montanide ISA 50V, Montanide ISA-51, OK-432, OM-174, OM-197-MP-EC, ONTAK, PEPTEL, vector system, PLGA microparticles, resiquimod, SRL172, Virosomes and other Virus-like particles, YF-17D, VEGF trap, R848, beta-glucan, Pam3Cys, and Aquila's QS21 stimulon.
  • the adjuvant comprises poly-ICLC.
  • TLR9 agonist CpG and the synthetic double-stranded RNA (dsRNA) TLR3 ligand poly-ICLC are two of the most promising vaccine adjuvants currently in clinical development.
  • poly-ICLC appears to be the most potent TLR adjuvant when compared to LPS and CpG. This appears due to its induction of pro-inflammatory cytokines and lack of stimulation of IL-10, as well as maintenance of high levels of co-stimulatory molecules in DCs.
  • Poly-ICLC is a synthetically prepared double- stranded RNA consisting of polyl and polyC strands of average length of about 5000 nucleotides, which has been stabilized to thermal denaturation and hydrolysis by serum nucleases by the addition of polylysine and carboxymethylcellulose.
  • the compound activates TLR3 and the RNA helicase-domain of MDA5, both members of the PAMP family, leading to DC and natural killer (NK) cell activation and mixed production of type I interferons, cytokines, and chemokines.
  • freeze-drying stabilizer may be for example carbohydrates such as sorbitol, mannitol, starch, sucrose, dextran or glucose, proteins such as albumin or casein, and derivatives thereof.
  • Vaccines may additionally comprise at least one immunogen from at least one additional pathogen, e.g., a pig pathogen such as Actinobacillus pleuropneunomia; Adenovirus; Alphavirus such as Eastern equine encephalomyelitis viruses; Balantidium coli; Bordetella bronchiseptica; Brachyspira spp., preferably B. hyodyentheriae, B. pilosicoli, B. innocens, Brucella suis, preferably biovars 1, 2 and 3; Classical swine fever virus, Chlamydia and Chlamydophila spp., preferably C. pecorum and C.
  • a pig pathogen such as Actinobacillus pleuropneunomia
  • Adenovirus such as Eastern equine encephalomyelitis viruses
  • Balantidium coli Bordetella bronchiseptica
  • Brachyspira spp. preferably B.
  • Clostridium spp. preferably Cl. difficile, Cl. perfringens types A, B and C, Cl. novyi, Cl. septicum, Cl. tetani; Digestive and respiratory Coronavirus; Cryptosporidium parvum; Eimeria spp.; Eperythrozoonis suis currently named Mycoplasma haemosuis; Erysipelothrix rhusiopathiae; Escherichia coli; Haemophilus parasuis, preferably subtypes 1, 7 and 14; Hemagglutinating encephalomyelitis virus; lsospora suis; Japanese Encephalitis virus; Lawsonia intr acellular s; Leptospira spp., preferably Leptospira australis, Leptospira canicola, Leptospira grippotyphosa, Leptospira icterohaemorrhagicae, Lepto
  • avium, M. intracellular and M. bovis Mycoplasma hyponeumoniae; Parvovirus; Pasteurella multocida; Porcine circovirus; Porcine cytomegolovirus; Porcine parovirus, Porcine reproductive and respiratory syndrome virus: Pseudorabies virus; Rotavirus; Sagiyama virus; Salmonella spp., preferably S. thyhimurium and S. choleraesuis; Staphylococcus spp., preferably S.
  • Streptococcus spp. preferably Strep suis; Swine cytomegalovirus; Swine herpes virus; Swine influenza virus; Swinepox virus; Toxoplasma gondii, ⁇ Vesicular stomatitis virus and virus of exanthema of swine; or other isolates and subtypes of porcine circovirus.
  • the vaccine compositions of the instant disclosure may be liquid formulations such as an aqueous solution, water-in-oil or oil-in-water emulsion, syrup, an elixir, a tincture, or a preparation for parenteral, subcutaneous, intradermal, intramuscular or intravenous administration (e.g., injectable administration), such as sterile suspensions or emulsions.
  • Such formulations are known in the art and are typically prepared by dissolution of the antigen and other typical additives in the appropriate carrier or solvent systems.
  • Liquid formulations also may include suspensions and emulsions that contain suspending or emulsifying agents.
  • the route of administration can be percutaneous, via mucosal administration, or via a parenteral route (intradermal, intramuscular, subcutaneous, intravenous, or intraperitoneal).
  • Vaccine compositions according to the present disclosure may be administered alone, or can be co-administered or sequentially administered with other treatments or therapies.
  • a vaccine of the present disclosure can conveniently be administered intranasally, transdermally (i.e., applied on or at the skin surface for systemic absorption), parenterally, ocularly, etc.
  • the parenteral route of administration includes, but is not limited to, intramuscular, intravenous, intradermal, and intraperitoneal routes and the like.
  • vaccines of the present disclosure are administered intradermally, e.g., by using a micro needle patch as is known in the art or by using a commercial needle-free high-pressure device such as Pulse NeedleFree technology (Pulse 50TM Micro Dose Injection System, Pulse NeedleFree Systems; Lenexa, KS, USA).
  • Pulse NeedleFree technology Pulse 50TM Micro Dose Injection System, Pulse NeedleFree Systems; Lenexa, KS, USA.
  • the present disclosure also relates to methods of immunizing or inducing an immune response in animals (e.g., humans) comprising administering to said animal a peptide, polypeptide, concatemeric peptide, chimeric or fusion polypeptide, nucleic acid, cell, vector, pharmaceutical, or vaccine as described above.
  • the present disclosure also relates to methods of treating and/or preventing beta- coronavirus infection and/or related diseases caused by beta-coronavirus diseases in animals (e.g., humans) comprising administering to said animal a peptide, polypeptide, concatemeric peptide, chimeric or fusion polypeptide, nucleic acid, cell, vector, pharmaceutical, or vaccine as disclosed herein.
  • a vaccine of the present disclosure can conveniently be administered intranasally, transdermally (i.e., applied on or at the skin surface for systemic absorption), parenterally, ocularly, etc.
  • the parenteral route of administration includes, but is not limited to, intramuscular, intravenous, and intraperitoneal routes and the like.
  • the dosage of the vaccines of the present disclosure will depend on the species, breed, age, size, vaccination history, and health status of the animal (e.g., human) to be vaccinated, as well as the route of administration, e.g., subcutaneous, intradermal, oral intramuscular or intravenous administration.
  • the vaccines of the instant disclosure can be administered as single doses or in repeated doses.
