WO2024036313A2 - Anti-sars-cov-2 antibodies and methods of use thereof - Google Patents

Anti-sars-cov-2 antibodies and methods of use thereof Download PDF

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Publication number
WO2024036313A2
WO2024036313A2 PCT/US2023/072092 US2023072092W WO2024036313A2 WO 2024036313 A2 WO2024036313 A2 WO 2024036313A2 US 2023072092 W US2023072092 W US 2023072092W WO 2024036313 A2 WO2024036313 A2 WO 2024036313A2
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seq
amino acid
acid sequence
antibody
antigen
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PCT/US2023/072092
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French (fr)
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WO2024036313A3 (en
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Michel C. Nussenzweig
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The Rockefeller University
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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K16/00—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies
    • C07K16/08—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from viruses
    • C07K16/10—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from viruses from RNA viruses
    • C07K16/1002—Coronaviridae
    • C07K16/1003—Severe acute respiratory syndrome coronavirus 2 [SARS‐CoV‐2 or Covid-19]

Definitions

  • the present invention relates to antibodies directed to epitopes of SARS-CoV-2 Coronavirus 2 (“SARS-CoV-2”).
  • SARS-CoV-2 SARS-CoV-2 coronavirus 2
  • the present invention further relates to the preparation and use of neutralizing antibodies directed to the SARS-CoV-2 spike (S) glycoproteins for the prevention and treatment of SARS-CoV-2 infection.
  • SARS-CoV-2 spike (S) glycoproteins for the prevention and treatment of SARS-CoV-2 infection.
  • VoC variants of concern
  • Several of these circulating variants have been designated variants of concern (VoC) and have led to successive waves of infection, most notably by VoCs Alpha (Supasa et al., 2021), Delta (Liu et al., 2021), Omicron (Dejnirattisai et al., 2022).
  • BA.2.12.1 variant (a BA.2 lineage) contributes 59% of new cases in the United States, while BA.4 and BA.5 caused a fifth wave of COVID-19 infection in South Africa. Nevertheless, vaccine- elicited immunity continues to provide robust protection against severe disease, even in the face of viral variants(Andrews et al., 2022; Madhi et al., 2022; Wolter et al., 2022; World Health, 2022).
  • the present disclosure provides an antibody or antigen-binding fragment thereof that binds to the S polypeptide of a Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) wherein the antibody or antigen-binding fragment thereof comprises: three heavy chain complementarity determining regions (HCDRs) (HCDR1, HCDR2, and HCDR3) of a heavy chain variable region (HCVR) having the amino acid sequence of SEQ ID NO: 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31, 33, 35, 37, 39, 41, 43, 45, 47, 49, 51, 53, 55, 57, 59, 61, 63, 65, 67, 69, 71, 73, 75, 77, 79, 81, 83, 85, 87, 89, 91, 93, 95, 97, 99, 101, 103, 105, 107, 109, 111, 113, 115, 117, 119, 121
  • HCDRs heavy chain complementarity
  • the antibody or antigen-binding fragment thereof comprises: a heavy chain variable region comprising an amino acid sequence with at least 75% identity to the amino acid sequence of SEQ ID NO: 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31, 33, 35, 37, 39, 41, 43, 45, 47, 49, 51, 53, 55, 57, 59, 61, 63, 65, 67, 69, 71, 73, 75, 77, 79, 81, 83, 85, 87, 89, 91, 93, 95, 97, 99, 101, 103, 105, 107, 109, 111, 113, 115, 117, 119, 121, 123, 125, 127, 129, 131, 133, 135, 137, 139, 141, 143, 145, 147, 149, 151, 153, 155, 157, 159, 161, 163, 165, 167, 169, 171, 17
  • the antibody or antigen-binding fragment thereof comprises a HCVR and a LCVR that comprise a HCVR/LCVR amino acid sequence pair of SEQ ID NOs: 1- 2, 3-4, 5-6, 7-8, 9-10, 11-12, 13-14, 15-16, 17-18, 19-20, 21-22, 23-24, 25-26, 27-28, 29-30, 31- 32, 33-34, 35-36, 37-38, 39-40, 41-42, 43-44, 45-46, 47-48, 49-50, 51-52, 53-54, 55-56, 57-58, 59-60, 61-62, 63-64, 65-66, 67-68, 69-70, 71-72, 73-74, 75-76, 77-78, 79-80, 81-82, 83-84, 85-86, 87-88, 89-90, 91-92, 93-94, 95-96, 97-98, 99-100, 101-102
  • the antibody or antigen-binding fragment thereof comprises a HCVR and a LCVR that comprise a HCVR/LCVR amino acid sequence pair with at least 75% identity to the amino acid sequence of SEQ ID NOs: 1-2, 3-4, 5-6, 7-8, 9-10, 11-12, 13-14, 15-16, 17-18, 19-20, 21-22, 23-24, 25-26, 27-28, 29-30, 31-32, 33-34, 35-36, 37-38, 39-40, 41-42, 43-44, 45-46, 47-48, 49-50, 51-52, 53-54, 55-56, 57-58, 59-60, 61-62, 63-64, 65-66, 67-68, 69-70, 71-72, 73-74, 75-76, 77-78, 79-80, 81-82, 83-84, 85-86, 87-88, 89-90, 91-92, 93-94, 95-96,
  • the present disclosure provides an antibody or antigen-binding fragment thereof that binds to the S polypeptide of a Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), wherein the antibody or antigen-binding fragment thereof comprises: (a) a variable heavy chain CDR1 (HCDR1) comprising an amino acid sequence selected from SEQ ID NO: 1354, 1360, 1366, 1372, and 1378; (b) a variable heavy chain CDR2 (HCDR2) comprising an amino acid sequence of SEQ ID NO: 1355, 1361, 1367, 1373, and 1379; (c) a variable heavy chain CDR3 (HCDR3) comprising an amino acid sequence selected from SEQ ID NO: 1356, 1362, 1368, 1374, and 1380; (d) a variable light chain CDR1 (LCDR1) comprising an amino acid sequence selected from SEQ ID NO: 1357, 1363, 1369, 1375, and 1381; (e) a variable light chain CDR
  • HCDR1 comprising
  • the antibody or antigen-binding fragment thereof comprises: a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1354, a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1355, a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1356, a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1357, a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1358, and a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1359.
  • the antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising an amino acid sequence that is at least about 75%, at least 85%, at least 90%, at least 95%, at least 99%, or 100% identical to SEQ ID NO: 25, and a light chain variable domain (VL) comprising an amino acid sequence that is at least about 75%, at least 85%, at least 90%, at least 95%, at least 99%, or 100% identical to SEQ ID NO: 26.
  • VH heavy chain variable domain
  • VL light chain variable domain
  • the antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising or consisting of the amino acid sequence of SEQ ID NO: 25, and a light chain variable domain (VL) comprising or consisting of the amino acid sequence of SEQ ID NO: 26.
  • VH heavy chain variable domain
  • VL light chain variable domain
  • the antibody or antigen-binding fragment thereof comprises a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1360, a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1361, a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1362, a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1363, a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1364, and a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1365.
  • the antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising an amino acid sequence that is at least about 75%, at least 85%, at least 90%, at least 95%, at least 99%, or 100% identical to SEQ ID NO: 395, and a light chain variable domain (VL) comprising an amino acid sequence that is at least about 75%, at least 85%, at least 90%, at least 95%, at least 99%, or 100% identical to SEQ ID NO: 396.
  • VH heavy chain variable domain
  • VL light chain variable domain
  • the antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising or consisting of the amino acid sequence of SEQ ID NO: 395, and a light chain variable domain (VL) comprising or consisting of the amino acid sequence of SEQ ID NO: 396.
  • VH heavy chain variable domain
  • VL light chain variable domain
  • the antibody or antigen-binding fragment thereof comprises a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1366, a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1367, a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1368, a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1369, a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1370, and a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1371.
  • the antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising an amino acid sequence that is at least about 75%, at least 85%, at least 90%, at least 95%, at least 99%, or 100% identical to SEQ ID NO: 397, and a light chain variable domain (VL) comprising an amino acid sequence that is at least about 75%, at least 85%, at least 90%, at least 95%, at least 99%, or 100% identical to SEQ ID NO: 398.
  • VH heavy chain variable domain
  • VL light chain variable domain
  • the antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising or consisting of the amino acid sequence of SEQ ID NO: 397, and a light chain variable domain (VL) comprising or consisting of the amino acid sequence of SEQ ID NO: 398.
  • VH heavy chain variable domain
  • VL light chain variable domain
  • the antibody or antigen-binding fragment thereof comprises a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1372, a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1373, a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1374, a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1375, a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1376, and a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1377.
  • the antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising an amino acid sequence that is at least about 75%, at least 85%, at least 90%, at least 95%, at least 99%, or 100% identical to SEQ ID NO: 191, and a light chain variable domain (VL) comprising an amino acid sequence that is at least about 75%, at least 85%, at least 90%, at least 95%, at least 99%, or 100% identical to SEQ ID NO: 192.
  • VH heavy chain variable domain
  • VL light chain variable domain
  • the antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising or consisting of the amino acid sequence of SEQ ID NO: 191, and a light chain variable domain (VL) comprising or consisting of the amino acid sequence of SEQ ID NO: 192.
  • VH heavy chain variable domain
  • VL light chain variable domain
  • the antibody or antigen-binding fragment thereof comprises a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1378, a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1379, a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1380, a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1381, a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1382, and a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1383.
  • the antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising an amino acid sequence that is at least about 75%, at least 85%, at least 90%, at least 95%, at least 99%, or 100% identical to SEQ ID NO: 219, and a light chain variable domain (VL) comprising an amino acid sequence that is at least about 75%, at least 85%, at least 90%, at least 95%, at least 99%, or 100% identical to SEQ ID NO: 220.
  • VH heavy chain variable domain
  • VL light chain variable domain
  • the antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising or consisting of the amino acid sequence of SEQ ID NO: 219, and a light chain variable domain (VL) comprising or consisting of the amino acid sequence of SEQ ID NO: 220.
  • VH heavy chain variable domain
  • VL light chain variable domain
  • the antibody or antigen-binding fragment thereof is a fully human antibody or antigen-binding fragment thereof.
  • the antibody or antigen-binding fragment thereof binds to an epitope within the receptor binding domain (RBD) or the N-terminal domain (NTD) of the S polypeptide.
  • the antibody or antigen-binding fragment thereof binds to an epitope selected from the following: (a) the supersite b-hairpin (residues 152-158 of SEQ ID NO: 1353); (b) WKH ⁇ -strand (residue 97-102 of SEQ ID NO: 1353); (c) N4-loop (residues 178-188 of SEQ ID NO: 1353); (d) N-linked glycans at positions N122 and N149 of the spike polypeptide of SEQ ID NO: 1353; (e) residues 27-32, 57-60, 210-218, and/or 286-303 of the spike polypeptide of SEQ ID NO: 1353; and/or (f) residues 600-606 of the spike polypeptide of SEQ ID NO: 1353.
  • an epitope selected from the following: (a) the supersite b-hairpin (residues 152-158 of SEQ ID NO: 1353); (b) WKH ⁇
  • the present disclosure provides an antibody or antigen-binding fragment thereof, wherein the antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising an amino acid sequence that is at least about 75%, at least 85%, at least 90%, at least 95%, at least 99%, or 100% identical to a SEQ ID NO selected from column 7 of Table 2, and a light chain variable domain (VL) comprising an amino acid sequence that is at least about 75%, at least 85%, at least 90%, at least 95%, at least 99%, or 100% identical to a SEQ ID NO selected from column 11 of Table 2, wherein the VH SEQ ID NO and the VL SEQ ID NO are selected from the same row.
  • VH heavy chain variable domain
  • VL light chain variable domain
  • the antibody or antigen- binding fragment thereof comprises a heavy chain variable domain (VH) comprising or consisting of a SEQ ID NO selected from column 7 of Table 2, and a light chain variable domain (VL) comprising or consisting of a SEQ ID NO selected from column 11 of Table 2, wherein the VH SEQ ID No and the VL SEQ ID NO are selected from the same row.
  • VH heavy chain variable domain
  • VL light chain variable domain
  • the antibody or antigen-binding fragment thereof neutralizes SARS-CoV-2.
  • the antibody or antigen-binding fragment thereof inhibits the fusion of SARS-CoV-2 and a host cell membrane.
  • the antibody or antigen-binding fragment thereof is cytotoxic to a SARS-CoV-2 infected host cell.
  • the antibody or antigen-binding fragment thereof is a multivalent antibody comprising (a) a first target binding site that specifically binds to an epitope within the spike polypeptide, and (b) a second target binding site that binds to an epitope on a different epitope on the spike polypeptide or a different molecule.
  • the antibody or antigen-binding fragment thereof further comprises a variant Fc constant region.
  • the antibody or antigen-binding fragment thereof is a monoclonal antibody, a chimeric antibody, a humanized antibody, or humanized antibody. In some embodiments, the antibody or antigen-binding fragment thereof is a single-chain antibody, Fab, or Fab2 fragment. [0030] In some embodiments, the antibody or antigen-binding fragment thereof is detectably labeled or conjugated to a toxin, a therapeutic agent, a polymer, a receptor, an enzyme, or a receptor ligand, preferably wherein the polymer is polyethylene glycol (PEG).
  • PEG polyethylene glycol
  • the present disclosure provides a polynucleotide encoding the antibody or antigen-binding fragment thereof described herein.
  • the present disclosure provides a vector comprising a polynucleotide described herein.
  • the present disclosure provides a cultured host cell comprising a vector described herein.
  • the present disclosure provides a pharmaceutical composition comprising an antibody or antigen-binding fragment thereof, polynucleotide, or vector described herein.
  • the present disclosure provides a pharmaceutical composition
  • a pharmaceutical composition comprising two or more antibodies or antigen-binding fragments thereof, wherein the two or more antibodies or antigen-binding fragments thereof are selected from: a HCVR/LCVR amino acid sequence pair of SEQ ID NOs: 1-2, 3-4, 5-6, 7-8, 9-10, 11-12, 13-14, 15-16, 17-18, 19-20, 21-22, 23-24, 25-26, 27-28, 29-30, 31-32, 33-34, 35-36, 37-38, 39-40, 41-42, 43-44, 45-46, 47-48, 49-50, 51-52, 53-54, 55-56, 57-58, 59-60, 61-62, 63-64, 65-66, 67-68, 69-70, 71-72, 73-74, 75-76, 77-78, 79-80, 81-82, 83-84, 85-86, 87-88, 89-90, 91-92,
  • the two or more of the antibody or antigen-binding fragment thereof comprise: a first antibody or antigen-binding fragment thereof comprising a HCVR/LCVR amino acid sequence pair with at least 75% identity to the amino acid sequence of SEQ ID NOs: 1-2, 3-4, 5-6, 7-8, 9-10, 11-12, 13-14, 15-16, 17-18, 19-20, 21-22, 23-24, 25-26, 27-28, 29-30, 31- 32, 33-34, 35-36, 37-38, 39-40, 41-42, 43-44, 45-46, 47-48, 49-50, 51-52, 53-54, 55-56, 57-58, 59-60, 61-62, 63-64, 65-66, 67-68, 69-70, 71-72, 73-74, 75-76, 77-78, 79-80, 81-82, 83-84, 85-86, 87-88, 89-90, 91-92, 93
  • the present disclosure provides a pharmaceutical composition comprising two or more antibodies or antigen-binding fragments thereof, wherein the two or more antibodies or antigen-binding fragments thereof are selected from: (a) an antibody or antigen- binding fragment thereof comprising: a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1354, a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1355, a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1356, a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1357, a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1358, and a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1359; (b) an antibody or antigen-binding fragment thereof comprising: a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1360, a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1361, a HCDR
  • the two or more monoclonal antibodies or antigen-binding fragments thereof are selected from: (a) an antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising or consisting of the amino acid sequence of SEQ ID NO: 25, and a light chain variable domain (VL) comprising or consisting of the amino acid sequence of SEQ ID NO: 26; (b) an antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising or consisting of the amino acid sequence of SEQ ID NO: 395, and a light chain variable domain (VL) comprising or consisting of the amino acid sequence of SEQ ID NO: 396; (c) an antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising or consisting of the amino acid sequence of SEQ ID NO: 397, and a light chain variable domain (VL) comprising or consisting of the amino acid sequence of SEQ ID NO: 398; (d) an antibody or antigen
  • the pharmaceutical composition further comprises a pharmaceutically acceptable carrier.
  • the present disclosure provides a pharmaceutical composition for use in treating a SARS-CoV-2 infection in a subject.
  • the present disclosure provides a method of treating a SARS-CoV- 2 infection in a subject, comprising administering to the subject therapeutically effective amount of an antibody or antigen-binding fragment thereof or a pharmaceutical composition thereof.
  • the present disclosure provides a method of neutralizing SARS-CoV-2 in a subject, comprising administering to a subject in need thereof a therapeutically effective amount of an antibody or antigen-binding fragment thereof or a pharmaceutical composition thereof.
  • the present disclosure provides a method of preventing a SARS-CoV-2 infection in a subject, comprising administering to a subject in need thereof a therapeutically effective amount of an antibody or antigen-binding fragment thereof or pharmaceutical composition thereof.
  • the subject is immunocompromised.
  • the method further comprises administering a second therapeutic agent, preferably wherein the second therapeutic agent comprises an anti-inflammatory drug or an antiviral compound.
  • the antiviral compound comprises: a nucleoside analog, a peptoid, an oligopeptide, a polypeptide, a protease inhibitor, a 3C-like protease inhibitor, a papain-like protease inhibitor, or an inhibitor of an RNA dependent RNA polymerase; or (b) the antiviral compound is selected from: acyclovir, ganciclovir, vidarabine, foscarnet, cidofovir, amantadine, ribavirin, trifluorothymidine, zidovudine, didanosine, zalcitabine, and an interferon, and preferably wherein the interferon is an interferon- ⁇ RU ⁇ DQ ⁇ LQWHUIHURQ- ⁇ In some embodiments, the second therapeutic agent is administered before, after, or concurrently with the antibody or pharmaceutical composition thereof.