  • the vaccines of the instant disclosure can be administered alone, or can be administered simultaneously or sequentially administered with one or more further compositions, such as other porcine immunogenic or vaccine compositions. Where the compositions are administered at different times, the administrations may be separate from one another or overlapping in time.
  • the present disclosure includes multiple rounds of administration of the instantly-disclosed vaccine compositions.
  • the vaccine can be boosted at one, two, three, and/or four week intervals. Such are known in the art to improve or boost the immune system to improve protection against the pathogen.
  • the present disclosure may also include assessing a subject’s immune system to determine if further administrations of the instantly-disclosed vaccine compositions is warranted.
  • multiple administrations may include the development of a prime boosting strategy of vaccination using the instantly-discloed vaccines (e.g.., polypeptide based or nucleic acid based as disclosed herein).
  • the vaccine can be boosted at 1, 2, 3, 4, 5, or 6 week intervals. In some aspects, the vaccine is boosted at 2 week intervals. In some apsects, the vaccine is boosted at 3 week intervals. In some aspects, peptide based vaccine and nucleice acid (e.g., RNA or DNA) vaccinations can be achieved in an alternative manner to provide a regimen of immunization with the same immunogen presented in different fashions to the subject’s immune system.
  • nucleice acid e.g., RNA or DNA
  • the vaccine compositions of the present disclosure are administered to a subject susceptible to or otherwise at risk for MERS-CoV infection and/or related diseases caused by MERS-CoV-2, including MERS, to enhance the subject own immune response capabilities.
  • the subject to which the vaccine is administered is, in one aspect, a human.
  • the animal may be susceptible to infection by MERS-CoV infection (or a closely related virus) and/or related diseases caused by MERS CoV, including MERS.
  • the vaccine compositions of the present disclosure are administered to a subject susceptible to or otherwise at risk for SARS-CoV infection and/or related diseases caused by SARS-CoV-2, including SARS, to enhance the subject own immune response capabilities.
  • the subject to which the vaccine is administered is, in one aspect, a human.
  • the animal may be susceptible to infection by SARS-CoV infection (or a closely related virus) and/or related diseases caused by SARS CoV, including SARS.
  • the vaccine compositions of the present disclosure are administered to a subject susceptible to or otherwise at risk for SARS-CoV-2 infection and/or related diseases caused by SARS-CoV-2, including COVID-19, to enhance the subject own immune response capabilities.
  • the subject to which the vaccine is administered is, in one aspect, a human.
  • the animal may be susceptible to infection by SARS-CoV-2 infection (or a closely related virus) and/or related diseases caused by SARS-CoV-2, including COVID-19.
  • the vaccine may comprise at least 2, at least 3, at least 4, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, or at least 20 of the instantly-disclosed peptides or polypetides (including concatemeric polypeptides) or nucleic acids encoding such peptides or polypeptides (including concatemeric polypeptides).
  • a vaccine can comprise at least 2, at least 3, at least 4, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, or at least 20 peptides or polypeptides (including up to 50 peptides or polypetides), including any value or range therebetween, comprising, consisting of, or consisting essentially of one or more peptides or polypeptides having an amino acid sequence comprising, consisting of, or consisting essentially of one or more of SEQ ID NOS: 4-1676, 1713-2595, 2605-2638, 2647-2718, and 2735-8692 and/or fragments and variants thereof, and optionally 1 to 12 additional amino acids distributed in any ratio on the N terminus and/or C-terminus of the polypeptide of SEQ ID NOS: 4-1676, 1713- 2595, 2605-2638, 2647-2718, and 2735-1305, 1317
  • the present disclosure also provides a container comprising an immunologically effective amount of a polypeptide, nucleic acid or vaccine as described above.
  • the present disclosure also provides vaccination kits comprising an optionally sterile container comprising an immunologically effective amount of the vaccine, means for administering the vaccine to animals, and optionally an instruction manual including information for the administration of the immunologically effective amount of the composition for treating and/or preventing beta- coronavirus infection (or a closely related virus) and/or related diseases caused by any beta- coronavirus including COVID-19.
  • Stimulating T-cells with T-cell epitope compounds and compositions of the present disclosure can stimulate, induce, and/or expand a corresponding naturally occurring immune response, e.g., stimulating, inducing, and/or expanding a corresponding naturally occurring immune response to any beta-coronavirus infection such as but not limited to SARS-CoV-2 infection (or a closely related virus such as Severe Acute Respiratory Syndrome (SARS) or Middle East respiratory syndrome coronavirus (MERS-CoV)) and/or related diseases caused by SARS-CoV-2, including MERS, SARS, and COVID-19, including CD4+ and/or CD8+ T cell responses, and in aspects results in increased secretion of one or more cytokines and chemokines.
  • SARS-CoV-2 infection or a closely related virus such as Severe Acute Respiratory Syndrome (SARS) or Middle East respiratory syndrome coronavirus (MERS-CoV)
  • SARS-CoV-2 infection or a closely related virus such as Se
  • T-cells activated by the T-cell epitope compounds and compositions of the present disclosure stimulate cell-mediated immunity against any beta-coronavirus infection and/or related diseases caused by any beta-coronavirus in a subject.
  • T cells activated by the T-cell epitope compounds and compositions of the present disclosure stimulate cell-mediated immunity against beta-coronavirus infection in a subject.
  • the present disclosure is directed to a method of stimulating, inducing, and/or expanding an immune response, e.g., against beta-coronavirus and/or related diseases caused by beta-coronavirus in a subject in need thereof by administering to the subject a therapeutically effect amount of a T-cell epitope composition (compound or composition of the present disclosure).
  • the present disclosure is directed to a method of preventing, treating, or ameliorating a disease by beta-coronavirus infection in a subject in need thereof by administering to the subject a therapeutically effect amount of a T-cell epitope compound or composition of the present disclosure.
  • T-cells specifically recognize epitopes presented by cells in the context of MHC (Major Histocompatibility Complex) Class I and II molecules. These T-cell epitopes can be represented as linear sequences comprising 7 to 30 contiguous amino acids that fit into the MHC Class I or II binding groove.
  • the conserveatrix system (EpiVax, Buffalo, Rhode Island) is an algorithm useful for identifying 9-mer polypeptide sequences from a larger set of data.
  • the conserveatrix system parses input sequences into 9-mer sequences that are conserved amongst multiple inputted whole sequences, such as multiple strains of the same pathogen, for even the most mutable of potential vaccine targets. These 9-mer sequences may be searched for identically matched 9- mer sequences across data sets.
  • the EpiMatrixTM system (EpiVax, Buffalo, Rhode Island) is a set of predictive algorithms encoded into computer programs useful for predicting class I and class II HLA ligands and T cell epitopes.
  • the EpiMatrixTM system uses matrices in order to model the interaction between specific amino acids and binding positions within the HLA molecule.
  • Each frame is then scored for predicted affinity to one or more common alleles of the HLA molecules.
  • n-mer peptide specific amino acid codes one for each of 20 naturally occurring amino acids
  • relative binding positions (1 to n) are used to select coefficients from the predictive matrix.
  • Individual coefficients are derived using a proprietary method similar to, but not identical to, the pocket profile method first developed by Sturniolo (Sturniolo T et al., 1999, Nat Biotechnol, 17(6):555-61). Individual coefficients are then summed to produce a raw score.
  • EpiMatrixTM raw scores are then normalized with respect to a score distribution derived from a very large set of randomly generated peptide sequences.
  • FIGS. 2-4 are EpiMatrix Cluster detail reports for identified MHC class II clusters of the envelope (SEQ ID NO: 1) of SARS-CoV-2.
  • FIGS. 6-15 are EpiMatrix Cluster detail reports for identified MHC class II clusters of the membrane (SEQ ID NO: 2) of SARS- CoV-2.
  • FIGS. 2-4 are EpiMatrix Cluster detail reports for identified MHC class II clusters of the membrane (SEQ ID NO: 2) of SARS- CoV-2.
  • FIG. 17-55 are EpiMatrix Cluster detail reports for identified MHC class II clusters of the spike (SEQ ID NO: 3) of SARS-CoV-2.
  • FIG. 59 is an EpiMatrix staircase report for identified MHC class I clusters of the envelope (SEQ ID NO: 1) of SARS-CoV-2.
  • FIG. 60 is an EpiMatrix staircase report for identified MHC class I clusters of the membrane (SEQ ID NO: 2) of SARS-CoV-2.
  • FIG. 61 is an EpiMatrix staircase report for identified MHC class I clusters of the spike (SEQ ID NO: 3) of SARS-CoV-2.
  • Peptides containing clusters of putative T cell epitopes are more likely to test positive in validating in vitro and in vivo assays.
  • the results of the initial EpiMatrixTM analysis is further screened for the presence of putative T cell epitope “clusters” using a second proprietary algorithm known as ClustimerTM algorithm.
  • the ClustimerTM algorithm identifies sub-regions contained within any given amino acid sequence that contains a statistically unusually high number of putative T cell epitopes.
  • Typical T-cell epitope “clusters” range from about 9 to roughly 30 amino acids in length and, considering their affinity to multiple alleles and across multiple 9-mer frames, can contain anywhere from about 4 to about 40 putative T cell epitopes.
  • T-cell epitopes of the instant disclosure contain several putative T-cell epitopes forming a pattern known as a T-cell epitope cluster.
  • JanusMatrix The JanusMatrix system (EpiVax, Buffalo, Rhode Island) useful for screening peptide sequences for cross-conservation with a host proteome.
  • JanusMatrix is an algorithm that predicts the potential for cross-reactivity between peptide clusters and the host genome or proteome, based on conservation of TCR-facing residues in their putative MHC ligands.
  • the JanusMatrix algorithm first considers all the predicted epitopes contained within a given protein sequence and divides each predicted epitope into its constituent agretope and epitope. Each sequence is then screened against a database of host proteins.
  • Peptides with a compatible MHC-facing agretope i.e., the agretopes of both the input peptide and its host counterparty are predicted to bind the same MHC allele
  • the JanusMatrix Homology Score suggests a bias towards immune tolerance.
  • cross-conservation between autologous human epitopes and epitopes in the therapeutic may increase the likelihood that such a candidate will be tolerated by the human immune system.
  • peptide clusters are screened against human genomes and proteomes, based on conservation of TCR-facing residues in their putative HLA ligands. The peptides are then scored using the JanusMatrix Homology Score. In aspects, peptides with a JanusMatrix Homology Score below 2.5 or below 3.0 indicate low tolerogenicity potential and may be useful for vaccines.
  • FIG. 56 is the JanusMatrix reports for identified MHC class II clusters of the envelope (SEQ ID NO: 1) of SARS-CoV-2.
  • FIG. 57 is the JanusMatrix reports for identified MHC class II clusters of the membrane (SEQ ID NO: 2) of SARS-CoV-2.
  • FIG. 58 is the JanusMatrix reports for identified MHC class II clusters of the spike (SEQ ID NO: 3) of SARS-CoV-2.
  • the VaccineCAD system is useful for arranging potential epitopic vaccine candidates into a string to avoid creation of novel epitopes upon joining of the vaccine candidate sequences.
  • VaccineCAD designs potential vaccine candidates into a string-of-beads vaccine while minimizing any deleterious junctional epitopes that may appear in the joining process.
  • VaccineCAD may use EpiMatrix to predict junctional epitopes.
  • Particularly concatemeric peptides of interest developed using VaccineCad are those of SEQ ID NOS: 1677-1692, 2593-2604, 2639-2646, and 2719-2734, and in aspects SEQ ID NOS: 1677-1684.
  • nucleic acids of interest are those encoding a peptide or polypeptide comprising, consisting of, or consisting essentially of one or more peptides or polypeptides having an amino acid sequence off SEQ ID NOS: 1677-1692, 2593-2604, 2639-2646, and 2719-2734.