  • the pharmaceutical composition is administered to the subject after the exposure to SARS-CoV-2.
  • the present disclosure provides a kit for detecting a SARS-CoV- 2 infection in a subject, comprising an antibody or antigen-binding fragment thereof described herein.
  • the present disclosure provides a method of detecting the presence of a SARS-CoV-2 in a sample, comprising (1) contacting the sample with an antibody or antigen- binding fragment thereof described herein, and (2) detecting the presence of an antibody-antigen complex, wherein the presence of the antibody-antigen complex indicates the presence of SARS- CoV-2.
  • the antibody or antigen-binding fragment thereof is conjugated to a label, preferably wherein the label is selected from a fluorescent label, a chemiluminescent label, a radiolabel, and an enzyme.
  • the method further comprises contacting a secondary antibody with the antibody or antigen-binding fragment thereof, wherein the secondary antibody comprises a label.
  • the method further comprises detecting fluorescence or chemiluminescence of the label, preferably wherein the step of detecting comprises a competitive binding assay or ELISA.
  • the method further comprises binding the sample to a solid support, preferably wherein the solid support is selected from microparticles, microbeads, magnetic beads, and an affinity purification column.
  • the sample is a blood sample, a nasal swab, or a throat swab.
  • the present disclosure provides a method of preparing an antibody, or antigen-binding fragment thereof, comprising: (a) obtaining a cultured host cell described herein; (b) culturing the cultured host cell in a medium under conditions permitting expression of a polypeptide encoded by the vector and assembling of an antibody or fragment thereof; and (c) purifying the antibody or fragment from the cultured cell or the medium of the cell.
  • the present disclosure provides a kit comprising a pharmaceutically acceptable dose unit of an antibody or antigen-binding fragment thereof or pharmaceutical composition described herein.
  • the present disclosure provides a kit for the diagnosis, prognosis or monitoring treatment of SARS-CoV-2 in a subject, comprising: an antibody or antigen-binding fragment thereof described herein; and a least one detection reagent that binds specifically to the antibody or antigen-binding fragment thereof described herein.
  • the present disclosure provides a method of neutralizing SARS- CoV-2 in a subject, comprising administering to a subject in need thereof a therapeutically effective amount of two or more antibodies or antigen-binding fragments thereof described herein.
  • the present disclosure provides a method of treating a SARS-CoV- 2 in a subject in need thereof, comprising administering to a subject in need thereof a therapeutically effective amount of two or more antibodies or antigen-binding fragments thereof described herein.
  • the present disclosure provides a method of preventing a SARS- CoV-2 in a subject in need thereof, comprising administering to a subject in need thereof a therapeutically effective amount of two or more antibodies or antigen-binding fragments thereof described herein.
  • the two or more antibodies or antigen-binding fragments thereof are selected from: (a) an antibody or antigen-binding fragment thereof comprising: a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1354, a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1355, a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1356, a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1357, a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1358, and a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1359; (b) an antibody or antigen-binding fragment thereof comprising: a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1360, a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1361, a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1362, a LCDR1 comprising an amino acid sequence of SEQ ID NO:
  • the two or more monoclonal antibodies or antigen- binding fragments thereof are selected from: (a) an antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising or consisting of the amino acid sequence of SEQ ID NO: 25, and a light chain variable domain (VL) comprising or consisting of the amino acid sequence of SEQ ID NO: 26; (b) an antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising or consisting of the amino acid sequence of SEQ ID NO: 395, and a light chain variable domain (VL) comprising or consisting of the amino acid sequence of SEQ ID NO: 396; (c) an antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising or consisting of the amino acid sequence of SEQ ID NO: 397, and a light chain variable domain (VL) comprising or consisting of the amino acid sequence of SEQ ID NO: 398; (d) an antibody or antigen-
  • the first antibody or antigen-binding fragment thereof and the second antibody or antigen-binding fragment thereof exhibit synergistic activity.
  • the method further comprises administering to the subject a therapeutically effective amount of a second therapeutic agent or therapy.
  • the second therapeutic agent comprises an anti-inflammatory drug or an antiviral compound.
  • the antiviral compound comprises: a nucleoside analog, a peptoid, an oligopeptide, a polypeptide, a protease inhibitor, a 3C-like protease inhibitor, a papain-like protease inhibitor, or an inhibitor of an RNA dependent RNA polymerase; or (b) the antiviral compound is selected from: acyclovir ganciclovir, vidarabine, foscarnet, cidofovir, amantadine, ribavirin, trifluorothymidine, zidovudine, didanosine, zalcitabine, and an interferon, and preferably wherein the interferon is an interferon- ⁇ RU ⁇ DQ ⁇ LQWHUIHURQ- ⁇ [0055]
  • the first antibody or antigen-binding fragment thereof is administered before, after, or concurrently with the second antibody or antigen-binding fragment thereof.
  • the antibody or antigen-binding fragment thereof is administered to the subject intravenously, subcutaneously, or intraperitoneally.
  • FIGs. 1a, 1b, and 1c show plasma ELISAs and neutralizing activity of the antibodies.
  • FIG. 1a, 1b, and 1c show plasma ELISAs and neutralizing activity of the antibodies.
  • 1a is a diagram showing blood donation schedules for vaccinated-only individuals 5 months after the 2nd dose (Vax2, top) (Cho et al., 2021), Delta breakthrough infection after Vax2 (Delta BT, 2nd from top), and vaccinated-only individuals 1 month after the 3rd dose (Vax3, 2nd from bottom)(Muecksch et al., 2022) and Omicron breakthrough infection after Vax3 (Omicron BT, bottom).
  • FIGs.1a, 1b, and 1c show characterization of anti-SARS-CoV-2 RBD memory B cells after breakthrough infection.
  • FIG.2a, 2b, 2c, 2d, and 2e show characterization of anti-SARS-CoV-2 RBD memory B cells after breakthrough infection.
  • FIG. 2a shows representative flow cytometry plots indicating PE-WT-RBD and AlexaFluor-647-WT-RBD binding memory B cells from 4 individuals after Delta breakthrough infection following Vax2 (Delta BT), 2 individuals 1 month after Vax3, and 6 individuals after Omicron BA.1 breakthrough infection following Vax3 (Omicron BT).
  • FIG. 2d depicts pie charts showing the distribution of IgG antibody sequences obtained from WT-specific memory B cells from: 2 individuals assayed sequentially 1 month after the 3rd mRNA dose (Vax3) an Omicron infection (left); 4 individuals after Delta breakthrough (Delta), and 4 individuals after Omicron breakthrough (Omicron).
  • the number inside the circle indicates the number of sequences analyzed for the individual denoted above the circle.
  • Pie slice size is proportional to the number of clonally related sequences.
  • the black outline and associated numbers indicate the percentage of clonal sequences detected at each time point.
  • Patterned slices indicate persisting clones (same IGHV and IGLV genes, with highly similar CDR3s) found at more than one timepoint within the same individual.
  • Grey slices indicate clones unique to the timepoint.
  • White slices indicate sequences isolated only once per time point.
  • FIG. 2e shows number of nucleotide somatic hypermutations (SHM) in IGHV + IGLV in WT- RBD-specific sequences after Delta or Omicron breakthrough infection, compared to 5 months after Vax2, and 1 month after Vax3. The bars and numbers in FIG. 2b and FIG.
  • FIG. 2c represent geometric mean, and in FIG. 2e, represent median values.
  • FIG. 2e shows the results of statistical analysis in FIG. 2b and FIG. 2c that were determined by a two-tailed Kruskal-Wallis test with subsequent Dunn’s multiple-comparisons test and in FIG.2e by two-tailed Mann-Whitney test.
  • FIGs. 3a, 3b, 3c, 3d, 3e, and 3f show characterization of anti-SARS-CoV-2 RBD monoclonal antibodies.
  • FIG. 3b is a graph showing anti-SARS-CoV-2 neutralizing activity of monoclonal antibodies measured by a SARS-CoV-2 pseudotype virus neutralization assay using WT SARS-CoV-2 pseudovirus. IC50 values for all antibodies, including the 288 reported and tested herein, and 350 previously reported (Cho et al., 2021; Muecksch et al., 2022).
  • FIGs. 3c-d are graphs showing IC50s of monoclonal antibodies against WT, Delta-RBD, and Omicron BA.1 SARS-CoV-2 pseudoviruses.
  • FIG. 3e shows ring plots depicting fraction of neutralizing (IC50 ⁇ 1000ng/ml) antibodies against WT, Delta-RBD, and Omicron BA.1 SARS- CoV-2 pseudoviruses, and non-neutralizing (IC50>1000 ng/ml) antibodies from each time point.
  • 3f shows ring plots depicting fraction of mAbs that are neutralizing (IC501-1000 ng/mL, white), or non-neutralizing (IC50>1000 ng/mL, black) against Omicron BA.4/5. Number in inner circles indicates number of antibodies tested.
  • the deletions/substitutions corresponding to viral variants used in Figs. 3c-f were incorporated into a spike protein that also includes the R683G substitution, which disrupts the furin cleavage site and increases particle infectivity.
  • Neutralizing activity against mutant pseudoviruses was compared to a wild-type (WT) SARS-CoV-2 spike sequence (NC_045512), carrying R683G where appropriate.
  • WT wild-type
  • NC_045512 SARS-CoV-2 spike sequence
  • FIGs. 3a, 3b, and 3d represent geometric mean values. Statistical significance in FIGs.3a and 3b was determined by a two-tailed Kruskal-Wallis test with subsequent Dunn’s multiple comparisons, in FIG.3c was determined by a two-tailed Wilcoxon test and in FIG. 3d was determined by a two-tailed Mann- Whitney test. [0060] FIGs. 4a, 4b, and 4c show the results of plasma ELISA. FIGs. 4a-b are graphs showing area under the curve (AUC) for plasma IgG binding to FIG.4a, SARS-CoV-2 Delta-RBD and FIG.
  • AUC area under the curve
  • FIGs. 4a, 5b, 5c, 5d, 5e, and 5f show the results of flow cytometry.
  • FIGs. 5a, 5b, 5c, 5d, 5e, and 5f show the results of flow cytometry.
  • FIGs.5c-e are graphs showing the frequency of IgM, IgG, and IgA isotype expression in FIG. 5c, WT RBD+ MBCs, FIG. 5d, WT+Delta+ RBD binding MBCs, FIG.
  • FIG. 5e WT+Omicron BA.1+ RBD binding MBCs cells.
  • FIG. 5f shows gating strategy for single-cell sorting for CD20+ B cells for WT RBD-PE and RBD-AF647.
  • Statistical significance in FIG. 5c was determined by a two-tailed Kruskal-Wallis test with subsequent Dunn’s multiple comparisons.
  • FIGs. 6a, 6b, 6c, 6d, 6e, 6f, 6g show frequency distribution of human V genes and antibody gene somatic hypermutations analysis.
  • FIG. 6a-c shows comparison of the frequency distribution of human V genes for heavy chain and light chains of anti-RBD antibodies from this study and from a database of shared clonotypes of human B cell receptor generated by Cinque Soto et al. (Soto et al., 2019).
  • the graphs show relative abundance of human IGHV (left panel), IGKV (middle panel) and IGLV (right panel) genes in Sequence Read Archive accession SRP010970 (bottom), antibodies obtained from Delta breakthrough infection (top), and Vax2 (middle).
  • FIGs.6d-f same as FIG.6a.
  • FIG. 6g shows number of nucleotide somatic hypermutations (SHM) in IGHV and IGLV in WT-RBD-specific sequences, separately after Delta or Omicron breakthrough infection, to Vax2 (Cho et al., 2021), and Vax3 (Muecksch et al., 2022).
  • the bars and numbers in FIG.6g represent the median value.
  • FIGs. 7a and 7b show mAb affinity and epitopes of the antibodies.
  • FIG. 7a is a graph showing affinity measurements (KDs) for WT RBD measured by BLI for antibodies cloned from vaccinated individuals after Delta or Omicron breakthrough infection, compared to Vax2 (Cho et al., 2021), and Vax3(Muecksch et al., 2022).
  • FIG.7a shows neutralizing breadth of the antibodies. Ring plots show fraction of mAbs in FIGs.
  • 3c-e that are neutralizing (IC50 1-1000 ng/mL, white), or non-neutralizing (IC50>1000 ng/mL, black) for mutant or variant SARS-CoV-2 pseudovirus indicated across the top at the time point indicated to the left.
  • the number inside the circle indicates the number of antibodies tested.
  • the deletions/substitutions corresponding to viral variants were incorporated into a spike protein that also includes the R683G substitution, which disrupts the furin cleavage site and increases particle infectivity.
  • Neutralizing activity against mutant pseudoviruses was compared to a wild- type (WT) SARS-CoV-2 spike sequence (NC_045512), carrying R683G where appropriate. All experiments were performed at least in duplicate and repeated twice.
  • SARS-CoV-2 represents a serious public health concern. Methods to diagnose and treat persons who are infected with SARS-CoV-2 provide the opportunity to either prevent or control further spread of infection by SARS-CoV-2. These methods are especially important due to the ability of SARS-CoV-2 to infect persons through an airborne route.
  • This invention is based, at least in part, on unexpected neutralizing activities of the disclosed anti-SARS-CoV-2 antibodies or antigen-binding fragments thereof. These antibodies and antigen-binding fragments constitute a novel therapeutic strategy in protection from SARS-CoV-2 infections. 1.1.
  • a “neutralizing antibody” is one that can neutralize the ability of that pathogen to initiate and/or perpetuate an infection in a host and/or in target cells in vitro.
  • “Broadly neutralizing anti- SARS-CoV-2 antibodies” refer to antibodies that neutralize more than one SARS-CoV-2 virus strains/variants in a neutralization assay.
  • a broad neutralizing anti-SARS-CoV-2 antibody may neutralize at least 2, 3, 4, 5, 6, 7, 8, 9 or more different strains/variants of SARS-CoV-2.
  • the term “antibody” as referred to herein includes whole antibodies and any antigen- binding fragment or single chains thereof.
  • Whole antibodies are glycoproteins comprising at least two heavy (H) chains and two light (L) chains inter-connected by disulfide bonds.
  • Each heavy chain is comprised of a heavy chain variable region (abbreviated herein as VH) and a heavy chain constant region.
  • the heavy chain constant region is comprised of three domains, CH1, CH2, and CH3.
  • Each light chain is comprised of a light chain variable region (abbreviated herein as VL) and a light chain constant region.
  • the light chain constant region is comprised of one domain, CL.
  • the VH and VL regions can be further subdivided into regions of hypervariability, termed complementarity determining regions (CDR), interspersed with regions that are more conserved, termed framework regions (FR).
  • CDR complementarity determining regions
  • Each VH and VL is composed of three CDRs and four FRs, arranged from amino-terminus to carboxy-terminus in the following order: FR1, CDR1, FR2, CDR2, FR3, CDR3, FR4.
  • the heavy chain variable region CDRs and FRs are HFRl, HCDRl, HFR2, HCDR2, HFR3, HCDR3, HFR4.
  • the light chain variable region CDRs and FRs are LFR1, LCDR1, LFR2, LCDR2, LFR3, LCDR3, LFR4.
  • the variable regions of the heavy and light chains contain a binding domain that interacts with an antigen.
  • the constant regions of the antibodies can mediate the binding of the immunoglobulin to host tissues or factors, including various cells of the immune system (e.g., effector cells) and the first component (CIq) of the classical complement system.
  • the term “antigen-binding fragment or portion” of an antibody refers to one or more fragments of an antibody that retain the ability to specifically bind to an antigen (e.g., a spike or S protein of SARS-CoV-2 virus). It has been shown that the antigen-binding function of an antibody can be performed by fragments of a full-length antibody.
  • binding fragments encompassed within the term “antigen- binding fragment or portion” of an antibody include (i) a Fab fragment, a monovalent fragment consisting of the VL, VH, CL, and CHI domains; (ii) a F(ab')2 fragment, a bivalent fragment comprising two Fab fragments linked by a disulfide bridge at the hinge region; (iii) a Fab' fragment, which is essentially a Fab with part of the hinge region (see, FUNDAMENTAL IMMUNOLOGY (Paul ed., 3rd ed.
  • the two domains of the Fv fragment, VL and VH are coded for by separate genes, they can be joined, using recombinant methods, by a synthetic linker that enables them to be made as a single protein chain in which the VL and VH regions pair to form monovalent molecules (known as single chain Fv or scFv); see, e.g., Bird et al. (1988) Science 242:423-426; and Huston et al. (1988) Proc. Natl. Acad. Sci. USA 85:5879-5883).
  • Such single chain antibodies are also intended to be encompassed within the term “antigen-binding fragment or portion” of an antibody.
  • antigen refers to a molecule or a portion of a molecule capable of being bound by an antibody or an antigen-binding fragment thereof and additionally capable of being used in an animal to produce antibodies capable of binding to an epitope of that antigen.
  • An antigen may be a polypeptide, carbohydrate, nucleic acid, lipid, hapten, or other naturally occurring or synthetic compound.
  • an “isolated antibody,” as used herein, is intended to refer to an antibody that is substantially free of other antibodies having different antigenic specificities (e.g., an isolated antibody that specifically binds to a spike or S protein of SARS-CoV-2 virus is substantially free of antibodies that specifically bind antigens other than the neuraminidase).
  • An isolated antibody can be substantially free of other cellular material and/or chemicals.
  • the terms “monoclonal antibody” or “monoclonal antibody composition” as used herein refer to a preparation of antibody molecules of single molecular composition. A monoclonal antibody composition displays a single binding specificity and affinity for a particular epitope.
  • human antibody is intended to include antibodies having variable regions in which both the framework and CDR regions are derived from human germline immunoglobulin sequences. Furthermore, if the antibody contains a constant region, the constant region also is derived from human germline immunoglobulin sequences.