  • the selected epitope clusters for HLA Class I and Class II used to produce the concatemeric peptides of SEQ ID NOS: 1677-1681, 2593-2604, 2641-2646, and 2719-2722 (and associated nucleic acid constructs encoding such) include the below sequences in TABLE 2.
  • the selected epitope clusters for HLA Class I and Class II used to produce the concatemeric peptides of SEQ ID NOS: 1682-1684 (and associated nucleic acid constructs encoding such) include the below sequences in TABLE 3.
  • Predicted epitope sequences were concatenated to form 22 multi-epitope pseudoproteins.
  • Vaccine constructs predicted to have no junctional epitopes were designed.
  • VaccineCAD was used to rearrange the peptides to avoid creation of novel epitopes at peptide junctions, and used JanusMatrix to predict junctional epitopes. Where reordering did not sufficiently reduce the potential for junctional immunogenicity, spacers (e.g., Gly-Pro-Gly- Pro-Gly) were introduced.
  • spacers e.g., Gly-Pro-Gly- Pro-Gly
  • a cleavage promoting motif or a binding inhibiting ‘breaker’ sequence could be introduced between peptides to optimize epitope processing.
  • T cell epitope cluster flanking residues were extended or removed to further minimize junctional T cell epitope content.
  • These post- VaccineCAD, optimized multi-epitope constructs are represented by sequences SEQ ID NOS: 1677-1684,2593-2604, 2641-2646, and 2719-2722. These constructs are located below in Tables 4-11, Tables 35-40, and Tables 41- 44. Additional epitope concatemers containing coronavirus cross-conserved sequences are represented by SEQ ID NOS: 1685-1692 and 2723-2734, and are found in Tables 12-19 and 45-56.
  • EpiMatrix Cluster Scores were calculated for each identified T cell epitope cluster. T cell epitope clusters were then screened for crossconservation against the human proteome using the JanusMatrix algorithm. T cell epitope clusters with JanusMatrix Human Homology Scores above two were considered as potentially tolerogenic (Tregitopes). To identify T cell epitope cluster cross-conserved with other coronaviruses, both the standard and a less stringent version of the JanusMatrix algorithm were applied. Finally, the Class I T cell epitope content for 9-mer and 10-mer frames of each T cell epitope clusters was identified for a set of six class I HLA supertype alleles using EpiMatrix.
  • SARS-CoV-2 T cell epitope clusters were next ranked based on Cluster Scores, JanusMatrix Human Homology Scores, and JanusMatrix Coronavirus Homology Scores. Peptides with Cysteines in their cores were excluded. The top 22 T cell epitope clusters were selected for vaccine design.
  • T cell epitope clusters were then aggregated into epitope concatemers using the VaxCAD algorithm.
  • VaxCAD also optimized the sequence arrangement to minimize Class I and II junctional epitope content.
  • Gly-Pro-Gly-Pro-Gly spacer sequences were introduced between epitopes to remove junctional epitopes where reordering did not sufficiently reduce potential junctional immunogenicity.
  • T cell epitope cluster flanking residues were extended or removed to further minimize junctional T cell epitope content.
  • Nine concatemers were constructed with two or three T cell epitope clusters per concatemer.
  • the peptides with identified originating protein and starting position utilized to generate the concatemers of SEQ ID NOS: 2593-2604 (FIG. 120) are presented in Tables 20-28.
  • Table 29 also presents 9 concatemer constructs.
  • Vaccine construct designs are developed, such as is demonstrated in the specification, and in aspects as specifically exemplified in Example 2. In aspects, this results in a concatemeric polypeptide vaccine or an “epistring” that consists of overlapping T-cell epitopes.
  • such vaccines may be used for stimulating, inducing, and/or expanding an immune response against SARS-CoV-2 infection (or a closely related virus such as Severe Acute Respiratory Syndrome (SARS) or Middle East respiratory syndrome coronavirus (MERS-CoV)) and/or related diseases caused by SARS-CoV-2, including COVID-19.
  • SARS Severe Acute Respiratory Syndrome
  • MERS-CoV Middle East respiratory syndrome coronavirus
  • such vaccines initiate a strong T-cell mediated immune response, and has potential of inducing a humoral immune response.
  • a vaccine containing a combination of the epistring together with either live attenuated virus (LAV) or inactivated virus is administered in an immunization trial in an appropriate animal model , e.g., mice, rats, rabbits, hamsters, etc., or even humans, as are known in the art. Data from administration of this combination vaccine provides positive results on the safety and effectiveness of the vaccine.
  • LAV live attenuated virus
  • This vaccination approach is expected to induce both cellular and humoral immune responses, thereby stimulating, inducing, and/or expanding an immune response against SARS-CoV-2 infection (or a closely related virus such as Severe Acute Respiratory Syndrome (SARS) or Middle East respiratory syndrome coronavirus (MERS-CoV)) and/or related diseases caused by SARS-CoV-2, including COVID-19, in humans.
  • SARS-CoV-2 infection or a closely related virus such as Severe Acute Respiratory Syndrome (SARS) or Middle East respiratory syndrome coronavirus (MERS-CoV)
  • SARS-CoV-2 infection or a closely related virus such as Severe Acute Respiratory Syndrome (SARS) or Middle East respiratory syndrome coronavirus (MERS-CoV)
  • SARS-CoV-2 infection or a closely related virus such as Severe Acute Respiratory Syndrome (SARS) or Middle East respiratory syndrome coronavirus (MERS-CoV)
  • SARS-CoV-2 infection or a closely related virus
  • polypeptides of the present disclosure e.g., but not limited to, a peptide or polypeptide comprising, consisting, or consisting essentially of an amino acid sequence of SEQ ID NOS: 4-1676, 1713-2595, 2605-2638, 2647-2718, and 2735-8692 (and/or fragments or variants thereof), and optionally 1 to 12 additional amino acids distributed in any ratio on the N terminus and/or C-terminus of the polypeptide of SEQ ID NOS: 4-1676, 1713- 2595, 2605-2638, 2647-2718, and 2735-8692) are produced by direct chemical synthesis or by recombinant methods (J Sambrook et ah, Molecular Cloning: A Laboratory Manual, (2ED, 1989), Cold Spring Harbor Laboratory Press, Cold Springs Harbor, NY (Publ)).
  • polypeptides as disclosed herein may be capped with an n- terminal acetyl and/or c-terminal amino group. HPLC, mass spectrometry and UV scan (ensuring purity, mass and spectrum, respectively) analysis of the selected polypeptides will indicate > 80% purity.
  • Binding activity is analyzed at EpiVax (Providence, Rhode Island) and is conducted for any polypeptides of the present disclosure (e.g., but not limited to, a peptide or polypeptide comprising, consisting, or consisting essentially of an amino acid sequence of SEQ ID NOS: 4-1676, 1713-2595, 2605-2638, 2647-2718, and 2735-8692 (and/or fragments or variants thereof), and optionally 1 to 12 additional amino acids distributed in any ratio on the N terminus and/or C-terminus of the polypeptide of SEQ ID NOS: 4-1676, 1713- 2595, 2605-2638, 2647-2718, and 2735-8692).
  • a polypeptide or polypeptide comprising, consisting, or consisting essentially of an amino acid sequence of SEQ ID NOS: 4-1676, 1713-2595, 2605-2638, 2647-2718, and 2735-8692 (and/or fragments or variants thereof), and optionally 1 to 12
  • the binding assay used yields an indirect measure of peptide-MHC affinity.
  • Soluble HLA molecules are loaded onto a 96-well plate with the unlabeled experimental polypeptides and labeled control peptide. Once the binding mixture reaches steady equilibrium (at 24 hours), the HLA-polypeptide complexes are captured on an ELISA plate coated with anti-human DR antibody and detected with a Europium-linked probe for the label (PerkinElmer, Waltham, MA). Time-resolved fluorescence measuring bound labeled control peptide is assessed by a SpectraMax® M5 unit (Spectramax, Radnor, PA).
  • Binding of experimental polypeptides is expressed as the percent inhibition of the labeled control peptide (experimental fluorescence / control fluorescence multiplied by 100). The percent inhibition values for each experimental polypeptide (across a range of molar concentrations) is used to calculate the concentration at which it inhibits 50% of the labeled control polypeptide’s specific binding, i.e., the polypeptides’s IC50. Select experimental polypeptides are solvated in DMSO. The diluted polypeptide is mixed with binding reagents in aqueous buffering solution, yielding a range of final concentrations from 100,000 nM down to 100 nM.
  • the select polypeptides are assayed against a panel of eight common Class II HLA alleles: DRB 1*0101, DRB 1*0301, DRB 1*0401, DRB1*0701, DRB1*0801, DRB1*1101, DRB1*1301, and DRB1*1501. From the percent inhibition of labeled control peptide at each concentration, IC50 values are derived for each polypeptide/allele combination using linear regression analysis.
  • the experimental polypeptides are considered to bind with very high affinity if they inhibit 50% of control peptide binding at a concentration of 100 nM or less, high affinity if they inhibit 50% of control peptide binding at a concentration between 100 nM and 1,000 nM, and moderate affinity if they inhibit 50% of control peptide binding at a concentration between 1,000 nM and 10,000 nM.
  • Low affinity peptides inhibit 50% of control peptide binding at concentrations between 10,000 nM and 100,000 nM.
  • Peptides that fail to inhibit at least 50% of control peptide binding at any concentration below 100,000 nM and do not show a dose response are considered non-binders (NB).
  • Soluble MHC binding assays are performed on any of the instantly disclosed polypeptides (e.g., but not limited to, a peptide or polypeptide comprising, consisting, or consisting essentially of an amino acid sequence of SEQ ID NOS: 4-1676, 1713-2595, 2605- 2638, 2647-2718, and 2735-8692 (and/or fragments or variants thereof), and optionally 1 to 12 additional amino acids distributed in any ratio on the N terminus and/or C-terminus of the polypeptide of SEQ ID NOS: 4-1676, 1713-2595, 2605-2638, 2647-2718, and 2735-8692).
  • a peptide or polypeptide comprising, consisting, or consisting essentially of an amino acid sequence of SEQ ID NOS: 4-1676, 1713-2595, 2605- 2638, 2647-2718, and 2735-8692 (and/or fragments or variants thereof), and optionally 1 to 12 additional amino acids distributed in any ratio on the N termin
  • Soluble MHC binding assays are performed on selected polypeptides as disclosed herein according to the methods described previously.
  • IC50 values (nM) will be derived from a six- point inhibition curve.
  • EpiMatrixTM Predictions, calculated IC50 values, and results classifications are reported for each polypeptide and HLA allele.
  • Binding curves are generated for certain polypeptides against the selected Class II HLA alleles, such as for the HLA DRB 1 *0801 assay and the HLA DRBl *1501 assay.
  • HLA-DR Class II HLA
  • CD86 surface expression of Class II HLA (HLA-DR) and CD86 by professional antigen presenting cells (APCs) is one way APCs modulate T cell response.
  • candidate polypeptides are tested for their ability to effect (e.g., upregulate) the expression of Class II HLA and the co-stimulatory molecule CD86 on the surface of professional APCs, specifically dendritic cells.
  • Polypeptides of the present disclosure e.g., but not limited to, a peptide or polypeptide comprising, consisting, or consisting essentially of an amino acid sequence of SEQ ID NOS: 4-1676, 1713-2595, 2605-2638, 2647-2718, and 2735-8692 (and/or fragments or variants thereof), and optionally 1 to 12 additional amino acids distributed in any ratio on the N terminus and/or C-terminus of the polypeptide of SEQ ID NOS: 4-1676, 1713-2595, 2605-2638, 2647- 2718, and 2735-8692) are individually tested for effector potential using a proprietary APC phenotyping assay previously developed at EpiVax (EpiVax, Buffalo, Rhode Island).
  • PBMC Previously harvested and frozen PBMC are thawed and suspended in chRPMI by conventional means.
  • HLA typing is conducted on small, extracted samples of cellular material, provided by EpiVax, by Hartford Hospital (Hartford, Connecticut).
  • 0.5x106 cells are extracted, screened for the presence of surface marker CD1 lc (a marker specific to dendritic cells) and are analyzed for the presence of surface markers HLA-DR and CD86 by flow cytometry.
  • CD1 lc a marker specific to dendritic cells
  • the remaining cells are plated (4.0x106 cell per ml in chRPMI plus 800ul media) and are stimulated (50 ⁇ g/mL) with one of the selected peptides or positive and negative controls including buffer only (negative control), Tregitope 167 (negative control for effector activity) (21st Century Biochemicals, Marlboro, MA), Flu- HA 306-318 (positive control) (21ST Century Biochemicals, Marlboro, MA) and Ova 323-339 (negative control) (21st Century Biochemicals, Marlboro, MA). Plated cells are incubated for seven days at 37°C. On assay day 7, incubated cells are screened by flow cytometry for the presence of surface marker CD1 lc. CD1 lc positive cells are then analyzed for the presence of surface markers HLA-DR and CD86. The experimental peptides are tested in samples drawn from five different human donors.