  • the human antibodies of the invention can include amino acid residues not encoded by human germline immunoglobulin sequences (e.g., mutations introduced by random or site-specific mutagenesis in vitro or by somatic mutation in vivo).
  • the term “human antibody,” as used herein is not intended to include antibodies in which CDR sequences derived from the germline of another mammalian species, such as a mouse, have been grafted onto human framework sequences.
  • human monoclonal antibody refers to antibodies displaying a single binding specificity, which have variable regions in which both the framework and CDR regions are derived from human germline immunoglobulin sequences.
  • the human monoclonal antibodies can be produced by a hybridoma that includes a B cell obtained from a transgenic nonhuman animal, e.g., a transgenic mouse, having a genome comprising a human heavy chain transgene and a light chain transgene fused to an immortalized cell.
  • recombinant human antibody includes all human antibodies that are prepared, expressed, created, or isolated by recombinant means, such as (a) antibodies isolated from an animal (e.g., a mouse) that is transgenic or transchromosomal for human immunoglobulin genes or a hybridoma prepared therefrom (described further below), (b) antibodies isolated from a host cell transformed to express the human antibody, e.g., from a transfectoma, (c) antibodies isolated from a recombinant, combinatorial human antibody library, and (d) antibodies prepared, expressed, created or isolated by any other means that involve splicing of human immunoglobulin gene sequences to other DNA sequences.
  • Such recombinant human antibodies have variable regions in which the framework and CDR regions are derived from human germline immunoglobulin sequences.
  • such recombinant human antibodies can be subjected to in vitro mutagenesis (or, when an animal transgenic for human Ig sequences is used, in vivo somatic mutagenesis) and thus the amino acid sequences of the VH and VL regions of the recombinant antibodies are sequences that, while derived from and related to human germline VH and VL sequences, may not naturally exist within the human antibody germline repertoire in vivo.
  • the term “isotype” refers to the antibody class (e.g., IgM or IgG1) that is encoded by the heavy chain constant region genes.
  • the phrases “an antibody recognizing an antigen” and “an antibody specific for an antigen” are used interchangeably herein with the term “an antibody which binds specifically to an antigen.”
  • the term “human antibody derivatives” refers to any modified form of the human antibody, e.g., a conjugate of the antibody and another agent or antibody.
  • the term “humanized antibody” is intended to refer to antibodies in which CDR sequences derived from the germline of another mammalian species, such as a mouse, have been grafted onto human framework sequences. Additional framework region modifications can be made within the human framework sequences.
  • chimeric antibody is intended to refer to antibodies in which the variable region sequences are derived from one species, and the constant region sequences are derived from another species, such as an antibody in which the variable region sequences are derived from a mouse antibody, and the constant region sequences are derived from a human antibody.
  • the term can also refer to an antibody in which its variable region sequence or CDR(s) is derived from one source (e.g., an IgA1 antibody), and the constant region sequence or Fc is derived from a different source (e.g., a different antibody, such as an IgG, IgA2, IgD, IgE or IgM antibody).
  • an effective amount refers to an amount of a therapeutic (e.g., a pharmaceutical composition provided herein) which is sufficient to treat, diagnose, prevent, delay the onset of, reduce and/or ameliorate the severity and/or duration of a given condition, disorder or disease and/or a symptom related thereto.
  • the term also encompasses an amount necessary for the reduction, slowing, or amelioration of the advancement or progression of a given disease, reduction, slowing, or amelioration of the recurrence, development or onset of a given disease, and/or to improve or enhance the prophylactic or therapeutic effect (s) of another therapy or to serve as a bridge to another therapy.
  • the invention encompasses isolated or substantially purified nucleic acids, peptides, polypeptides, or proteins.
  • an “isolated” nucleic acid, DNA or RNA molecule or an “isolated” polypeptide is a nucleic acid, DNA molecule, RNA molecule, or polypeptide that exists apart from its native environment and is therefore not a product of nature.
  • An isolated nucleic acid, DNA molecule, RNA molecule, or polypeptide may exist in a purified form or may exist in a non-native environment such as, for example, a transgenic host cell.
  • a “purified” nucleic acid molecule, peptide, polypeptide, or protein, or a fragment thereof, is substantially free of other cellular material, or culture medium when produced by recombinant techniques, or substantially free of chemical precursors or other chemicals when chemically synthesized.
  • an ⁇ isolated ⁇ nucleic acid is free of sequences that naturally flank the nucleic acid (i.e., sequences located at the 5' and 3’ ends of the nucleic acid) in the genomic DNA of the organism from which the nucleic acid is derived.
  • the isolated nucleic acid molecule can contain less than about 5 kb, 4 kb, 3 kb, 2 kb, 1 kb, 0.5 kb, or 0.1 kb of nucleotide sequences that naturally flank the nucleic acid molecule in genomic DNA of the cell from which the nucleic acid is derived.
  • a protein, peptide, or polypeptide that is substantially free of cellular material includes preparations of protein, peptide, or polypeptide having less than about 30%, 20%, 10%, or 5% (by dry weight) of contaminating protein.
  • culture medium represents less than about 30%, 20%, 10%, or 5% (by dry weight) of chemical precursors or non-protein-of-interest chemicals.
  • amino acid polymer that has been modified; for example, disulfide bond formation, glycosylation, lipidation, acetylation, phosphorylation, pegylation, or any other manipulation, such as conjugation with a labeling component.
  • amino acid includes natural and/or unnatural or synthetic amino acids, including glycine and both the D or L optical isomers, and amino acid analogs and peptidomimetics.
  • a peptide or polypeptide “fragment” as used herein refers to a less than full-length peptide, polypeptide, or protein.
  • a peptide or polypeptide fragment can have at least about 3, at least about 4, at least about 5, at least about 10, at least about 20, at least about 30, at least about 40 amino acids in length, or single unit lengths thereof.
  • fragment may be 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, or more amino acids in length.
  • peptide fragments can be less than about 500 amino acids, less than about 400 amino acids, less than about 300 amino acids, or less than about 250 amino acids in length.
  • the peptide fragment can elicit an immune response when used to inoculate an animal.
  • a peptide fragment may be used to elicit an immune response by inoculating an animal with a peptide fragment in combination with an adjuvant, a peptide fragment that is coupled to an adjuvant, or a peptide fragment that is coupled to arsanilic acid, sulfanilic acid, an acetyl group, or a picryl group.
  • a peptide fragment can include a non- amide bond and can be a peptidomimetic.
  • a conjugate encompasses both peptide-small molecule conjugates as well as peptide-protein/peptide conjugates.
  • the term “recombinant,” as used herein, refers to antibodies or antigen-binding fragments thereof of the invention created, expressed, isolated, or obtained by technologies or methods known in the art as recombinant DNA technology, which include, e.g., DNA splicing and transgenic expression.
  • the term refers to antibodies expressed in a non-human mammal (including transgenic non-human mammals, e.g., transgenic mice), or a cell (e.g., CHO cells) expression system or isolated from a recombinant combinatorial human antibody library.
  • a “nucleic acid” or “polynucleotide” refers to a DNA molecule (for example, but not limited to, a cDNA or genomic DNA) or an RNA molecule (for example, but not limited to, an mRNA), and includes DNA or RNA analogs.
  • a DNA or RNA analog can be synthesized from nucleotide analogs.
  • the DNA or RNA molecules may include portions that are not naturally occurring, such as modified bases, modified backbone, deoxyribonucleotides in an RNA, etc.
  • the nucleic acid molecule can be single-stranded or double-stranded.
  • nucleic acid or fragment thereof indicates that, when optimally aligned with appropriate nucleotide insertions or deletions with another nucleic acid (or its complementary strand), there is nucleotide sequence identity in at least about 90%, and more preferably at least about 95%, 96%, 97%, 98% or 99% of the nucleotide bases, as measured by any well-known algorithm of sequence identity, such as FASTA, BLAST or GAP, as discussed below.
  • a nucleic acid molecule having substantial identity to a reference nucleic acid molecule may, in certain instances, encode a polypeptide having the same or substantially similar amino acid sequence as the polypeptide encoded by the reference nucleic acid molecule.
  • the term “substantial similarity” or “substantially similar” means that two peptide sequences, when optimally aligned, such as by the programs GAP or BESTFIT using default gap weights, share at least 90% sequence identity, even more preferably at least 95%, 98% or 99% sequence identity.
  • a “conservative amino acid substitution” is one in which an amino acid residue is substituted by another amino acid residue having a side chain (R group) with similar chemical properties (e.g., charge or hydrophobicity).
  • R group side chain
  • a conservative amino acid substitution will not substantially change the functional properties of a protein.
  • the percent or degree of similarity may be adjusted upwards to correct for the conservative nature of the substitution. Means for making this adjustment are well known to those of skill in the art. See, e.g., Pearson (1994) Methods Mol. Biol. 24: 307-331, which is herein incorporated by reference.
  • Examples of groups of amino acids that have side chains with similar chemical properties include 1) aliphatic side chains: glycine, alanine, valine, leucine, and isoleucine; 2) aliphatic- hydroxyl side chains: serine and threonine; 3) amide-containing side chains: asparagine and glutamine; 4) aromatic side chains: phenylalanine, tyrosine, and tryptophan; 5) basic side chains: lysine, arginine, and histidine; 6) acidic side chains: aspartate and glutamate, and 7) sulfur-containing side chains: cysteine and methionine.
  • Preferred conservative amino acids substitution groups are: valine-leucine-isoleucine, phenylalanine-tyrosine, lysine-arginine, alanine-valine, glutamate-aspartate, and asparagine-glutamine.
  • a conservative replacement is any change having a positive value in the PAM250 log-likelihood matrix disclosed in Gonnet et al. (1992) Science 256: 1443 45, herein incorporated by reference.
  • a “moderately conservative” replacement is any change having a nonnegative value in the PAM250 log- likelihood matrix.
  • GCG software matches similar sequences using measures of similarity assigned to various substitutions, deletions, and other modifications, including conservative amino acid substitutions.
  • GCG software contains programs such as GAP and BESTFIT, which can be used with default parameters to determine sequence homology or sequence identity between closely related polypeptides, such as homologous polypeptides from different species of organisms or between a wild-type protein and a mutein thereof. See, e.g., GCG Version 6.1. Polypeptide sequences also can be compared using FASTA with default or recommended parameters; a program in GCG Version 6.1.
  • FASTA e.g., FASTA2 and FASTA3
  • FASTA2 and FASTA3 provides alignments and percent sequence identity of the regions of the best overlap between the query and search sequences (Pearson (2000) supra).
  • Another preferred algorithm when comparing a sequence of the invention to a database containing a large number of sequences from different organisms is the computer program BLAST, especially BLASTP or TBLASTN, using default parameters. See, e.g., Altschul et al. (1990) J. Mol. Biol. 215: 403-410 and (1997) Nucleic Acids Res. 25:3389- 3402, each of which is herein incorporated by reference.
  • affinity refers to the strength of the sum total of noncovalent interactions between a single binding site of a molecule (e.g., an antibody) and its binding partner (e.g., an antigen).
  • binding affinity refers to intrinsic binding affinity, which reflects a 1:1 interaction between members of a binding pair (e.g., antibody and antigen).
  • the affinity of a molecule X for its partner Y can generally be represented by the dissociation constant (KD). Affinity can be measured by common methods known in the art, including those described herein.
  • the term “specifically binds,” or “binds specifically to,” or the like, refers to an antibody that binds to a single epitope, e.g., under physiologic conditions., but which does not bind to more than one epitope. Accordingly, an antibody that specifically binds to a polypeptide will bind to an epitope that is present on the polypeptide, but which is not present on other polypeptides. Specific binding can be characterized by an equilibrium dissociation constant of at least about l x 10-8 M or less (e.g., a smaller KD denotes a tighter binding).
  • the antibody binds to a spike or S protein with “high affinity,” namely with a KD of 1 X 10-7 M or less, more preferably 5 x 10-8 M or less, more preferably 3 x 10-8 M or less, more preferably 1 x 10-8 M or less, more preferably 5 x 10-9 M or less or even more preferably 1 x 10-9 M or less, as determined by surface plasmon resonance, e.g., BIACORETM.
  • does not substantially bind to a protein or cells, as used herein, means does not bind or does not bind with a high affinity to the protein or cells, i.e., binds to the protein or cells with a KD of 1 x 10-6 M or more, more preferably 1 x 10-5 M or more, more preferably 1 x 10-4 M or more, more preferably 1 x 10-3 M or more, even more preferably 1 x 10-2 M or more.
  • Kassoc or “Ka,” as used herein, is intended to refer to the association rate of a particular antibody-antigen interaction
  • Kdis or “Kd,” as used herein, is intended to refer to the dissociation rate of a particular antibody-antigen interaction
  • KD is intended to refer to the dissociation constant, which is obtained from the ratio of Kd to Ka (i.e., Kd/Ka) and is expressed as a molar concentration (M). KD values for antibodies can be determined using methods well established in the art.
  • a preferred method for determining the KD of an antibody is by using surface plasmon resonance, preferably using a biosensor system such as a BIACORETM system.
  • Antibodies that “compete with another antibody for binding to a target” refer to antibodies that inhibit (partially or completely) the binding of the other antibody to the target. Whether two antibodies compete with each other for binding to a target, i.e., whether and to what extent one antibody inhibits the binding of the other antibody to a target, may be determined using known competition experiments. In some embodiments, an antibody competes with, and inhibits binding of another antibody to a target by at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% or 100%.
  • the level of inhibition or competition may be different depending on which antibody is the “blocking antibody” (i.e., the cold antibody that is incubated first with the target).
  • Competition assays can be conducted as described, for example, in Ed Harlow and David Lane, Cold Spring Harb Protoc; 2006 or in Chapter 11 of “Using Antibodies” by Ed Harlow and David Lane, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, USA 1999. Competing antibodies bind to the same epitope, an overlapping epitope or to adjacent epitopes (e.g., as evidenced by steric hindrance).
  • epitope refers to an antigenic determinant that interacts with a specific antigen-binding site in the variable region of an antibody molecule known as a paratope.
  • a single antigen may have more than one epitope.
  • different antibodies may bind to different areas on an antigen and may have different biological effects.
  • epitopes also refers to a site on an antigen to which B and/or T cells respond. It also refers to a region of an antigen that is bound by an antibody. Epitopes may be defined as structural or functional. Functional epitopes are generally a subset of the structural epitopes and have those residues that directly contribute to the affinity of the interaction. Epitopes may also be conformational, that is, composed of non- linear amino acids. In some embodiments, epitopes may include determinants that are chemically active surface groupings of molecules such as amino acids, sugar side chains, phosphoryl groups, or sulfonyl groups, and, In some embodiments, may have specific three-dimensional structural characteristics, and/or specific charge characteristics.
  • An epitope typically includes at least 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15 amino acids in a unique spatial conformation.
  • Methods for determining what epitopes are bound by a given antibody i.e., epitope mapping
  • epitope mapping include, for example, immunoblotting and immune-precipitation assays, wherein overlapping or contiguous peptides from a spike or S protein are tested for reactivity with a given antibody.
  • Methods of determining spatial conformation of epitopes include techniques in the art and those described herein, for example, x-ray crystallography and 2-dimensional nuclear magnetic resonance (see, e.g., Epitope Mapping Protocols in Methods in Molecular Biology, Vol. 66, G. E.
  • epitope mapping refers to the process of identification of the molecular determinants for antibody-antigen recognition.
  • the term “binds to an epitope” or “recognizes an epitope” with reference to an antibody or antibody fragment refers to continuous or discontinuous segments of amino acids within an antigen. Those of skill in the art understand that the terms do not necessarily mean that the antibody or antibody fragment is in direct contact with every amino acid within an epitope sequence.
  • the term “binds to the same epitope” with reference to two or more antibodies means that the antibodies bind to the same, overlapping, or encompassing continuous or discontinuous segments of amino acids.
  • the phrase “binds to the same epitope” does not necessarily mean that the antibodies bind to or contact exactly the same amino acids.
  • the precise amino acids that the antibodies contact can differ.
  • a first antibody can bind to a segment of amino acids that is completely encompassed by the segment of amino acids bound by a second antibody.
  • a first antibody binds one or more segments of amino acids that significantly overlap the one or more segments bound by the second antibody.
  • such antibodies are considered to “bind to the same epitope.”
  • the term “immune response” refers to a biological response within a vertebrate against foreign agents, which response protects the organism against these agents and diseases caused by them.
  • An immune response is mediated by the action of a cell of the immune system (for example, a T lymphocyte, B lymphocyte, natural killer (NK) cell, macrophage, eosinophil, mast cell, dendritic cell, or neutrophil) and soluble macromolecules produced by any of these cells or the liver (including antibodies, cytokines, and complement) that results in selective targeting, binding to, damage to, destruction of, and/or elimination from the vertebrate's body of invading pathogens, cells or tissues infected with pathogens, cancerous or other abnormal cells, or, in cases of autoimmunity or pathological inflammation, normal human cells or tissues.
  • a cell of the immune system for example, a T lymphocyte, B lymphocyte, natural killer (NK) cell, macrophage, eosinophil, mast cell, dendritic cell, or neutrophil
  • soluble macromolecules produced by any of these cells or the liver (including antibodies, cytokines, and complement) that results in selective targeting, binding to, damage
  • An immune reaction includes, e.g., activation or inhibition of a T cell, e.g., an effector T cell or a Th cell, such as a CD4+ or CD8+ T cell, or the inhibition of a Treg cell.
  • a T cell e.g., an effector T cell or a Th cell, such as a CD4+ or CD8+ T cell, or the inhibition of a Treg cell.
  • the term “detectable label” as used herein refers to a molecule capable of detection, including, but not limited to, radioactive isotopes, fluorescers, chemiluminescers, chromophores, enzymes, enzyme substrates, enzyme cofactors, enzyme inhibitors, chromophores, dyes, metal ions, metal sols, ligands (e.g., biotin, avidin, streptavidin or haptens), intercalating dyes and the like.
  • fluorescer refers to a substance or a portion thereof that is capable of exhibiting fluorescence in the detectable range.
  • the terms “subject” and “patient” are used interchangeably irrespective of whether the subject has or is currently undergoing any form of treatment.
  • the terms “subject” and “subjects” may refer to any vertebrate, including, but not limited to, a mammal (e.g., cow, pig, camel, llama, horse, goat, rabbit, sheep, hamsters, guinea pig, cat, dog, rat, and mouse, a non-human primate (for example, a monkey, such as a cynomolgus monkey, chimpanzee, etc.) and a human).
  • the subject may be a human or a non-human.
  • the mammal is a human.
  • a subject in need thereof or “a patient in need thereof” means a human or non-human mammal that exhibits one or more symptoms or indications of disorders (e.g., neuronal disorders, autoimmune diseases, and cardiovascular diseases), and/or who has been diagnosed with inflammatory disorders.
  • the subject is a mammal.
  • the subject is human.
  • Progeny of such a cell may not be identical to the parent cell comprising the nucleic acid molecule due to mutations or environmental influences that may occur in succeeding generations or integration of the nucleic acid molecule into the host cell genome.
  • disease is intended to be generally synonymous and is used interchangeably with the terms “disorder” and “condition” (as in medical condition), in that all reflect an abnormal condition (e.g., inflammatory disorder) of the human or animal body or of one of its parts that impairs normal functioning, is typically manifested by distinguishing signs and symptoms, and causes the human or animal to have a reduced duration or quality of life.
  • the term “treating” or “treatment” of any disease or disorder refers in one embodiment, to ameliorating the disease or disorder (i.e., arresting or reducing the development of the disease or at least one of the clinical symptoms thereof). In another embodiment, “treating” or “treatment” refers to ameliorating at least one physical parameter, which may not be discernible by the patient. In yet another embodiment, “treating” or “treatment” refers to modulating the disease or disorder, either physically, (e.g., stabilization of a discernible symptom), physiologically, (e.g., stabilization of a physical parameter), or both. In yet another embodiment, “treating” or “treatment” refers to preventing or delaying the onset or development or progression of the disease or disorder.
  • the terms “prevent,” “preventing,” “prevention,” “prophylactic treatment” and the like refer to reducing the probability of developing a disorder or condition in a subject, who does not have, but is at risk of or susceptible to developing a disorder or condition.
  • the terms “decrease,” “reduced,” “reduction,” “decrease,” or “inhibit” are all used herein generally to mean a decrease by a statistically significant amount.
  • “reduced,” “reduction,” “decrease,” or “inhibit” means a decrease by at least 10% as compared to a reference level, for example, a decrease by at least about 20%, or at least about 30%, or at least about 40%, or at least about 50%, or at least about 60%, or at least about 70%, or at least about 80%, or at least about 90% or up to and including a 100% decrease (e.g., absent level as compared to a reference sample), or any decrease between 10-100% as compared to a reference level.
  • the term “agent” denotes a chemical compound, a mixture of chemical compounds, a biological macromolecule (such as a nucleic acid, an antibody, a protein or portion thereof, e.g., a peptide), or an extract made from biological materials such as bacteria, plants, fungi, or animal (particularly mammalian) cells or tissues.
  • a biological macromolecule such as a nucleic acid, an antibody, a protein or portion thereof, e.g., a peptide
  • an extract made from biological materials such as bacteria, plants, fungi, or animal (particularly mammalian) cells or tissues.
  • the activity of such agents may render it suitable as a “therapeutic agent,” which is a biologically, physiologically, or pharmacologically active substance (or substances) that acts locally or systemically in a subject.
  • the terms “therapeutic agent,” “therapeutic capable agent,” or “treatment agent” are used interchangeably and refer to a molecule or compound that confers some beneficial effect upon administration to a subject.
  • the beneficial effect includes enablement of diagnostic determinations; amelioration of a disease, symptom, disorder, or pathological condition; reducing or preventing the onset of a disease, symptom, disorder, or condition; and generally counteracting a disease, symptom, disorder or pathological condition.
  • the term “therapeutic effect” is art-recognized and refers to a local or systemic effect in animals, particularly mammals, and more particularly humans caused by a pharmacologically active substance.
  • an effective amount is defined as an amount sufficient to achieve or at least partially achieve a desired effect.
  • a “therapeutically effective amount” or “therapeutically effective dosage” of a drug or therapeutic agent is any amount of the drug that, when used alone or in combination with another therapeutic agent, promotes disease regression evidenced by a decrease in severity of disease symptoms, an increase in frequency and duration of disease symptom-free periods, or a prevention of impairment or disability due to the disease affliction.
  • a “prophylactically effective amount” or a “prophylactically effective dosage” of a drug is an amount of the drug that, when administered alone or in combination with another therapeutic agent to a subject at risk of developing a disease or of suffering a recurrence of disease, inhibits the development or recurrence of the disease.
  • the ability of a therapeutic or prophylactic agent to promote disease regression or inhibit the development or recurrence of the disease can be evaluated using a variety of methods known to the skilled practitioner, such as in human subjects during clinical trials, in animal model systems predictive of efficacy in humans, or by assaying the activity of the agent in in vitro assays. [00116] Doses are often expressed in relation to bodyweight.
  • composition or “pharmaceutical composition” refers to a mixture of at least one component useful within the invention with other components, such as carriers, stabilizers, diluents, dispersing agents, suspending agents, thickening agents, and/or excipients.
  • the pharmaceutical composition facilitates administration of one or more components of the invention to an organism.
  • the term “pharmaceutically acceptable” refers to a material, such as a carrier or diluent, which does not abrogate the biological activity or properties of the composition, and is relatively non-toxic, i.e., the material may be administered to an individual without causing undesirable biological effects or interacting in a deleterious manner with any of the components of the composition in which it is contained.
  • the term “pharmaceutically acceptable carrier” includes a pharmaceutically acceptable salt, pharmaceutically acceptable material, composition or carrier, such as a liquid or solid filler, diluent, excipient, solvent or encapsulating material, involved in carrying or transporting a compound(s) of the present invention within or to the subject such that it may perform its intended function. Typically, such compounds are carried or transported from one organ, or portion of the body, to another organ, or portion of the body. Each salt or carrier must be “acceptable” in the sense of being compatible with the other ingredients of the formulation and not injurious to the subject.
  • materials that may serve as pharmaceutically acceptable carriers include: sugars, such as lactose, glucose, and sucrose; starches, such as corn starch and potato starch; cellulose, and its derivatives, such as sodium carboxymethyl cellulose, ethyl cellulose, and cellulose acetate; powdered tragacanth; malt; gelatin; talc; excipients, such as cocoa butter and suppository waxes; oils, such as peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil, and soybean oil; glycols, such as propylene glycol; polyols, such as glycerin, sorbitol, mannitol, and polyethylene glycol; esters, such as ethyl oleate and ethyl laurate; agar; buffering agents, such as magnesium hydroxide and aluminum hydroxide; alginic acid; pyrogen- free water; isotonic saline
  • “pharmaceutically acceptable carrier” also includes any and all coatings, antibacterial and antifungal agents, and absorption delaying agents, and the like that are compatible with the activity of one or more components of this disclosure, and are physiologically acceptable to the subject. Supplementary active compounds may also be incorporated into the compositions.
  • “Combination” therapy is meant to encompass administration of two or more therapeutic agents in a coordinated fashion and includes, but is not limited to, concurrent dosing.
  • combination therapy encompasses both co-administration (e.g., administration of a co-formulation or simultaneous administration of separate therapeutic compositions) and serial or sequential administration, provided that administration of one therapeutic agent is conditioned in some way on the administration of another therapeutic agent.
  • one therapeutic agent may be administered only after a different therapeutic agent has been administered and allowed to act for a prescribed period of time. See, e.g., Kohrt et al. (2011) Blood 117:2423.
  • co-administration or “co-administered” refers to the administration of at least two agent(s) or therapies to a subject. In some embodiments, the co- administration of two or more agents/therapies is concurrent.
  • a first agent/therapy is administered prior to a second agent/therapy.
  • a second agent/therapy is administered prior to a second agent/therapy.
  • the term “contacting,” when used in reference to any set of components includes any process whereby the components to be contacted are mixed into the same mixture (for example, are added into the same compartment or solution), and does not necessarily require actual physical contact between the recited components.
  • the recited components can be contacted in any order or any combination (or sub-combination) and can include situations where one or some of the recited components are subsequently removed from the mixture, optionally prior to addition of other recited components.
  • “contacting A with B and C” includes any and all of the following situations: (i) A is mixed with C, then B is added to the mixture; (ii) A and B are mixed into a mixture; B is removed from the mixture, and then C is added to the mixture; and (iii) A is added to a mixture of B and C.
  • sample can be a sample of serum, urine plasma, amniotic fluid, cerebrospinal fluid, cells, or tissue.
  • sample can be used directly as obtained from a patient or can be pre-treated, such as by filtration, distillation, extraction, concentration, centrifugation, inactivation of interfering components, addition of reagents, and the like, to modify the character of the sample in some manner as discussed herein or otherwise as is known in the art.
  • sample and biological sample as used herein generally refer to a biological material being tested for and/or suspected of containing an analyte of interest, such as antibodies.
  • the sample may be any tissue sample from the subject.
  • the sample may comprise protein from the subject.
  • the term “in vitro” refers to events that occur in an artificial environment, e.g., in a test tube or reaction vessel, in cell culture, etc., rather than within a multi-cellular organism.
  • the term “in vivo” refers to events that occur within a multi-cellular organism, such as a non-human animal.
  • the singular forms “a,” “an,” and “the” include plural references unless the context clearly dictates otherwise.
  • the terms “including,” “comprising,” “containing,” or “having” and variations thereof are meant to encompass the items listed thereafter and equivalents thereof as well as additional subject matter unless otherwise noted.
  • the phrases “in one embodiment,” “in various embodiments,” “in some embodiments,” and the like are used repeatedly. Such phrases do not necessarily refer to the same embodiment, but they may unless the context dictates otherwise.
  • the terms “and/or” or “/” means any one of the items, any combination of the items, or all of the items with which this term is associated.
  • the word “substantially” does not exclude “completely,” e.g., a composition that is “substantially free” from Y may be completely free from Y. Where necessary, the word “substantially” may be omitted from the definition of the invention.
  • each when used in reference to a collection of items, is intended to identify an individual item in the collection but does not necessarily refer to every item in the collection. Exceptions can occur if explicit disclosure or context clearly dictates otherwise.
  • the term “approximately” or “about” refers to a range of values that fall within 25%, 20%, 19%, 18%, 17%, 16%, 15%, 14%, 13%, 12%, 11%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, or less in either direction (greater than or less than) of the stated reference value unless otherwise stated or otherwise evident from the context (except where such number would exceed 100% of a possible value).
  • the term “about” is intended to include values, e.g., weight percents, proximate to the recited range that are equivalent in terms of the functionality of the individual ingredient, the composition, or the embodiment.
  • this disclosure provides novel isolated and recombinantly modified anti- SARS-CoV-2 antibodies or antigen-binding fragments thereof.
  • antibodies or antigen-binding fragments thereof described herein are broadly neutralizing antibodies that neutralize multiple SARS-CoV-2 virus strains. The antibodies are able to protect a subject prophylactically and therapeutically against a lethal challenge with a SARS-CoV-2 virus.
  • this disclosure provides an isolated anti-SARS-CoV-2 antibody or antigen-binding fragment thereof that binds specifically to a SARS-CoV-2 antigen.
  • the SARS-CoV-2 antigen comprises a portion of a spike (S) polypeptide, such as a S polypeptide of a human or an animal SARS-CoV-2.
  • the antibodies described herein are monoclonal antibodies.
  • the monoclonal antibodies, or antigen-binding fragments thereof specifically bind to the S polypeptide of a SARS-CoV-2, or a domain of the spike protein.
  • the monoclonal antibodies or antigen-binding fragments thereof bind to the receptor binding domain (RBD) of the S polypeptide (also referred to herein as the Spike protein). In some embodiments, the monoclonal antibodies or antigen-binding fragments thereof bind to the N- terminal domain (NTD) of the Spike protein. In some embodiments, the monoclonal antibodies or antigen-binding fragments thereof bind to the S2 domain of the Spike protein. [00141] In some embodiments, the monoclonal antibodies or antigen-binding fragments thereof bind to the RBD of the Spike protein (e.g., amino acids 319-541 of SEQ ID NO: 1353).
  • the monoclonal antibodies or antigen-binding fragments thereof bind to the N- terminal domain (NTD) (e.g., residues 14–305 of SEQ ID NO: 1353) of the S polypeptide. In some embodiments, the monoclonal antibodies or antigen-binding fragments thereof bind to the supersite b-hairpin (residues 152-158 of SEQ ID NO: 1353).
  • NTD N- terminal domain
  • the monoclonal antibodies or antigen-binding fragments thereof bind to the supersite b-hairpin (residues 152-158 of SEQ ID NO: 1353).
  • the monoclonal antibodies or antigen-binding fragments thereof bind to WKH ⁇ -strand (residue 97-102 of SEQ ID NO: 1353), N4-loop (residues 178-188 of SEQ ID NO: 1353), and/or N-linked glycans at positions N122 and N149 of the S polypeptide. In some embodiments, the monoclonal antibodies or antigen-binding fragments thereof bind to residues 27-32, 57-60, 210-218, and/or 286-303 of the S polypeptide of SEQ ID NO: 1353.
  • the monoclonal antibodies or antigen-binding fragments thereof bind to residues 600-606 of the S polypeptide of SEQ ID NO: 1353.
  • the antibody or antigen-binding fragment thereof is capable of neutralizing a SARS-CoV-2 virus at an IC50 concentration of less than 50 (e.g., 1, 5, 10, 20, 30, 40, 50) ⁇ g/ml.
  • the monoclonal antibodies or antigen-binding fragment thereof disclosed herein can be used for detecting and treating SARS-CoV-2. For instance, they can be useful in a cocktail approach designed to minimize the chances of escape that can readily occur with single antibody approaches.
  • the spike protein is important because it is present on the outside of intact SARS-CoV- 2. Thus, it presents a target that can be used to inhibit or eliminate an intact virus before the virus has an opportunity to infect a cell.
  • a representative amino acid sequence is provided below: [00145]
  • the total length of SARS-CoV-2 S is 1273 amino acids and consists of a signal peptide (amino acids 1–13) located at the N-terminus, the S1 subunit (14–685 residues), and the S2 subunit (686–1273 residues); the last two regions are responsible for receptor binding and membrane fusion, respectively.
  • S1 subunit there is an N-terminal domain (14–305 residues) and a receptor-binding domain (RBD, 319–541 residues); the fusion peptide (FP) (788–806 residues), heptapeptide repeat sequence 1 (HR1) (912–984 residues), HR2 (1163–1213 residues), TM domain (1213–1237 residues), and cytoplasm domain (1237–1273 residues) comprise the S2 subunit.
  • S protein trimers visually form a characteristic bulbous, crown-like halo surrounding the viral particle. Based on the structure of coronavirus S protein monomers, the S1 and S2 subunits form the bulbous head and stalk region.
  • HC variable regions HCVR
  • LCVR light chain variable regions
  • the antibody or antigen-binding fragment thereof comprises: (i) a heavy chain variable region having an amino acid sequence with at least 75% (e.g., 75%, 78%, 80%, 82%, 84%, 85%, 86%, 88%, 90%, 92%, 94%, 95%, 96%, 97%, 98%, 99%) identity to one selected from those in Table 2; and/or (ii) a light chain variable region having an amino acid sequence with at least 75% (e.g., 75%, 78%, 80%, 82%, 84%, 85%, 86%, 88%, 90%, 92%, 94%, 95%, 96%, 97%, 98%, 99%) identity to one selected from those in Table 2.
  • a heavy chain variable region having an amino acid sequence with at least 75% e.g., 75%, 78%, 80%, 82%, 84%, 85%, 86%, 88%, 90%, 92%, 94%, 95%, 96%, 97%, 98%, 99%
  • the antibody or antigen-binding fragment thereof comprises: (i) a heavy chain variable region having an amino acid sequence selected from those in Table 2; and/or (ii) a light chain variable region having an amino acid sequence selected from those in Table 2. [00150] In some embodiments, the antibody or antigen-binding fragment thereof comprises a heavy chain variable region and a light chain variable region that comprises an amino acid sequence pair selected from those in Table 2.
  • the antibody or antigen-binding fragment thereof comprises three heavy chain complementarity determining regions (HCDRs) (HCDR1, HCDR2, and HCDR3) of a heavy chain variable region having the amino acid sequence of SEQ ID NO: 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31, 33, 35, 37, 39, 41, 43, 45, 47, 49, 51, 53, 55, 57, 59, 61, 63, 65, 67, 69, 71, 73, 75, 77, 79, 81, 83, 85, 87, 89, 91, 93, 95, 97, 99, 101, 103, 105, 107, 109, 111, 113, 115, 117, 119, 121, 123, 125, 127, 129, 131, 133, 135, 137, 139, 141, 143, 145, 147, 149, 151, 153, 155, 157, 159, 16
  • HCDRs heavy chain
  • the antibody or antigen-binding fragment thereof comprises: a heavy chain variable region comprising an amino acid sequence with at least 75% (e.g., 75%, 78%, 80%, 82%, 84%, 85%, 86%, 88%, 90%, 92%, 94%, 95%, 96%, 97%, 98%, 99%) identity to the amino acid sequence of SEQ ID NO: 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31, 33, 35, 37, 39, 41, 43, 45, 47, 49, 51, 53, 55, 57, 59, 61, 63, 65, 67, 69, 71, 73, 75, 77, 79, 81, 83, 85, 87, 89, 91,
  • the antibody or antigen-binding fragment thereof comprises a heavy chain variable region (HCVR) and a light chain variable region (LCVR) that comprise a HCVR/LCVR amino acid sequence pair of SEQ ID NOs: 1-2, 3-4, 5-6, 7-8, 9-10, 11-12, 13-14, 15-16, 17-18, 19-20, 21-22, 23-24, 25-26, 27-28, 29-30, 31-32, 33-34, 35-36, 37-38, 39-40, 41-42, 43-44, 45-46, 47-48, 49-50, 51-52, 53-54, 55-56, 57-58, 59-60, 61-62, 63-64, 65-66, 67-68, 69-70, 71-72, 73-74, 75-76, 77-78, 79-80, 81-82, 83-84, 85-86, 87-88, 89-90, 91-92, 93-94, 95-96,
  • the antibody is selected from B1030, B1032, B2014, B2058, and B2117 antibodies, as listed in Table 2. Sequences for exemplary antibodies are provided in Table 3 below: Table 3: Exemplary Antibody Sequences [00156]
  • the antibody or antigen-binding fragment thereof comprises a variable heavy chain CDR1 (HCDR1) comprising an amino acid sequence of SEQ ID NO: 1354.