  • Leukocyte Reduction Filters are obtained from the Rhode Island Blood Center (Providence, RI) to filter white blood cells from whole blood obtained from healthy donors. After the whole blood is run through the filters, the filters are flushed in the opposite direction to push collected white blood cells out of the filter. The white blood cells are isolated using a conventional FicollTM separation gradient (GE Healthcare). The collected white blood cells are thereafter frozen for future use. When needed for use in an assay, the frozen white blood cells are thawed using conventional methods. For the GvHD studies discussed below, PBMCs are obtained (e.g., from HemaCare, VanNuys, CA). Exposure to the instantly disclosed polypeptides as disclosed herein on the phenotypes of dendritic cells is measured by multiple means.
  • RI Rhode Island Blood Center
  • dot-plots contrasting surface expression of CD1 lc and HLA-DR
  • dot-plots of cells exposed to all control and experimental peptides are overlaid onto dot-plots produced from control cells exposed to only the culture media.
  • the overlay provides an effective method to visually observe shifts in HLA-DR distribution between polypeptide stimulated, and unstimulated CD1 lc-high cells (data not shown). Observed shifts in the distribution of HLA- DR are reported as a qualitative measure.
  • the change in intensity of HLA-DR expression for the CD11 c-high segment of each dot-plot is calculated.
  • Percent change in intensity of HLA- DR expression equals Mean Florescence Index (MFI) of HLA-DR expression for peptide exposed cells minus MFI of HLA-DR expression for media exposed cells divided by MFI of HLA-DR expression for media exposed cells, times 100 (HLA-DRMFIpeptide - HLA- DRMFImedia / HLA-DRMFImedia * 100).
  • MFI Mean Florescence Index
  • Percent change in the percentage of HLA-DR-low cells is calculated, and equals the percent of HLA-DR-low for peptide exposed cells minus the percent of HLA- DR-low for media exposed cells divided by percent of HLA-DR-low for media exposed cells times 100 (HLA-DR-low%peptide - HLA-DR-low%media / HLA-DR-low%media * 100).
  • HLA-DR-low%peptide - HLA-DR-low%media / HLA-DR-low%media * 100 a negative change in observed HLA-DR MFI and a positive change in percentage of HLA-DR-low cells present in the CD 1 lc-high population indicates reduced expression of HLA and a shift to a regulatory APC phenotype.
  • CD86 is a costimulatory molecule known to promote T cell activation.
  • dot plots contrasting surface expression of CD1 lc and CD86 are produced.
  • Dot plots of cells exposed to all control and experimental Tregitopes are overlaid onto dots plots produced from control cells exposed to only the culture media.
  • the overlay provides an effective method to visually observe shifts in CD86 distribution between polypeptide stimulated and un-stimulated CD 1 lc- high cells. Observed shifts in the distribution of CD86 are reported as a qualitative measure.
  • Percent change in intensity of CD86-high expression equals Mean Florescence Index (MFI) of CD86 expression for peptide exposed cells minus MFI of CD86- high expression for media exposed cells divided by MFI of CD86 expression for media exposed cells, times 100 (CD86-highMFIpeptide - CD86-highMFImedia / CD86-highMFImedia * 100).
  • MFI Mean Florescence Index
  • Percent change in the percentage of CD86-high cells equals the percent of CD86-high for peptide exposed cells minus the percent of CD86-high for media exposed cells divided by percent of CD86-high for media exposed cells, times 100 (CD86- low%Ipeptide - CD86-low%media / CD86-low%media * 100).
  • a negative change in observed CD86 MFI and a positive change in percentage of CD86-low cells present in the CD1 lc-high population indicates reduced expression of CD86 and a shift to a regulatory APC phenotype.
  • a positive change in observed CD86 MFI and a negative change in percentage of CD86-low cells present in the CD 1 lc-high population indicates increased expression of CD86 and a shift to an effector APC phenotype.
  • Dendritic cell phenotyping assays are performed on the polypeptides of the instant disclosure according to the methods described previously.
  • CD1 lc+/HLA-DR+ population will be analyzed on assay day 7 across the five donors in the presence of various peptide stimulants.
  • the effector polypeptides of the instant disclosure e.g., select polypeptides of the instant disclosure (e.g., but not limited to, a peptide or polypeptide comprising, consisting, or consisting essentially of an amino acid sequence of SEQ ID NOS: 4-1676, 1713- 2595, 2605-2638, 2647-2718, and 2735-8692 (and/or fragments or variants thereof), and optionally 1 to 12 additional amino acids distributed in any ratio on the N terminus and/or C- terminus of the polypeptide of SEQ ID NOS: 4-1676, 1713-2595, 2605-2638, 2647-2718, and 2735-8692), including the concatemeric peptides of SEQ ID NOS: 4-1676, 1713-2595, 2605-2638, 2647-2718, and 2735-8692), including
  • CD86-hi cells present in the samples treated with effector polypeptides of the instant disclosure (e.g., a peptide or polypeptide comprising, consisting, or consisting essentially of an amino acid sequence of SEQ ID NOS: 4-1676, 1713-2595, 2605-2638, 2647- 2718, and 2735-8692 (and/or fragments or variants thereof), and optionally 1 to 12 additional amino acids distributed in any ratio on the N terminus and/or C-terminus of the polypeptide of SEQ ID NOS: 4-1676, 1713-2595, 2605-2638, 2647-2718, and 2735-8692), including the concatemeric peptides of SEQ ID NOS: 1677-1692, 2593-2604, 2639-2646, and 2719-2734)), excluding one or more
  • EXAMPLE 6 Memory T cell responses to SARS-CoV-2 in COVID-19 Convalescents Materials and Methods
  • SARS-CoV-2 convalescent donors SARS-CoV-2 convalescent donors.
  • Convalescent patients were recruited by Sanguine Biosciences, a clinical services group that identified, consented and enrolled participants. Inclusion criteria included subjects (i) willing and able to provide written informed consent and photo identification, (ii) aged 18-60, both male or female, (iii) confirmed COVID-19 diagnosis (recovered) with date of diagnosis a minimum of 30 days from blood collection, and (iv) positive COVID-19 PCR based-kit documented by time-stamped medical record and/or diagnostic test report and test kit used identified.