  • the antibody or antigen-binding fragment thereof comprises a HCDR1 comprising an amino acid sequence of SEQ ID NO:1360.
  • the antibody or antigen-binding fragment thereof comprises a HCDR1 comprising an amino acid sequence of SEQ ID NO:1366.
  • the antibody or antigen-binding fragment thereof comprises a HCDR1 comprising an amino acid sequence of SEQ ID NO:1372. In some embodiments, the antibody or antigen-binding fragment thereof comprises a HCDR1 comprising an amino acid sequence of SEQ ID NO:1378. In some embodiments, the HCDR1 comprises an amino acid sequence that is at least 75% or at least 87% identical to one of SEQ ID NOs: 1354, 1360, 1366, 1372, or 1378. In some embodiments, the HCDR1 comprises an amino acid sequence that is 100% identical to one of SEQ ID NOs: 1354, 1360, 1366, 1372, or 1378 except for the substitution or deletion of one or two amino acids.
  • the antibody or antigen-binding fragment thereof comprises a variable heavy chain CDR2 HCDR2 comprising an amino acid sequence of SEQ ID NO: 1355. In some embodiments, the antibody or antigen-binding fragment thereof comprises a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1361. In some embodiments, the antibody or antigen-binding fragment thereof comprises a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1367. In some embodiments, the antibody or antigen-binding fragment thereof comprises a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1373.
  • the antibody or antigen-binding fragment thereof comprises a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1379. In some embodiments, the antibody or antigen-binding fragment thereof comprises a HCDR2 comprising an amino acid sequence that is at least 71%, at least 75%, at least 85%, or at least 87% identical to one of SEQ ID NOs: 1355, 1361, 1367, 1373, or 1379. In some embodiments, the HCDR2 comprises an amino acid sequence that is at least 100% identical to one of SEQ ID NOs: 1355, 1361, 1367, 1373, or 1379 except for the substitution or deletion of one or two amino acids.
  • the antibody or antigen-binding fragment thereof comprises a variable heavy chain CDR3 (HCDR3) comprising an amino acid sequence of SEQ ID NO: 1356. In some embodiments, the antibody or antigen-binding fragment thereof comprises a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1362. In some embodiments, the antibody or antigen-binding fragment thereof comprises a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1368. In some embodiments, the antibody or antigen-binding fragment thereof comprises a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1374.
  • HCDR3 variable heavy chain CDR3
  • the antibody or antigen-binding fragment thereof comprises a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1380. In some embodiments, the antibody or antigen-binding fragment thereof comprises a HCDR3 comprising an amino acid sequence that is at least at least 75%, at least 81%, at least 85%, at least 86%, at least 87%, at least 89%, at least 93%, at least 94%, or at least 95% identical to one of SEQ ID NOs: 1356, 1362, 1368, 1374, or 1380.
  • the HCDR3 comprises an amino acid sequence that is at least 100% identical to one of SEQ ID NOs: 1356, 1362, 1368, 1374, or 1380 except for the substitution or deletion of one or two amino acids.
  • the antibody or antigen-binding fragment thereof comprises a variable light chain CDR1 (LCDR1) comprising an amino acid sequence of SEQ ID NO: 1357.
  • the antibody or antigen-binding fragment thereof comprises a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1363.
  • the antibody or antigen-binding fragment thereof comprises a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1369.
  • the antibody or antigen-binding fragment thereof comprises a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1375. In some embodiments, the antibody or antigen-binding fragment thereof comprises a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1381. In some embodiments, the antibody or antigen-binding fragment thereof comprises an LCDR1 comprises an amino acid sequence that is at least 77% or at least 88% identical to one of SEQ ID NOs: 1357, 1363, 1369, 1375, or 1381. In some embodiments, the LCDR1 comprises an amino acid sequence that is at least 100% identical to one of SEQ ID NOs: 1357, 1363, 1369, 1375, or 1381 except for the substitution or deletion of one or two amino acids.
  • the antibody or antigen-binding fragment thereof comprises a variable light chain CDR2 (LCDR2) comprising an amino acid sequence of SEQ ID NO: 1358.
  • the antibody or antigen-binding fragment thereof comprises a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1364.
  • the antibody or antigen-binding fragment thereof comprises a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1370.
  • the antibody or antigen-binding fragment thereof comprises a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1379.
  • the antibody or antigen-binding fragment thereof comprises a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1382.
  • the antibody or antigen-binding fragment thereof comprises an LCDR2 comprises an amino acid sequence that is at least 66% identical to one of SEQ ID NOs: 1358, 1364, 1370, 1376, or 1382. In some embodiments, the LCDR2 comprises an amino acid sequence that is at least 100% identical to one of SEQ ID NOs: 1358, 1364, 1370, 1376, or 1382 except for the substitution or deletion of one amino acid. [00161] In some embodiments, the antibody or antigen-binding fragment thereof comprises a variable light chain CDR3 (LCDR3) comprising an amino acid sequence of SEQ ID NO: 1359.
  • LCDR3 variable light chain CDR3
  • the antibody or antigen-binding fragment thereof comprises a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1365. In some embodiments, the antibody or antigen-binding fragment thereof comprises a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1371. In some embodiments, the antibody or antigen-binding fragment thereof comprises a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1377. In some embodiments, the antibody or antigen-binding fragment thereof comprises a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1383.
  • the antibody or antigen-binding fragment thereof comprises an LCDR3 comprises an amino acid sequence that is at least 50%, at least 75%, at least 77%, at least 81%, at least 84%, at least 87%, at least 88%, at least 90%, or at least 92% identical to one of SEQ ID NOs: 1359, 1365, 1371, 1377, or 1383.
  • the LCDR3 comprises an amino acid sequence that is at least 100% identical to one of SEQ ID NOs: 1359, 1365, 1371, 1377, or 1383 except for the substitution or deletion of one or two amino acids.
  • the antibody or antigen-binding fragment thereof comprises: 1) a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1354, 1360, 1366, 1372, or 1378; 2) a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1355, 1361, 1367, 1373, or 1379; 3) a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1356, 1362, 1368, 1374, or 1380; 4) a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1357, 1363, 1369, 1375, or 1381; 5) a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1358, 1364, 1370, 1376, or 1382; and 6) a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1359, 1365, 1371, 1377, or 1383.
  • the antibody or antigen-binding fragment thereof comprises a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1354, a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1355, a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1356, a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1357, a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1358, and a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1359.
  • the antibody or antigen-binding fragment thereof comprises a HCDR1 consisting of SEQ ID NO: 1354, a HCDR2 consisting of SEQ ID NO: 1355, a HCDR3 consisting of SEQ ID NO: 1356, a LCDR1 consisting of SEQ ID NO: 1357, a LCDR2 consisting of SEQ ID NO: 1358, and a LCDR3 consisting of SEQ ID NO: 1359.
  • the antibody or antigen-binding fragment thereof comprises a VH comprising an amino acid sequence that is at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% identical to SEQ ID NO: 25.
  • the antibody or antigen-binding fragment thereof comprises a VH comprising an amino acid sequence of SEQ ID NO: 25.
  • the antibody or antigen-binding fragment thereof comprises a VH consisting of SEQ ID NO: 25.
  • the antibody or antigen-binding fragment thereof comprises a VL comprising an amino acid sequence that is at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% identical to SEQ ID NO: 26.
  • the antibody or antigen-binding fragment thereof comprises a VL comprising an amino acid sequence of SEQ ID NO: 26.
  • the antibody or antigen-binding fragment thereof comprises a VH consisting of SEQ ID NO: 26.
  • the antibody or antigen-binding fragment thereof comprises a VH comprising an amino acid sequence that is at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% identical to SEQ ID NO: 25 and a VL comprising an amino acid sequence that is at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% identical to SEQ ID NO: 26.
  • the antibody or antigen-binding fragment thereof comprises a VH comprising an amino acid sequence of SEQ ID NO: 25 and a VL comprising an amino acid sequence of SEQ ID NO: 26. In some embodiments, the antibody or antigen-binding fragment thereof comprises a VL consisting of SEQ ID NO: 25 and a VL consisting of SEQ ID NO: 26.
  • the antibody or antigen-binding fragment thereof comprises a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1360, a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1361, a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1362, a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1363, a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1364, and a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1365.
  • the antibody or antigen-binding fragment thereof comprises a HCDR1 consisting of SEQ ID NO: 1360, a HCDR2 consisting of SEQ ID NO: 1361, a HCDR3 consisting of SEQ ID NO: 1362, a LCDR1 consisting of SEQ ID NO: 1363, a LCDR2 consisting of SEQ ID NO: 1364, and a LCDR3 consisting of SEQ ID NO: 1365.
  • the antibody or antigen-binding fragment thereof comprises a VH comprising an amino acid sequence that is at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% identical to SEQ ID NO: 395.
  • the antibody or antigen-binding fragment thereof comprises a VH comprising an amino acid sequence of SEQ ID NO: 395.
  • the antibody or antigen-binding fragment thereof comprises a VH consisting of SEQ ID NO: 395.
  • the antibody or antigen-binding fragment thereof comprises a VL comprising an amino acid sequence that is at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% identical to SEQ ID NO: 396.
  • the antibody or antigen-binding fragment thereof comprises a VL comprising an amino acid sequence of SEQ ID NO: 396.
  • the antibody or antigen- binding fragment thereof comprises a VL consisting of SEQ ID NO: 396.
  • the antibody or antigen-binding fragment thereof comprises a VH comprising an amino acid sequence that is at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% identical to SEQ ID NO: 395 and a VL comprising an amino acid sequence that is at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% identical to SEQ ID NO: 396.
  • the antibody or antigen-binding fragment thereof comprises a VH comprising an amino acid sequence of SEQ ID NO: 395 and a VL comprising an amino acid sequence of SEQ ID NO: 396. In some embodiments, the antibody or antigen-binding fragment thereof comprises a VH consisting of SEQ ID NO: 395 and a VL consisting of SEQ ID NO: 396.
  • the antibody or antigen-binding fragment thereof comprises a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1366, a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1367, a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1368, a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1369, a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1370, and a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1371.
  • the antibody or antigen-binding fragment thereof comprises a HCDR1 consisting of SEQ ID NO: 1366, a HCDR2 consisting of SEQ ID NO: 1367, a HCDR3 consisting of SEQ ID NO: 1368, a LCDR1 consisting of SEQ ID NO: 1369, a LCDR2 consisting of SEQ ID NO: 1370, and a LCDR3 consisting of SEQ ID NO: 1371.
  • the antibody or antigen-binding fragment thereof comprises a VH comprising an amino acid sequence that is at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% identical to SEQ ID NO: 397.
  • the antibody or antigen-binding fragment thereof comprises a VH comprising an amino acid sequence of SEQ ID NO: 397.
  • the antibody or antigen-binding fragment thereof comprises a VH consisting of SEQ ID NO: 397.
  • the antibody or antigen-binding fragment thereof comprises a VL comprising an amino acid sequence that is at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% identical to SEQ ID NO: 398.
  • the antibody or antigen-binding fragment thereof comprises a VL comprising an amino acid sequence of SEQ ID NO: 398.
  • the antibody or antigen- binding fragment thereof comprises a VL consisting of SEQ ID NO: 398.
  • the antibody or antigen-binding fragment thereof comprises a VH comprising an amino acid sequence that is at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% identical to SEQ ID NO: 397 and a VL comprising an amino acid sequence that is at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% identical to SEQ ID NO: 398.
  • the antibody or antigen-binding fragment thereof comprises a VH comprising an amino acid sequence of SEQ ID NO: 397 and a VL comprising an amino acid sequence of SEQ ID NO: 398. In some embodiments, the antibody or antigen-binding fragment thereof comprises a VH consisting of SEQ ID NO: 397 and a VL consisting of SEQ ID NO: 398.
  • the antibody or antigen-binding fragment thereof comprises a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1372, a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1373, a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1374, a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1375, a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1376, and a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1377.
  • the antibody or antigen-binding fragment thereof comprises a HCDR1 consisting of SEQ ID NO: 1372, a HCDR2 consisting of SEQ ID NO: 1373, a HCDR3 consisting of SEQ ID NO: 1374, a LCDR1 consisting of SEQ ID NO: 1375, a LCDR2 consisting of SEQ ID NO: 1376, and a LCDR3 consisting of SEQ ID NO: 1377.
  • the antibody or antigen-binding fragment thereof comprises a VH comprising an amino acid sequence that is at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% identical to SEQ ID NO: 191.
  • the antibody or antigen-binding fragment thereof comprises a VH comprising an amino acid sequence of SEQ ID NO: 191.
  • the antibody or antigen-binding fragment thereof comprises a VH consisting of SEQ ID NO: 191.
  • the antibody or antigen-binding fragment thereof comprises a VL comprising an amino acid sequence that is at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% identical to SEQ ID NO: 192.
  • the antibody or antigen-binding fragment thereof comprises a VL comprising an amino acid sequence of SEQ ID NO: 192.
  • the antibody or antigen- binding fragment thereof comprises a VL consisting of SEQ ID NO: 192.
  • the antibody or antigen-binding fragment thereof comprises a VH comprising an amino acid sequence that is at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% identical to SEQ ID NO: 191 and a VL comprising an amino acid sequence that is at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% identical to SEQ ID NO: 192.
  • the antibody or antigen-binding fragment thereof comprises a VH comprising an amino acid sequence of SEQ ID NO: 191 and a VL comprising an amino acid sequence of SEQ ID NO: 192. In some embodiments, the antibody or antigen-binding fragment thereof comprises a VH consisting of SEQ ID NO: 191 and a VL consisting of SEQ ID NO: 192.
  • the antibody or antigen-binding fragment thereof comprises a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1378, a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1379, a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1380, a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1381, a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1382, and a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1383.
  • the antibody or antigen-binding fragment thereof comprises a HCDR1 consisting of SEQ ID NO: 1378, a HCDR2 consisting of SEQ ID NO: 1379, a HCDR3 consisting of SEQ ID NO: 1380, a LCDR1 consisting of SEQ ID NO: 1381, a LCDR2 consisting of SEQ ID NO: 1382, and a LCDR3 consisting of SEQ ID NO: 1383.
  • the antibody or antigen-binding fragment thereof comprises a VH comprising an amino acid sequence that is at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% identical to SEQ ID NO: 219.
  • the antibody or antigen-binding fragment thereof comprises a VH comprising an amino acid sequence of SEQ ID NO: 219.
  • the antibody or antigen-binding fragment thereof comprises a VH consisting of SEQ ID NO: 219.
  • the antibody or antigen-binding fragment thereof comprises a VL comprising an amino acid sequence that is at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% identical to SEQ ID NO: 220.
  • the antibody or antigen-binding fragment thereof comprises a VL comprising an amino acid sequence of SEQ ID NO: 220.
  • the antibody or antigen- binding fragment thereof comprises a VL consisting of SEQ ID NO: 220.
  • the antibody or antigen-binding fragment thereof comprises a VH comprising an amino acid sequence that is at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% identical to SEQ ID NO: 219 and a VL comprising an amino acid sequence that is at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% identical to SEQ ID NO: 220.
  • the antibody or antigen-binding fragment thereof comprises a VH comprising an amino acid sequence of SEQ ID NO: 219 and a VL comprising an amino acid sequence of SEQ ID NO: 220. In some embodiments, the antibody or antigen-binding fragment thereof comprises a VH consisting of SEQ ID NO: 219 and a VL consisting of SEQ ID NO: 220. [00178] In some embodiments, the antibody or antigen-binding fragment thereof binds to an epitope within the N-terminal domain (NTD) of the S polypeptide. In some embodiments, the antibody or antigen-binding fragment thereof binds to an epitope within the receptor binding domain (RBD) of the S polypeptide.
  • NTD N-terminal domain
  • RBD receptor binding domain
  • the antibody or antigen-binding fragment thereof binds to an epitope within the subdomain (SD) of the S polypeptide. In some embodiments, the antibody or antigen-binding fragment thereof binds to an epitope within the S2 subunit of the S polypeptide. [00179] In some embodiments, the antibody or antigen-binding fragment thereof binds to an epitope outside of the N-terminal domain (NTD) of the S polypeptide. In some embodiments, the antibody or antigen-binding fragment thereof binds to an epitope outside of the receptor binding domain (RBD) of the S polypeptide.
  • NTD N-terminal domain
  • RBD receptor binding domain
  • the antibody or antigen-binding fragment thereof binds to an epitope outside of the subdomain (SD) of the S polypeptide. In some embodiments, the antibody or antigen-binding fragment thereof binds to an epitope outside of the S2 subunit of the S polypeptide. [00180] In some embodiments, the antibody or antigen-binding fragment thereof is a neutralizing antibody or antigen-binding fragment thereof. In some embodiments, the antibody or antigen-binding fragment thereof neutralizes SARS-CoV-2. In some embodiments, the antibody or antigen-binding fragment thereof inhibits the fusion of SARS-CoV-2 and host cell membrane.
  • the antibody or antigen-binding fragment thereof is a non- neutralizing antibody or antigen-binding fragment thereof. In some embodiments, the antibody or antigen-binding fragment thereof is cytotoxic to a SARS-CoV-2 infected host cell.