  • Exclusion criteria included subjects who (i) are pregnant or nursing, (ii) have a known history of HIV, hepatitis or other infectious diseases, (iii) have autoimmune diseases, (iv) in vulnerable patient population (prisoners, mentally impaired), (v) have medical conditions impacting their ability to donate blood (i.e. anemia, acute illness) (vi) received immunosuppressive therapy or steroids within the last 6 months , (vii) received an investigational product in the last 30 days, (viii) experienced excess blood loss including blood donation defined as 250 mL in the last month or 500 mL in the last two months, or (ix) had a positive COVID-19 PCR test, but were asymptomatic. Samples were collected in accordance with NIH regulations and with IRB approval.
  • PBMC culture Thawed whole PBMCs (normal healthy donors) were rested overnight and expanded by antigen stimulation (including select polypeptides of the instant disclosure (e.g., but not limited to, a peptide or polypeptide comprising, consisting, or consisting essentially of an amino acid sequence of SEQ ID NOS: 4-1676, 1713-2595, 2605-2638, 2647-2718, and 2735-8692 (and/or fragments or variants thereof), and optionally 1 to 12 additional amino acids distributed in any ratio on the N terminus and/or C-terminus of the polypeptide of SEQ ID NOS: 4-1676, 1713-2595, 2605-2638, 2647-2718, and 2735-8692), including the concatemeric peptides of SEQ ID NOS
  • Peptides were added individually at 10 ⁇ g/ml and pooled at 10 ⁇ g/ml (8 peptides, 1.25 ⁇ g/mL) to triplicate wells containing 250,000 PBMCs (ex vivo) or 100,000 PBMCs (cultured) in RPMI medium supplemented with 10% human AB serum.
  • Triplicate wells were plated with ConA (10 ⁇ g/ml) as a positive control, and six wells containing no antigen stimulus were used for background determination. Cells were incubated for 40-48 hours at 37°C under a 5% C02 atmosphere. Plates were developed according to the manufacturer’s directions using FITC-labeled anti-IFN-g detection antibody.
  • Raw spot counts were recorded by ZellNet Consulting, Inc. using a FluoroSpot reader system (iSpot Spectrum, AID, Strassberg, Germany) with software version 7.0, build 14790, where fluorescent spots were counted utilizing separate filters for FITC, Cy3, and Cy5. Camera exposure and gain settings were adapted for each filter to obtain high quality spot images preventing over- or underexposure. Fluorophore-specific spot parameters were defined using spot size, spot intensity and spot gradient (fading of staining intensity from center to periphery of spot), and a spot separation algorithm was applied for optimal spot detection.
  • Results were calculated as the average number of spots in the peptide wells, adjusted to spots per one million cells. Responses meeting the following criteria are positive when the number of spots is (i) at least twice background, (ii) greater than 50 spot forming cells per well above background (1 response per 20,000 PBMCs), and (iii) statistically different (p ⁇ 0.05) from the media-only control by the Student’s t test.
  • peptide 1 (or rank 1) is SEQ ID NO: 1091 (cluster SEQ ID NO: 1401); peptide 2 (or rank 2) is SEQ ID NO: 1062 (cluster SEQ ID NO: 1373); peptide 3 (or rank 3) is SEQ ID NO: 1085 (cluster SEQ ID NO: 1395); peptide 4 (or rank 4) is SEQ ID NO: 1066 (cluster SEQ ID NO: 1377); peptide 5 (or rank 5) is SEQ ID NO: 1080 (cluster SEQ ID NO: 1391); peptide 6 (or rank 6) is SEQ ID NO: 1081 (cluster SEQ ID NO: 1392); peptide 7 (or rank 7) is SEQ ID NO: 1065 (cluster SEQ ID NO: 1376); peptide 8 (or rank 8) is SEQ ID NO: 1092 (cluster SEQ ID NO: 1403); peptide 9 (or rank 9) is SEQ ID NO: 1104 (cluster SEQ ID NO: 1091 (cluster SEQ
  • pool A includes the following: SEQ ID NO: 1091 (cluster SEQ ID NO: 1401); SEQ ID NO: 1062 (cluster SEQ ID NO: 1373); SEQ ID NO: 1085 (cluster SEQ ID NO: 1395); SEQ ID NO: 1066 (cluster SEQ ID NO: 1377); SEQ ID NO: 1080 (cluster SEQ ID NO: 1391); SEQ IDNO: 1081 (cluster SEQ ID NO: 1392); SEQ ID NO: 1065 (cluster SEQ ID NO: 1376); SEQ ID NO: 1092 (cluster SEQ ID NO: 1403).
  • Pool B includes the following: SEQ ID NO: 1104 (cluster SEQ ID NO: 1415); SEQ ID NO: 1071 (cluster SEQ ID NO: 1382); SEQ ID NO: 1107 (cluster SEQ ID NO: 1418); SEQ ID NO: 1072 (cluster SEQ ID NO: 1383); SEQ ID NO: 1074 (cluster SEQ ID NO: 1384); SEQ ID NO: 1115 (cluster SEQ ID NO: 1426); SEQ ID NO: 1096 (cluster SEQ ID NO: 1407); SEQ ID NO: 1110 (cluster SEQ ID NO: 1421).
  • Pool C includes the following: peptide SEQ ID NO: 1116 (cluster SEQ ID NO: 1427); SEQ ID NO: 1105 (cluster SEQ ID NO: 1416); SEQ ID NO: 1055 (cluster SEQ ID NO:
  • SEQ ID NO: 1120 cluster SEQ ID NO: 1430
  • Pool D includes the following: SEQ ID NO: 1120 (cluster SEQ ID NO: 1430)
  • ex vivo immune recall responses differentiate SARS-CoV-2 naive and experienced individuals and exhibit different COVID-19 immunotypes.
  • Robust and failed immune responses in convalescent donors may represent different immunotypes characterized in a deep immune profiling study of SARS-CoV-2 experienced humans (Giles et al. Deep immune profiling of COVID-19 patients reveals distinct immunotypes with therapeutic implications. Science. 2020 Jul 15:eabc8511. doi: 10.1126/science. abc8511. PMID: 32669297, herein incorporated by reference in its entirety).
  • a polypeptide of the instant disclosure e.g., a peptide or polypeptide comprising, consisting, or consisting essentially of an amino acid sequence of SEQ ID NOS: 4-1676, 1713- 2595, 2605-2638, 2647-2718, and 2735-8692 (and/or fragments or variants thereof), and optionally 1 to 12 additional amino acids distributed in any ratio on the N terminus and/or C- terminus of the polypeptide of SEQ ID NOS: 4-1676, 1713-2595, 2605-2638, 2647-2718, and 2735-8692), including the concatemeric peptides of SEQ ID NOS: 1677-1692, 2593-2604, 2639-2646, and 2719-2734)), excluding one or more of a peptide or polypeptide having an amino acid sequence comprising, consisting of, or consisting essentially of one or more of SEQ ID NOS: 1056-1057, 1059, 1367-1368, 1370, 1063, 1068
  • one or more of a peptide or polypeptide having an amino acid sequence comprising, consisting of, or consisting essentially of one or more of SEQ ID NOS: 1056-1057, 1059, 1367-1368, 1370, 1063, 1068, 101, 1374, 1379, 1382, 1073, 1075, 1083-1084, 1094, 1101, 1103, 1108, 1111,