  • the monoclonal antibodies, or antigen-binding fragments thereof, described herein are an IgG1, IgG2, IgG3, or IgG4 isotype. In some embodiments, the monoclonal antibodies, or antigen-binding fragments thereof, described herein are IgG1 isotypes.
  • the antibody or antigen-binding fragment thereof comprises (a) a first target binding site that specifically binds to an epitope within the S polypeptide, and (b) a second target binding site that binds to a different epitope on the S polypeptide or on a different molecule.
  • the multivalent antibody is a bivalent or bispecific antibody.
  • the antibody or the antigen-binding fragment thereof further comprises a variant Fc constant region.
  • the antibody is a monoclonal antibody.
  • the antibody is a chimeric antibody, a humanized antibody, or a humanized monoclonal antibody.
  • the antibody or antigen-binding fragment thereof is a single-chain antibody, Fab or Fab2 fragment.
  • the antibody or the antigen-binding fragment thereof further comprises a variant Fc constant region.
  • the antibody can be a monoclonal antibody.
  • the antibody can be a chimeric antibody, a humanized antibody, or a humanized monoclonal antibody.
  • the antibody or antigen-binding fragment thereof can be a single-chain antibody, Fab or Fab2 fragment.
  • the antibody or antigen-binding fragment thereof can be detectably labeled or conjugated to a toxin, a therapeutic agent, a polymer (e.g., polyethylene glycol (PEG)), a receptor, an enzyme, or a receptor ligand.
  • a toxin e.g., a tetanus toxin
  • Such antibodies may be used to treat animals, including humans, that are infected with the virus that is etiologically linked to SARS- CoV-2.
  • the toxin-coupled antibody is thought to bind to a portion of a spike protein presented on an infected cell, and then kill the infected cell.
  • an antibody of the present invention may be coupled to a detectable tag.
  • detectable tags include fluorescent proteins (i.e., green fluorescent protein, red fluorescent protein, yellow fluorescent protein), fluorescent markers (i.e., fluorescein isothiocyanate, rhodamine, texas red), radiolabels (i.e., 3H, 32P, 125I), enzymes (i.e. ⁇ -JDODFWRVLGDVH ⁇ KRUVHUDGLVK ⁇ SHUR[LGDVH ⁇ -glucuronidase, alkaline phosphatase), or an affinity tag (i.e., avidin, biotin, streptavidin).
  • fluorescent proteins i.e., green fluorescent protein, red fluorescent protein, yellow fluorescent protein
  • fluorescent markers i.e., fluorescein isothiocyanate, rhodamine, texas red
  • radiolabels i.e., 3H, 32P, 125I
  • enzymes i.e. ⁇ -JDODF
  • an antibody provided herein is an antibody fragment.
  • Antibody fragments include, but are not limited to, Fab, Fab', Fab'-SH, F(ab')2, Fv, and single-chain Fv (scFv) fragments, and other fragments described below, e.g., diabodies, triabodies tetrabodies, and single- domain antibodies.
  • Fab fragment antigen binding protein
  • Diabodies are antibody fragments with two antigen-binding sites that may be bivalent or bispecific.
  • Single-domain antibodies are antibody fragments comprising all or a portion of the heavy chain variable domain or all or a portion of the light chain variable domain of an antibody.
  • a single-domain antibody is a human single-domain antibody (DOMANTIS, Inc., Waltham, Mass.; see, e.g., U.S.
  • Antibody fragments can be made by various techniques, including but not limited to proteolytic digestion of an intact antibody as well as production by recombinant host cells (e.g., E. coli or phage), as described herein.
  • Chimeric and Humanized Antibodies [00192] In some embodiments, an antibody provided herein is a chimeric antibody. Certain chimeric antibodies are described, e.g., in U.S. Pat. No.4,816,567; and Morrison et al., Proc. Natl. Acad. Sci. USA, 81:6851-6855 (1984)).
  • a chimeric antibody comprises a non-human variable region (e.g., a variable region derived from a mouse, rat, hamster, rabbit, or non- human primate, such as a monkey) and a human constant region.
  • a chimeric antibody is a “class switched” antibody in which the class or subclass has been changed from that of the parent antibody. Chimeric antibodies include antigen-binding fragments thereof.
  • a chimeric antibody is a humanized antibody.
  • a non- human antibody is humanized to reduce immunogenicity to humans, while retaining the specificity and affinity of the parental non-human antibody.
  • a humanized antibody comprises one or more variable domains in which HVRs, e.g., CDRs, (or portions thereof) are derived from a non-human antibody, and FRs (or portions thereof) are derived from human antibody sequences.
  • a humanized antibody optionally will also comprise at least a portion of a human constant region.
  • some FR residues in a humanized antibody are substituted with corresponding residues from a non-human antibody (e.g., the antibody from which the HVR residues are derived), e.g., to restore or improve antibody specificity or affinity.
  • Human framework regions that may be used for humanization include but are not limited to: framework regions selected using the “best-fit” method (see, e.g., Sims et al. J. Immunol.
  • framework regions derived from the consensus sequence of human antibodies of a particular subgroup of light or heavy chain variable regions see, e.g., Carter et al. Proc. Natl. Acad. Sci. USA, 89:4285 (1992); and Presta et al. J. Immunol., 151:2623 (1993)); human mature (somatically mutated) framework regions or human germline framework regions (see, e.g., Almagro and Fransson, Front. Biosci.13:1619-1633 (2008)); and framework regions derived from screening FR libraries (see, e.g., Baca et al., J. Biol.
  • an antibody provided herein is a human antibody.
  • Human antibodies can be produced using various techniques known in the art or using techniques described herein. Human antibodies are described generally in van Dijk and van de Winkel, Curr. Opin. Pharmacol.5: 368-74 (2001) and Lonberg, Curr. Opin. Immunol.20:450-459 (2008).
  • Human antibodies may be prepared by administering an immunogen to a transgenic animal that has been modified to produce intact human antibodies or intact antibodies with human variable regions in response to antigenic challenge.
  • Such animals typically contain all or a portion of the human immunoglobulin loci, which replace the endogenous immunoglobulin loci, or which are present extrachromosomally or integrated randomly into the animal's chromosomes.
  • the endogenous immunoglobulin loci have generally been inactivated.
  • Human variable regions from intact antibodies generated by such animals may be further modified, e.g., by combining with a different human constant region.
  • Human antibodies can also be made by hybridoma-based methods. Human myeloma and mouse-human heteromyeloma cell lines for the production of human monoclonal antibodies have been described. (See, e.g., Kozbor J. Immunol., 133: 3001 (1984); Brodeur et al., Monoclonal Antibody Production Techniques and Applications, pp.
  • Human hybridoma technology (Trioma technology) is also described in Vollmers and Brandlein, Histology and Histopathology, 20(3):927-937 (2005) and Vollmers and Brandlein, Methods and Findings in Experimental and Clinical Pharmacology, 27(3):185-91 (2005).
  • Human antibodies may also be generated by isolating Fv clone variable domain sequences selected from human-derived phage display libraries. Such variable domain sequences may then be combined with a desired human constant domain. Techniques for selecting human antibodies from antibody libraries are described below.
  • Antibodies of the invention may be isolated by screening combinatorial libraries for antibodies with the desired activity or activities.
  • phage display methods repertoires of VH and VL genes are separately cloned by polymerase chain reaction (PCR) and recombined randomly in phage libraries, which can then be screened for antigen-binding phage as described in Winter et al., Ann. Rev. Immunol., 12: 433- 455 (1994). Phage typically displays antibody fragments, either as scFv fragments or as Fab fragments. Libraries from immunized sources provide high-affinity antibodies to the immunogen without the requirement of constructing hybridomas.
  • PCR polymerase chain reaction
  • naive repertoire can be cloned (e.g., from human) to provide a single source of antibodies to a wide range of non-self and also self-antigens without any immunization as described by Griffiths et al., EMBO J, 12: 725-734 (1993).
  • naive libraries can also be made synthetically by cloning unrearranged V-gene segments from stem cells and using PCR primers containing random sequences to encode the highly variable CDR3 regions and to accomplish rearrangement in vitro, as described by Hoogenboom and Winter, J. Mol. Biol., 227: 381-388 (1992).
  • Patent publications describing human antibody phage libraries include, for example, U.S. Pat. No.
  • amino acid sequence variants of the antibodies provided herein are contemplated. For example, it may be desirable to improve the binding affinity and/or other biological properties of the antibody. Amino acid sequence variants of an antibody may be prepared by introducing appropriate modifications into the nucleotide sequence encoding the antibody, or by peptide synthesis.
  • Such modifications include, for example, deletions from, and/or insertions into and/or substitutions of residues within the amino acid sequences of the antibody. Any combination of deletion, insertion, and substitution can be made to arrive at the final construct, provided that the final construct possesses the desired characteristics, e.g., antigen- binding. [00203] Substitution of one or more CDR residues or omission of one or more CDRs is also possible. Antibodies have been described in the scientific literature in which one or two CDRs can be dispensed with for binding. Padlan et al. (1995 FASEB J.
  • CDR residues not contacting antigen can be identified based on previous studies (for example residues H60-H65 in CDRH2 are often not required), from regions of Kabat CDRs lying outside Chothia CDRs, by molecular modeling and/or empirically.
  • a CDR or residue(s) thereof is omitted, it is usually substituted with an amino acid occupying the corresponding position in another human antibody sequence or a consensus of such sequences. Positions for substitution within CDRs and amino acids to substitute can also be selected empirically.
  • the fragments, whether attached to other sequences or not, can also include insertions, deletions, substitutions, or other selected modifications of particular regions or specific amino acids residues, provided the activity of the antibody or antibody fragment is not significantly altered or impaired compared to the non-modified antibody or antibody fragment. These modifications can provide for some additional property, such as to remove/add amino acids capable of disulfide bonding, to increase its bio-longevity, to alter its secretory characteristics, etc.
  • the antibody or antibody fragment must possess a bioactive property, such as specific binding to its cognate antigen.
  • Functional or active regions of the antibody or antibody fragment may be identified by mutagenesis of a specific region of the protein, followed by expression and testing of the expressed polypeptide. Such methods are readily apparent to a skilled practitioner in the art and can include site-specific mutagenesis of the nucleic acid encoding the antibody or antibody fragment. (Zoller, M.J. Curr. Opin. Biotechnol.3:348-354, 1992).
  • Protein variants and derivatives are well understood to those of skill in the art and in can involve amino acid sequence modifications.
  • amino acid sequence modifications typically fall into one or more of three classes: substitutional, insertional or deletional variants.
  • insertions refer to a change in an amino acid or nucleotide sequence resulting in the addition of one or more amino acid or nucleotide residues, respectively, as compared to the parent, often the naturally occurring, molecule. Insertions include amino and/or carboxyl terminal fusions as well as intrasequence insertions of single or multiple amino acid residues. Insertions ordinarily will be smaller insertions than those of amino or carboxyl terminal fusions, for example, on the order of one to four residues.
  • Immunogenic fusion protein derivatives are made by fusing a polypeptide sufficiently large to confer immunogenicity to the target sequence by cross-linking in vitro or by recombinant cell culture transformed with DNA encoding the fusion.
  • Deletions are characterized by the removal of one or more amino acid residues from the protein sequence. Typically, no more than about from 2 to 6 residues are deleted at any one site within the protein molecule.
  • These variants ordinarily are prepared by site specific mutagenesis of nucleotides in the DNA encoding the protein, thereby producing DNA encoding the variant, and thereafter expressing the DNA in recombinant cell culture.
  • substitution mutations at predetermined sites in DNA having a known sequence are well known, for example M13 primer mutagenesis and PCR mutagenesis.
  • Amino acid substitutions are typically of single residues but can occur at a number of different locations at once; insertions usually will be on the order of about from 1 to 10 amino acid residues; and deletions will range about from 1 to 30 residues.
  • Deletions or insertions preferably are made in adjacent pairs, i.e., a deletion of 2 residues or insertion of 2 residues. Substitutions, deletions, insertions, or any combination thereof may be combined to arrive at a final construct.
  • the mutations must not place the sequence out of reading frame and preferably will not create complementary regions that could produce secondary mRNA structure.
  • Substitutional variants are those in which at least one residue has been removed and a different residue inserted in its place. Such substitutions generally are made in accordance with the following Table B and are referred to as conservative substitutions.
  • Table A Amino Acid Abbreviations
  • Table B Amino Acid Substitutions - Exemplary Conservative Substitutions
  • Substantial changes in function or immunological identity are made by selecting substitutions that are less conservative than those in Table B, i.e., selecting residues that differ more significantly in their effect on maintaining (a) the structure of the polypeptide backbone in the area of the substitution, for example as a sheet or helical conformation, (b) the charge or hydrophobicity of the molecule at the target site or (c) the bulk of the side chain.
  • Conservative amino acid substitutions include the ones in which the amino acid residue is replaced with an amino acid residue having similar structural or chemical properties. Families of amino acid residues having similar side chains have been defined in the art. These families include amino acids with basic side chains (e.g., lysine, arginine, histidine), acidic side chains (e.g., aspartic acid, glutamic acid), uncharged polar side chains (e.g., glycine, asparagine, glutamine, serine, threonine, tyrosine, cystine, tryptophan), nonpolar side chains (e.g., alanine, valine, leucine, isoleucine, proline, phenylalanine, methionine), beta-branched side chains (e.g., threonine, valine, isoleucine) and aromatic side chains (e.g., tyrosine, phenylalanine, tryptophan, histidine).
  • basic side chains e.
  • substitutions which in general are expected to produce the greatest changes in the protein properties will be those in which (a) a hydrophilic residue, e.g. seryl or threonyl, is substituted for (or by) a hydrophobic residue, e.g.
  • an electropositive side chain e.g., lysyl, arginyl, or histidyl
  • an electronegative residue e.g., glutamyl or aspartyl
  • substitutions include combinations such as, for example, Gly, Ala; Val, Ile, Leu; Asp, Glu; Asn, Gln; Ser, Thr; Lys, Arg; and Phe, Tyr. Such conservatively substituted variations of each explicitly disclosed sequence are included within the mosaic polypeptides provided herein.
  • substitution, Insertion, and Deletion Variants [00210] In some embodiments, antibody variants having one or more amino acid substitutions are provided.
  • Sites of interest for substitutional mutagenesis include the HVRs and FRs.
  • Conservative substitutions are defined herein.
  • Amino acid substitutions may be introduced into an antibody of interest and the products screened for a desired activity, e.g., retained/improved antigen-binding, decreased immunogenicity, or improved antibody-dependent cell-mediated cytotoxicity (ADCC) and complement-dependent cytotoxicity (CDC).
  • ADCC antibody-dependent cell-mediated cytotoxicity
  • CDC complement-dependent cytotoxicity
  • an antibody of the invention can comprise one or more conservative modifications of the CDRs, heavy chain variable region, or light variable regions described herein.
  • a conservative modification or functional equivalent of a peptide, polypeptide, or protein disclosed in this invention refers to a polypeptide derivative of the peptide, polypeptide, or protein, e.g., a protein having one or more point mutations, insertions, deletions, truncations, a fusion protein, or a combination thereof. It substantially retains the activity to of the parent peptide, polypeptide, or protein (such as those disclosed in this invention).
  • a conservative modification or functional equivalent is at least 60% (e.g., any number between 60% and 100%, inclusive, e.g., 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, and 99%) identical to a parent.
  • heavy chain variable region or light variable regions having one or more point mutations, insertions, deletions, truncations, a fusion protein, or a combination thereof, as well as antibodies having the variant regions.
  • percent homology between two amino acid sequences is equivalent to the percent identity between the two sequences.
  • the comparison of sequences and determination of percent identity between two sequences can be accomplished using a mathematical algorithm, as described in the non-limiting examples below.
  • the percent identity between two amino acid sequences can be determined using the algorithm of E. Meyers and W. Miller (Comput. Appl. Biosci., 4:11-17 (1988)) which has been incorporated into the ALIGN program (version 2.0), using a PAM120 weight residue table, a gap length penalty of 12 and a gap penalty of 4.
  • the percent identity between two amino acid sequences can be determined using the Needleman and Wunsch (J. Mol. Biol.
  • the protein sequences of the present invention can further be used as a “query sequence” to perform a search against public databases to, for example, identify related sequences. Such searches can be performed using the XBLAST program (version 2.0) of Altschul, et al. (1990) J. Mol. Biol. 215:403-10.
  • Gapped BLAST can be utilized as described in Altschul et al., (1997) Nucleic Acids Res. 25(17):3389-3402.
  • the default parameters of the respective programs e.g., XBLAST and NBLAST
  • XBLAST and NBLAST See ncbi.nlm.nih.gov).
  • conservative modifications refers to amino acid modifications that do not significantly affect or alter the binding characteristics of the antibody containing the amino acid sequence. Such conservative modifications include amino acid substitutions, additions, and deletions. Modifications can be introduced into an antibody of the invention by standard techniques known in the art, such as site-directed mutagenesis and PCR- mediated mutagenesis. Conservative amino acid substitutions are ones in which the amino acid residue is replaced with an amino acid residue having a similar side chain. Families of amino acid residues having similar side chains have been defined in the art.
  • amino acids with basic side chains e.g., lysine, arginine, histidine
  • acidic side chains e.g., aspartic acid, glutamic acid
  • uncharged polar side chains e.g., glycine, asparagine, glutamine, serine, threonine, tyrosine, cysteine, tryptophan
  • nonpolar side chains e.g., alanine, valine, leucine, isoleucine, proline, phenylalanine, methionine
  • beta-branched side chains e.g., threonine, valine, isoleucine
  • aromatic side chains e.g., tyrosine, phenylalanine, tryptophan, histidine
  • Non-conservative substitutions will entail exchanging a member of one of these classes for another class.
  • An exemplary substitutional variant is an affinity matured antibody, which may be conveniently generated, e.g., using phage display-based affinity maturation techniques such as those described in, e.g., Hoogenboom et al., in Methods in Molecular Biology 178:1-37 (O'Brien et al., ed., Human Press, Totowa, N.J., (2001).