Landscapes

  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Virology (AREA)
  • General Health & Medical Sciences (AREA)
  • Organic Chemistry (AREA)
  • Medicinal Chemistry (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Animal Behavior & Ethology (AREA)
  • Veterinary Medicine (AREA)
  • Molecular Biology (AREA)
  • Public Health (AREA)
  • Biochemistry (AREA)
  • Genetics & Genomics (AREA)
  • Oncology (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • General Chemical & Material Sciences (AREA)
  • Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
  • Gastroenterology & Hepatology (AREA)
  • Communicable Diseases (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Biophysics (AREA)
  • Immunology (AREA)
  • Microbiology (AREA)
  • Mycology (AREA)
  • Epidemiology (AREA)
  • Peptides Or Proteins (AREA)

Abstract

La présente invention concerne de manière générale de nouvelles compositions à base d'épitopes, comprenant des vaccins pan bêta-coronavirus et leur utilisation dans le traitement, le soulagement ou la prévention de maladies provoquées par des coronavirus ou des virus apparentés. L'invention concerne également des acides nucléiques, des vecteurs et des cellules qui expriment les polypeptides et leurs utilisations. Les compositions sont particulièrement appropriées pour produire des vaccins, en particulier pour vacciner contre une infection par de multiples bêta-coronavirus et contre des maladies provoquées par ces virus.
PCT/US2022/030726 2021-05-24 2022-05-24 Épitopes de lymphocytes t et compositions associées utiles dans la prévention, le diagnostic et le traitement de bêta-coronavirus WO2022251216A1 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
US202163192267P 2021-05-24 2021-05-24
US63/192,267 2021-05-24

Publications (1)

Publication Number Publication Date
WO2022251216A1 true WO2022251216A1 (fr) 2022-12-01

Family

ID=84229095

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/US2022/030726 WO2022251216A1 (fr) 2021-05-24 2022-05-24 Épitopes de lymphocytes t et compositions associées utiles dans la prévention, le diagnostic et le traitement de bêta-coronavirus

Country Status (1)

Country Link
WO (1) WO2022251216A1 (fr)

Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2004111081A2 (fr) * 2003-06-13 2004-12-23 Crucell Holland B.V. Peptides antigeniques de coronavirus de sars, et utilisations
US10973909B1 (en) * 2020-04-03 2021-04-13 Peptc Vaccines Limited Coronavirus vaccine
US20210260180A1 (en) * 2020-02-14 2021-08-26 Altimmune, Inc Coronavirus immunogenic compositions and uses thereof

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2004111081A2 (fr) * 2003-06-13 2004-12-23 Crucell Holland B.V. Peptides antigeniques de coronavirus de sars, et utilisations
US20210260180A1 (en) * 2020-02-14 2021-08-26 Altimmune, Inc Coronavirus immunogenic compositions and uses thereof
US10973909B1 (en) * 2020-04-03 2021-04-13 Peptc Vaccines Limited Coronavirus vaccine

Non-Patent Citations (3)

* Cited by examiner, † Cited by third party
Title
MOISE LEONARD, PRINCIOTTA MICHAEL F, MEYERS LAUREN M, GUTIERREZ ANDRES H, BOYLE CHRISTINE M, TERRY FRANCES, MCGONNIGAL BETHANY G, : "Highly conserved, non-human-like, and cross-reactive SARS-CoV-2 T cell epitopes for COVID-19 vaccine design and validation ", NPJ VACCINES, 13 May 2021 (2021-05-13), XP093006723, Retrieved from the Internet <URL:https://epivax.com/wp-content/uploads/2021/09/ISV_COVID-19_poster_02Sep21.pdf> [retrieved on 20221209] *
SLATHIA PARVEZ SINGH, SHARMA PREETI: "Prediction of T and B Cell Epitopes in the Proteome of SARS-CoV-2 for Potential Use in Diagnostics and Vaccine Design", CHEMRXIV, 15 April 2020 (2020-04-15), pages 1 - 20, XP055865887, Retrieved from the Internet <URL:https://chemrxiv.org/engage/chemrxiv/article-details/60c749deee301c4145c79b4f> [retrieved on 20211125], DOI: 10.26434/chemrxiv.12116943.v1 *
SYED FARAZ AHMED, AHMED A. QUADEER, MATTHEW R. MCKAY: "Preliminary Identification of Potential Vaccine Targets for the COVID-19 Coronavirus (SARS-CoV-2) Based on SARS-CoV Immunological Studies", VIRUSES, vol. 12, no. 3, pages 254, XP055716518, DOI: 10.3390/v12030254 *

Similar Documents

Publication Publication Date Title
US20230190915A1 (en) T cell epitopes and related compositions useful in the prevention, diagnosis, and treatment of covid-19
AU2021237720A1 (en) Coronavirus vaccines and methods of use
Lim et al. A comparative characterization of SARS-CoV-2-specific T cells induced by mRNA or inactive virus COVID-19 vaccines
JP2023524054A (ja) ベータコロナウイルスの予防と治療
US10144766B2 (en) Cytotoxic T Lymphocyte inducing immunogens for prevention treatment and diagnosis of dengue virus infection
Gupta et al. SARS coronavirus nucleocapsid immunodominant T-cell epitope cluster is common to both exogenous recombinant and endogenous DNA-encoded immunogens
JP2022513021A (ja) 向上した酸化還元酵素モチーフを有する免疫原性ペプチド
EP2953966B1 (fr) Induction d&#39;une réponse cellulaire à réactivité croisée contre des antigènes de rhinovirus
US20140302124A1 (en) Cytotoxic T Lymphocyte Inducing Immunogens For Prevention Treatment and Diagnosis of INFLUENZA VIRUS INFECTION
US20230192777A1 (en) Epitopic vaccine for african swine fever virus
US20100080817A1 (en) HUMAN PAPILLOMAVIRUS / Ii-KEY HYBRIDS AND METHODS OF USE
WO2022251216A1 (fr) Épitopes de lymphocytes t et compositions associées utiles dans la prévention, le diagnostic et le traitement de bêta-coronavirus
US20230181721A1 (en) Vaccine against sars-cov virus
WO2023004307A1 (fr) Épitopes de lymphocytes t et compositions associées utiles dans la prévention et le traitement d&#39;une infection par le virus respiratoire syncytial
CA3181635A1 (fr) Peptides immunogenes avec de nouveaux motifs d&#39;oxydoreductase
US20220023412A1 (en) Compositions Useful in Both Homologous And Heterologous Vaccine Regimens
US20220370604A1 (en) Peptide vaccine for virus infection
JP2023538002A (ja) 制御性ｔ細胞エピトープ
WO2023081861A1 (fr) Expression améliorée par l&#39;intermédiaire d&#39;autotransporteurs
JP2023520989A (ja) １型糖尿病の予防および治療に有用なｔレジトープ構築物
US20130345152A1 (en) Compositions comprising recombinant cowpox virus protein cpxv014
JPWO2022036193A5 (fr)
JPWO2021195508A5 (fr)

Legal Events

Date Code Title Description
121 Ep: the epo has been informed by wipo that ep was designated in this application

Ref document number: 22811984

Country of ref document: EP

Kind code of ref document: A1

NENP Non-entry into the national phase

Ref country code: DE