  • Amino acid sequence insertions include amino- and/or carboxyl-terminal fusions ranging in length from one residue to polypeptides containing a hundred or more residues, as well as intrasequence insertions of single or multiple amino acid residues.
  • terminal insertions include an antibody with an N-terminal methionyl residue.
  • Other insertional variants of the antibody molecule include the fusion to the N- or C- terminus of the antibody to an enzyme (e.g., for ADEPT) or a polypeptide which increases the serum half-life of the antibody.
  • Glycosylation Variants [00218]
  • an antibody provided herein is altered to increase or decrease the extent to which the antibody is glycosylated.
  • Addition or deletion of glycosylation sites to an antibody may be conveniently accomplished by altering the amino acid sequence such that one or more glycosylation sites are created or removed.
  • an aglycoslated antibody can be made (i.e., the antibody lacks glycosylation).
  • Glycosylation can be altered to, for example, increase the affinity of the antibody for antigen.
  • carbohydrate modifications can be accomplished by, for example, altering one or more sites of glycosylation within the antibody sequence.
  • one or more amino acid substitutions can be made that result in elimination of one or more variable region framework glycosylation sites to thereby eliminate glycosylation at that site.
  • Such aglycosylation may increase the affinity of the antibody for antigen.
  • Glycosylation of the constant region on N297 may be prevented by mutating the N297 residue to another residue, e.g., N297A, and/or by mutating an adjacent amino acid, e.g., 298 to thereby reduce glycosylation on N297.
  • an antibody can be made that has an altered type of glycosylation, such as a hypofucosylated antibody having reduced amounts of fucosyl residues or an antibody having increased bisecting GlcNac structures.
  • Such altered glycosylation patterns have been demonstrated to increase the ADCC ability of antibodies.
  • Such carbohydrate modifications can be accomplished by, for example, expressing the antibody in a host cell with altered glycosylation machinery.
  • Cells with altered glycosylation machinery have been described in the art and can be used as host cells in which to express recombinant antibodies described herein to thereby produce an antibody with altered glycosylation.
  • EP 1,176,195 by Hanai et al. describes a cell line with a functionally disrupted FUT8 gene, which encodes a fucosyltransferase, such that antibodies expressed in such a cell line exhibit hypofucosylation.
  • PCT Publication WO 03/035835 by Presta describes a variant Chinese Hamster Ovary cell line, Led 3 cells, with reduced ability to atach fucose to Asn(297)-lmked carbohydrates, also resulting in hypofucosylation of antibodies expressed in that host cell (see also Shields, R.L. et al. (2002) J . Biol. Chem. 277:26733-26740).
  • glycoprotein-modifying glycosyltransferases e.g., beta(l,4)-N- acetylglucosammyltransferase III (GnTIII)
  • GnTIII glycoprotein-modifying glycosyltransferases
  • variable regions of the antibody described herein can be linked (e.g., covalently linked or fused) to an Fc, e.g., an IgGl, IgG2, IgG3 or IgG4 Fc, which may be of any allotype or isoallotype, e.g., for IgGl: Glm, Glml(a), Glm2(x), Glm3(f), Glml7(z); for IgG2: G2m, G2m23(n); for IgG3: G3m, G3m21(gl), G3m28(g5), G3ml l(b0), G3m5(bl), G3ml3(b3), G3ml4(b4), G3ml0(b5), G3ml5(s), G3ml6(t), G3m6(c3), G3m24(c5), G3m26(u), G3m27(
  • the antibodies variable regions described herein are linked to an Fc that binds to one or more activating Fc receptors (Fcyl, Fcylla, or Fcyllla), and thereby stimulate ADCC and may cause T cell depletion. In some embodiments, the antibody variable regions described herein are linked to an Fc that causes depletion.
  • the antibody variable regions described herein may be linked to an Fc comprising one or more modifications, typically to alter one or more functional properties of the antibody, such as serum half-life, complement fixation, Fc receptor binding, and/or antigendependent cellular cytotoxicity.
  • an antibody described herein may be chemically modified (e.g., one or more chemical moieties can be atached to the antibody) or be modified to alter its glycosylation, to alter one or more functional properties of the antibody.
  • the numbering of residues in the Fc region is that of the EU index of Kabat.
  • the Fc region encompasses domains derived from the constant region of an immunoglobulin, preferably a human immunoglobulin, including a fragment, analog, variant, mutant, or derivative of the constant region.
  • Suitable immunoglobulins include IgGl, IgG2, IgG3, IgG4, and other classes such as IgA, IgD, IgE, and IgM.
  • the constant region of an immunoglobulin is defined as a naturally- occurring or synthetically-produced polypeptide homologous to the immunoglobulin C -terminal region, and can include a CHI domain, a hinge, a CH2 domain, a CH3 domain, or a CH4 domain, separately or in combination.
  • an antibody of this invention has an Fc region other than that of a wild-type IgAl .
  • the antibody can have an Fc region from that of IgG (e.g., IgGl, IgG2, IgG3, and IgG4) or other classes such as IgA2, IgD, IgE, and IgM.
  • the Fc can be a mutant form of IgAl.
  • the constant region of an immunoglobulin is responsible for many important antibody functions, including Fc receptor (FcR) binding and complement fixation.
  • FcR Fc receptor
  • IgG is separated into four subclasses known as IgGl, IgG2, IgG3, and IgG4.
  • Ig molecules interact with multiple classes of cellular receptors.
  • IgG molecules interact with three classes of Fey receptors (FcyR) specific for the IgG class of antibody, namely FcyRI, FcyRII, and FcyRUL.
  • FcyR Fey receptors
  • the important sequences for the binding of IgG to the FcyR receptors have been reported to be located in the CH2 and CH3 domains.
  • the serum half-life of an antibody is influenced by the ability of that antibody to bind to an FcR.
  • the Fc region is a variant Fc region, e.g., an Fc sequence that has been modified (e.g., by amino acid substitution, deletion and/or insertion) relative to a parent Fc sequence (e.g., an unmodified Fc polypeptide that is subsequently modified to generate a variant), to provide desirable structural features and/or biological activity.
  • a variant Fc region e.g., an Fc sequence that has been modified (e.g., by amino acid substitution, deletion and/or insertion) relative to a parent Fc sequence (e.g., an unmodified Fc polypeptide that is subsequently modified to generate a variant), to provide desirable structural features and/or biological activity.
  • modifications in the Fc region in order to generate an Fc variant that (a) has increased or decreased ADCC, (b) increased or decreased CDC, (c) has increased or decreased affinity for Clq and/or (d) has increased or decreased affinity for an Fc receptor relative to the parent Fc.
  • Such Fc region variants will generally comprise at least, one ammo acid modification in the Fc region. Combining amino acid modifications is thought to be particularly desirable.
  • the variant Fc region may include two, three, four, five, etc., substitutions therein, e.g., of the specific Fc region positions identified herein.
  • a variant Fc region may also comprise a sequence alteration wherein amino acids involved in disulfide bond formation are removed or replaced with other amino acids. Such removal may avoid reaction with other cysteine-containing proteins present in the host cell used to produce the antibodies described herein. Even when cy steine residues are removed, single chain Fc domains can still form a dimeric Fc domain that is held together non-covalently.
  • the Fc region may be modified to make it more compatible with a selected host cell. For example, one may remove the PA sequence near the N-terminus of a typical native Fc region, which may be recognized by a digestive enzyme in E. coli, such as proline iminopeptidase.
  • one or more glycosylation sites within the Fc domain may be removed. Residues that are typically glycosylated (e.g., asparagine) may confer cytolytic response. Such residues may be deleted or substituted with unglycosylated residues (e.g., alanine).
  • sites involved in interaction with complement such as the Clq binding site, may be removed from the Fc region. For example, one may delete or substitute the EKK sequence of human IgGl.
  • sites that affect binding to Fc receptors may be removed, preferably sites other than salvage receptor binding sites.
  • an Fc region may be modified to remove an ADCC site.
  • ADCC sites are known in the art; see, for example, Molec. Immunol. 29 (5): 633-9 (1992) with regard to ADCC sites in IgGl. Specific examples of variant Fc domains are disclosed, for example, in WO 97/34631 and WO 96/32478.
  • the hinge region of Fc is modified such that the number of cysteine residues in the hinge region is altered, e.g., increased or decreased. This approach is described further in U.S. Patent No. 5,677,425 by Bodmer et al. The number of cysteine residues in the hinge region of Fc is altered to, for example, facilitate assembly of the light and heavy chains or to increase or decrease the stability of the antibody.
  • the Fc hinge region of an antibody is mutated to decrease the biological half-life of the antibody. More specifically, one or more amino acid mutations are introduced into the CH2-CH3 domain interface region of the Fc- hinge fragment such that the antibody has impaired Staphylococcal protein A (SpA) binding relative to native Fc-hinge domain SpA binding.
  • SpA Staphylococcal protein A
  • the Fc region is altered by replacing at least one amino acid residue with a different amino acid residue to alter the effector function(s) of the antibody.
  • one or more amino acids selected from amino acid residues 234, 235, 236, 237, 297, 318, 320, and 322 can be replaced with a different amino acid residue such that the antibody has an altered affinity for an effector ligand but retains the antigen-binding ability of the parent antibody.
  • the effector ligand to which affinity is altered can be, for example, an Fc receptor or the CI component of complement. This approach is described in further detail in U.S. Patent Nos. 5,624,821 and 5,648,260, both by Winter et al.
  • one or more amino acids selected from amino acid residues 329, 331, and 322 can be replaced with a different ammo acid residue such that the antibody has altered Clq binding and/or reduced or abolished CDC.
  • This approach is described in further detail in U.S. Patent Nos. 6,194,551 by Idusogie et al.
  • one or more amino acid residues within amino acid positions 231 and 239 are altered to thereby alter the ability of the antibody to fix complement. This approach is described further in PCT Publication WO 94/29351 by Bodmer et al.
  • the Fc region may be modified to increase ADCC and/or to increase the affinity for an Fey receptor by modifying one or more amino acids at the following positions: 234, 235, 236, 238, 239, 240, 241 , 243, 244, 245, 247, 248, 249, 252, 254, 255, 256, 258, 262, 263, 264, 265, 267, 268, 269, 270, 272, 276, 278, 280, 283, 285, 286, 289, 290, 292, 293,
  • substitutions include 236A, 239D, 239E,
  • Fc modifications that increase binding to an Fey receptor include amino acid modifications at any one or more of amino acid positions 238, 239, 248, 249, 252, 254, 255, 256, 258, 265, 267, 268, 269, 270, 272, 279, 280, 283, 285, 298, 289, 290, 292, 293, 294, 295, 296, 298, 301, 303, 305, 307, 312, 315, 324, 327, 329, 330, 335, 337, 3338, 340, 360, 373, 376, 379, 382, 388, 389, 398, 414, 416, 419, 430, 434, 435, 437, 438 or 439 of the Fc region, wherein the numbering of the residues in the Fc region is that of the EU index as in Kabat (WO00/42072).
  • Fc modifications that can be made to Fes are those for reducing or ablating binding to FcyR and/or complement proteins, thereby reducing or ablating Fc-mediated effector functions such as ADCC, antibody-dependent cellular phagocytosis (ADCP), and CDC.
  • Exemplary modifications include but are not limited to substitutions, insertions, and deletions at positions 234, 235, 236, 237, 267, 269, 325, and 328, wherein numbering is according to the EU index.
  • Exemplary substitutions include but are not limited to 234G, 235G, 236R, 237K, 267R, 269R, 325L, and 328R, wherein numbering is according to the EU index.
  • An Fc variant may comprise 236R/328R.
  • Other modifications for reducing FcyR and complement interactions include substitutions 297A, 234 A, 235A, 237A, 318A, 228P, 236E, 268Q, 309L, 330S, 331 S, 220S, 226S, 229S, 238S, 233P, and 234V, as well as removal of the glycosylation at position 297 by mutational or enzymatic means or by production in organisms such as bacteria that do not glycosylate proteins.
  • the Fc region may comprise a non-naturally occurring amino acid residue at additional and/or alternative positions known to one skilled in the art (see, e.g., ELS. Pat. Nos. 5,624,821; 6,277,375; 6,737,056; 6,194,551; 7,317,091; 8,101,720; WO00/42072; WOOl/58957; W002/06919; WO04/016750; W004/029207; WO04/035752; WO04/074455; WO04/099249; W004/063351; W005/070963; W005/040217, WO05/092925 and W006/020114).
  • Fc variants that enhance affinity for an inhibitory receptor FcyRIIb may also be used. Such variants may provide an Fc fusion protein with immune-modulatory activities related to FcyRIIb cells, including, for example, B cells and monocytes. In one embodiment, the Fc variants provide selectively enhanced affinity to FcyRIIb relative to one or more activating receptors. Modifications for altering binding; to FcyRIIb include one or more modifications at a position selected from the group consisting of 234, 235, 236, 237, 239, 266, 267, 268, 325, 326, 327, 328, and 332, according to the EU index.
  • Exemplary substitutions for enhancing FcyRIIb affinity include but are not limited to 234D, 234E, 234F, 234W, 235D, 235F, 235R, 235Y, 236D, 236N, 237D, 237N, 239D, 239E, 266M, 267D, 267E, 268D, 268E, 327D, 327E, 328F, 328W, 328Y, and 332E.
  • Exemplary substitutions include 235Y, 236D, 239D, 266M, 267E, 268D, 268E, 328F, 328W, and 328Y.
  • Fc variants for enhancing binding to FcyRIIb include 235Y/267E, 236D/267E, 239D/268D, 239D/267E, 267E/268D, 267E/268E, and 267E/328F.
  • the affinities and binding properties of an Fc region for its ligand may be determined by a variety of in vitro assay methods (biochemical or immunological based assays) known in the art, including but not limited to equilibrium methods (e.g., ELISA, or radioimmunoassay), or kinetics (e.g., BIACORETM analysis), and other methods such as indirect binding assays, competitive inhibition assays, fluorescence resonance energy transfer (FRET), gel electrophoresis and chromatography (e.g., gel filtration).
  • in vitro assay methods biochemical or immunological based assays known in the art, including but not limited to equilibrium methods (e.g., ELISA, or radioimmunoassay), or kinetics (e.g., BIACORETM analysis), and other methods such as indirect binding assays, competitive inhibition assays, fluorescence resonance energy transfer (FRET), gel electrophoresis and chromatography (e.g., gel filtration).
  • these and other methods may utilize a label on one or more of the components being examined and/or employ a variety of detection methods, including but not limited to chromogenic, fluorescent, luminescent, or isotopic labels.
  • detection methods including but not limited to chromogenic, fluorescent, luminescent, or isotopic labels.
  • a detailed description of binding affinities and kinetics can be found in Paul, W. E., ed., Fundamental Immunology, 4th Ed., Lippincott-Raven, Philadelphia (1999), which focuses on antibody-immunogen interactions.
  • the antibody is modified to increase its biological half-life.
  • Various approaches are possible. For example, this may be done by increasing the binding affinity of the Fc region for FcRn.
  • one or more of the following residues can be mutated: 252, 254, 256, 433, 435, 436, as described in U.S. Pat. No. 6,277,375.
  • Specific exemplary substitutions include one or more of the following: T252L, T254S, and/or T256F.
  • the antibody can be altered within the CH1 or CL region to contain a salvage receptor binding epitope taken from two loops of a CH2 domain of an Fc region of an IgG, as described in U.S. Patent Nos. 5,869,046 and 6,121,022 by Presta et al.
  • variants that increase binding to FcRn and/or improve pharmacokinetic properties include substitutions at positions 259, 308, 428, and 434, including for example 259I, 308F, 428L, 428M, 434S, 434H, 434F, 434Y, and 434M.
  • variants that increase Fc binding to FcRn include: 250E, 250Q, 428L, 428F, 250Q/428L (Hinton et al friendship 2004, J. Biol. Chem.
  • hybrid IgG isotypes with particular biological characteristics may be used.
  • an IgGl/IgG3 hybrid variant may be constructed by substituting IgG 1 positions in the CH2 and/or CH3 region with the amino acids from IgG3 at positions where the two isotypes differ.
  • a hybrid variant IgG antibody may be constructed that comprises one or more substitutions, e.g., 274Q, 276K, 300F, 339T, 356E, 358M, 384S, 392N, 397M, 422I, 435R, and 436F.
  • an IgGl/IgG2 hybrid variant may be constructed by substituting IgG2 positions in the CH2 and/or CH3 region with amino acids from IgGl at positions where the two isotypes differ.
  • a hybrid variant IgG antibody may be constructed chat comprises one or more substitutions, e.g., one or more of the following ammo acid substitutions: 233E, 234L, 235L, 236G (referring to an insertion of a glycine at position 236), and 321 h.
  • IgGl variants with strongly enhanced binding to FcyRIIIa have been identified, including variants with S239D/I332E and S239D/I332E/A330L mutations which showed the greatest increase in affinity for FcyRIIIa, a decrease in FcyRIIb binding, and strong cytotoxic activity in cynomolgus monkeys (Lazar et al., 2006).
  • IgGl mutants containing L235V, F243L, R292P, Y300L, and P396L mutations which exhibited enhanced binding to FcyRIIIa and concomitantly enhanced ADCC activity' in transgenic mice expressing human FcyRIIIa in models of B cell malignancies and breast cancer have been identified (Stavenhagen et al., 2007; Nordstrom et al., 2011).
  • Other Fc mutants that, may be used include S298A/E333A/L334A, S239D/1332E, S239D/I332E/A330L, L235V/F243L/R292P/Y300L/ P396L, and M428L/N434S.
  • an Fc is chosen that has reduced binding to FcyRs.
  • An exemplary Fc e.g., IgGl Fc, with reduced FcyR binding, comprises the following three amino acid substitutions: L234A, L235E, and G237A.
  • an Fc is chosen that has reduced complement fixation.
  • An exemplary Fc e.g., IgGl Fc, with reduced complement fixation, has the following two ammo acid substitutions: A330S and P331S.
  • an Fc is chosen that has essentially no effector function, i.e., it has reduced binding to FcyRs and reduced complement fixation.
  • An exemplary Fc e.g., IgGl Fc, that is effectorless, comprises the following five mutations: L234A, L235E, G237A, A330S, and P331S.
  • substitution S228P which mimics the hinge sequence in IgGl and thereby stabilizes IgG4 molecules.
  • the antibodies of the invention may be monovalent or multivalent (e.g., bivalent, trivalent, etc.).
  • valency refers to the number of potential target binding sites associated with an antibody. Each target binding site specifically binds one target molecule or specific position or locus on a target molecule. When an antibody is monovalent, each binding site of the molecule will specifically bind to a single antigen position or epitope. When an antibody comprises more than one target binding site (multivalent), each target binding site may specifically bind to the same or different molecules (e.g., may bind to different ligands or different antigens, or different epitopes or positions on the same antigen).
  • the antibodies are bispecific antibodies in which the two chains have different specificities, as described in Millstein et al., 1983, Nature, 305:537-539.
  • Other embodiments include antibodies with additional specificities, such as trispecific antibodies.
  • Other more sophisticated compatible multispecific constructs and methods of their fabrication are set forth in U.S.P.N. 2009/0155255, as well as WO 94/04690; Suresh et al., 1986, Methods in Enzymology, 121:210; and WO96/27011.
  • multivalent antibodies may immunospecifically bind to different epitopes of the desired target molecule or may immunospecifically bind to both the target molecule as well as a heterologous epitope, such as a heterologous polypeptide or solid support material.
  • the multivalent antibodies may include bispecific antibodies or trispecific antibodies.
  • Bispecific antibodies also include cross-linked or “heteroconjugate” antibodies.
  • one of the antibodies in the heteroconjugate can be coupled to avidin, the other to biotin.
  • Such antibodies have, for example, been proposed to target immune system cells to unwanted cells (U.S. Pat.
  • Heteroconjugate antibodies may be made using any convenient cross-linking methods. Suitable cross-linking agents are well known in the art and are disclosed in U.S. Pat. No.4,676,980, along with a number of cross-linking techniques.
  • antibody variable domains with the desired binding specificities are fused to immunoglobulin constant domain sequences, such as an immunoglobulin heavy chain constant domain comprising at least part of the hinge, CH2, and/or CH3 regions, using methods well known to those of ordinary skill in the art.
  • Antibody Derivatives [00249] An antibody provided herein may be further modified to contain additional nonproteinaceous moieties that are known in the art and readily available.
  • the moieties suitable for derivatization of the antibody include but are not limited to water-soluble polymers.
  • Non-limiting examples of water-soluble polymers include, but are not limited to, PEG, copolymers of ethylene glycol/propylene glycol, carboxymethylcellulose, dextran, polyvinyl alcohol, polyvinyl pyrrolidone, poly-1,3-dioxolane, poly-1,3,6-trioxane, ethylene/maleic anhydride copolymer, polyaminoacids (either homopolymers or random copolymers), and dextran or poly(n-vinyl pyrrolidone)polyethylene glycol, propropylene glycol homopolymers, polypropylene oxide/ethylene oxide co-polymers, polyoxyethylated polyols (e.g., glycerol), polyvinyl alcohol, and mixtures thereof.
  • PEG polyethylene glycol/propylene glycol
  • carboxymethylcellulose dextran
  • dextran polyvinyl alcohol
  • polyvinyl pyrrolidone poly-1,3
  • Polyethylene glycol propionaldehyde may have advantages in manufacturing due to its stability in water.
  • the polymer may be of any molecular weight and may be branched or unbranched.
  • the number of polymers attached to the antibody may vary, and if more than one polymer is attached, they can be the same or different molecules. In general, the number and/or type of polymers used for derivatization can be determined based on considerations including, but not limited to, the particular properties or functions of the antibody to be improved, whether the antibody derivative will be used in a therapy under defined conditions, etc.
  • conjugates of an antibody and nonproteinaceous moiety that may be selectively heated by exposure to radiation are provided.
  • the nonproteinaceous moiety is a carbon nanotube (Kam et al., Proc. Natl. Acad. Sci. USA 102: 11600- 11605 (2005)).
  • the radiation may be of any wavelength and includes, but is not limited to, wavelengths that do not harm ordinary cells, but which heat the nonproteinaceous moiety to a temperature at which cells proximal to the antibody-nonproteinaceous moiety are killed.
  • Another modification of the antibodies described herein is pegylation.
  • An antibody can be pegylated to, for example, increase the biological (e.g., serum) half-life of the antibody.
  • the antibody, or fragment thereof typically is reacted with PEG, such as a reactive ester or aldehyde derivative of PEG, under conditions in which one or more PEG groups become attached to the antibody or antibody fragment.
  • PEG such as a reactive ester or aldehyde derivative of PEG
  • the pegylation is carried out via an acylation reaction or an alkylation reaction with a reactive PEG molecule (or an analogous reactive water-soluble polymer).
  • the term “polyethylene glycol” is intended to encompass any of the forms of PEG that have been used to derivatize other proteins, such as mono (CI -CIO) alkoxy- or aryloxy-polyethylene glycol or polyethylene glycol-maleimide.
  • the antibody to be pegylated is an aglycosylated antibody.
  • Methods for pegylating proteins are known in the art and can be applied to the antibodies described herein. See, for example, EP0154316 by Nishimura et al. and EP0401384 by Ishikawa et al.
  • the present invention also encompasses a human monoclonal antibody described herein conjugated to a therapeutic agent, a polymer, a detectable label, or enzyme.
  • the therapeutic agent is a cytotoxic agent.
  • the polymer is PEG. 1.3.
  • the present invention provides isolated nucleic acid segments that encode the polypeptides, peptide fragments, and coupled proteins of the invention.
  • the nucleic acid segments of the invention also include segments that encode for the same amino acids due to the degeneracy of the genetic code.
  • the amino acid threonine is encoded by ACU, ACC, ACA, and ACG and is therefore degenerate. It is intended that the invention includes all variations of the polynucleotide segments that encode for the same amino acids. Such mutations are known in the art (Watson et al., Molecular Biology of the Gene, Benjamin Cummings 1987).
  • nucleic acid segments of the invention may be contained within a vector.
  • a vector may include, but is not limited to, any plasmid, phagemid, F-factor, virus, cosmid, or phage in a double- or single-stranded linear or circular form which may or may not be self transmissible or mobilizable.
  • the vector can also transform a prokaryotic or eukaryotic host either by integration into the cellular genome or exist extra-chromosomally (e.g., autonomous replicating plasmid with an origin of replication).
  • the nucleic acid segment in the vector can be under the control of, and operably linked to, an appropriate promoter or other regulatory elements for transcription in vitro or in a host cell, such as a eukaryotic cell, or a microbe, e.g., bacteria.
  • the vector may be a shuttle vector that functions in multiple hosts.
  • the vector may also be a cloning vector that typically contains one or a small number of restriction endonuclease recognition sites at which foreign DNA sequences can be inserted in a determinable fashion.
  • a cloning vector may also contain a marker gene that is suitable for use in the identification and selection of cells transformed with the cloning vector. Examples of marker genes are tetracycline resistance or ampicillin resistance. Many cloning vectors are commercially available (Stratagene, New England Biolabs, Clonetech). [00257] The nucleic acid segments of the invention may also be inserted into an expression vector.
  • an expression vector contains prokaryotic DNA elements coding for a bacterial replication origin and an antibiotic resistance gene to provide for the amplification and selection of the expression vector in a bacterial host; regulatory elements that control initiation of transcription such as a promoter; and DNA elements that control the processing of transcripts such as introns, or a transcription termination/polyadenylation sequence.
  • Methods to introduce nucleic acid segment into a vector are available in the art (Sambrook et al., Molecular Cloning: A Laboratory Manual, 3rd edition, Cold Spring Harbor Press, Cold Spring Harbor, N.Y. (2001)).
  • a vector into which a nucleic acid segment is to be inserted is treated with one or more restriction enzymes (restriction endonuclease) to produce a linearized vector having a blunt end, a ⁇ sticky ⁇ end with a 5’ or a 3’ overhang, or any combination of the above.
  • the vector may also be treated with a restriction enzyme and subsequently treated with another modifying enzyme, such as a polymerase, an exonuclease, a phosphatase or a kinase, to create a linearized vector that has characteristics useful for ligation of a nucleic acid segment into the vector.
  • the nucleic acid segment that is to be inserted into the vector is treated with one or more restriction enzymes to create a linearized segment having a blunt end, a ⁇ sticky ⁇ end with a 5’ or a 3’ overhang, or any combination of the above.
  • the nucleic acid segment may also be treated with a restriction enzyme and subsequently treated with another DNA modifying enzyme.
  • DNA modifying enzymes include, but are not limited to, polymerase, exonuclease, phosphatase or a kinase, to create a nucleic acid segment that has characteristics useful for ligation of a nucleic acid segment into the vector.
  • the treated vector and nucleic acid segment are then ligated together to form a construct containing a nucleic acid segment according to methods available in the art (Sambrook et al., Molecular Cloning: A Laboratory Manual, 3rd edition, Cold Spring Harbor Press, Cold Spring Harbor, N.Y. (2001)). Briefly, the treated nucleic acid fragment, and the treated vector are combined in the presence of a suitable buffer and ligase. The mixture is then incubated under appropriate conditions to allow the ligase to ligate the nucleic acid fragment into the vector.
  • the invention also provides an expression cassette that contains a nucleic acid sequence capable of directing expression of a particular nucleic acid segment of the invention, either in vitro or in a host cell. Also, a nucleic acid segment of the invention may be inserted into the expression cassette such that an anti-sense message is produced.
  • the expression cassette is an isolatable unit such that the expression cassette may be in linear form and functional for in vitro transcription and translation assays. The materials and procedures to conduct these assays are commercially available from Promega Corp. (Madison, Wis.).
  • an in vitro transcript may be produced by placing a nucleic acid sequence under the control of a T7 promoter and then using T7 RNA polymerase to produce an in vitro transcript.
  • the expression cassette can be incorporated into a vector allowing for replication and amplification of the expression cassette within a host cell or also in vitro transcription and translation of a nucleic acid segment.
  • Such an expression cassette may contain one or a plurality of restriction sites allowing for placement of the nucleic acid segment under the regulation of a regulatory sequence.
  • the expression cassette can also contain a termination signal operably linked to the nucleic acid segment as well as regulatory sequences required for proper translation of the nucleic acid segment.
  • the expression cassette containing the nucleic acid segment may be chimeric, meaning that at least one of its components is heterologous with respect to at least one of its other components.
  • the expression cassette may also be one that is naturally occurring but has been obtained in a recombinant form useful for heterologous expression. Expression of the nucleic acid segment in the expression cassette may be under the control of a constitutive promoter or an inducible promoter, which initiates transcription only when the host cell is exposed to some particular external stimulus.
  • the expression cassette may include in the 5’-3’ direction of transcription, a transcriptional and translational initiation region, a nucleic acid segment and a transcriptional and translational termination region functional in vivo and/or in vitro.
  • the termination region may be native with the transcriptional initiation region, may be native with the nucleic acid segment, or may be derived from another source.
  • the regulatory sequence can be a polynucleotide sequence located upstream (5' non- coding sequences), within, or downstream (3’ non-coding sequences) of a coding sequence, and which influences the transcription, RNA processing or stability, or translation of the associated coding sequence.
  • Regulatory sequences can include, but are not limited to, enhancers, promoters, repressor binding sites, translation leader sequences, introns, and polyadenylation signal sequences. They may include natural and synthetic sequences as well as sequences which may be a combination of synthetic and natural sequences.
  • a promoter is a nucleotide sequence that controls the expression of the coding sequence by providing the recognition for RNA polymerase and other factors required for proper transcription.
  • a promoter includes a minimal promoter, consisting only of all basal elements needed for transcription initiation, such as a TATA-box and/or initiator that is a short DNA sequence comprised of a TATA-box and other sequences that serve to specify the site of transcription initiation, to which regulatory elements are added for control of expression.
  • a promoter may be derived entirely from a native gene, or be composed of different elements derived from different promoters found in nature, or even be comprised of synthetic DNA segments.
  • a promoter may contain DNA sequences that are involved in the binding of protein factors that control the effectiveness of transcription initiation in response to physiological or developmental conditions.
  • the invention also provides a construct containing a vector and an expression cassette.
  • the vector may be selected from, but not limited to, any vector previously described. Into this vector may be inserted an expression cassette through methods known in the art and previously described (Sambrook et al., Molecular Cloning: A Laboratory Manual, 3rd edition, Cold Spring Harbor Press, Cold Spring Harbor, N.Y. (2001)).
  • the regulatory sequences of the expression cassette may be derived from a source other than the vector into which the expression cassette is inserted.
  • a construct containing a vector and an expression cassette is formed upon insertion of a nucleic acid segment of the invention into a vector that itself contains regulatory sequences.
  • an expression cassette is formed upon insertion of the nucleic acid segment into the vector.
  • Vectors containing regulatory sequences are available commercially, and methods for their use are known in the art (Clonetech, Promega, Stratagene).
  • vectors comprising the polynucleotides or nucleic acid molecules disclosed herein.
  • the host cell may be co-transfected with two vectors provided herein, the first vector encoding a heavy chain derived polypeptide and the second vector encoding a light chain derived polypeptide.
  • the two vectors may contain identical selectable markers which enable equal expression of heavy and light chain polypeptides.
  • a single vector may be used which encodes, and is capable of expressing, both heavy and light chain polypeptides. In such situations, the light chain should be placed before the heavy chain to avoid an excess of toxic free heavy chain (Proudfoot, Nature, 1986, 322:52; and Kohler, Proc. Natl. Acad. Sci. USA , 1980, 77:2197-9).
  • vectors include, without limitation, plasmids, phagemids, cosmids, transposons, artificial chromosomes such as yeast artificial chromosome (YAC), bacterial artificial chromosome (BAC), or P1-derived artificial chromosome (PAC), bacteriophages such as lambda phage or M13 phage, and animal viruses.
  • artificial chromosomes such as yeast artificial chromosome (YAC), bacterial artificial chromosome (BAC), or P1-derived artificial chromosome (PAC)
  • bacteriophages such as lambda phage or M13 phage
  • animal viruses such as yeast artificial chromosome (YAC), bacterial artificial chromosome (BAC), or P1-derived artificial chromosome (PAC)
  • bacteriophages such as lambda phage or M13 phage
  • animal viruses such as lambda phage or M13 phage
  • the recombinant vector comprising a polynucleotide encoding the antibody or antigen-binding fragment thereof described herein is a plasmid.
  • suitable plasmid expression vectors are known to those of skill in the art, and many are commercially available. The following vectors are provided by way of example; for eukaryotic host cells: pXT1, pSG5 (Stratagene), pSVK3, pBPV, pMSG, and pSVLSV40 (Pharmacia). However, any other plasmid vector may be used so long as it is compatible with the host cell.
  • viral vectors are used to deliver one or more polynucleotides contemplated herein.
  • Suitable viral vectors include, but are not limited to, viral vectors based on adenovirus (see, e.g., Li et al., Invest Opthalmol Vis Sci 35:25432549, 1994; Borras et al., Gene Ther 6:515524, 1999; Li and Davidson, PNAS 92:77007704, 1995; Sakamoto et al., H Gene Ther 5:10881097, 1999; WO 94/12649, WO 93/03769; WO 93/19191 ; WO 94/28938; WO 95/11984 and WO 95/00655); adeno-associated virus (see, e.g., U.S. Patent No.
  • alphaviruses alphaviruses; arenaviruses; baculovirus; herpes simplex virus; human immunodeficiency virus (see, e.g., Miyoshi et al., PNAS 94:10319 23, 1997; Takahashi et al., J Virol 73:7812 7816, 1999); poliovirus; poxvirus; retrovirus (e.g., Murine Leukemia Virus, spleen necrosis virus, and vectors derived from retroviruses such as Rous Sarcoma Virus, Harvey Sarcoma Virus, avian leukosis virus, a lentivirus, human immunodeficiency virus, myeloproliferative sarcoma virus, and mammary tumor virus); SV40; vaccinia virus; and the like.
  • retrovirus e.g., Murine Leukemia Virus, spleen necrosis virus, and vectors derived from retroviruses such as Rous Sarcoma
  • vectors are pClneo vectors (Promega) for expression in mammalian cells; pLenti4/V5-DESTTM, pLenti6/V5-DESTTM, and pLenti6.2/V5-GW/lacZ (Invitrogen) for lentivirus-mediated gene transfer and expression in mammalian cells.
  • the vector is a non-integrating vector, including but not limited to, an episomal vector or a vector that is maintained extrachromosomally.
  • the term “episomal” refers to a vector that is able to replicate without integration into host’s chromosomal DNA and without gradual loss from a dividing host cell also meaning that said vector replicates extrachromosomally or episomally.
  • the vector is engineered to harbor the sequence coding for the origin of DNA replication or “ori” from a lymphotrophic herpes virus or a gamma herpesvirus, an adenovirus, SV40, a bovine papilloma virus, or a yeast, specifically a replication origin of a lymphotrophic herpes virus or a gamma herpesvirus corresponding to oriP of EBV.
  • the lymphotrophic herpes virus may be Epstein Barr virus (EBV), Kaposi’s sarcoma herpes virus (KSHV), Herpes virus saimiri (HS), or Marek’s disease virus (MDV).
  • Epstein Barr virus (EBV) and Kaposi’s sarcoma herpes virus (KSHV) are also examples of a gamma herpesvirus.
  • a viral vector delivered by such viruses or viral particles may be referred to by the type of virus to deliver the viral vector (e.g., a lentiviral vector is a viral vector that is to be delivered by a lentivirus).
  • a viral vector can contain viral elements (e.g., nucleotide sequences) necessary for packaging of the viral vector into the virus or viral particle, replicating the virus, or other desired viral activities.
  • a virus containing a viral vector may be replication competent, replication deficient or replication defective.
  • the vector is an integrating vector.
  • a polynucleotide is introduced into a target or host cell using a transposon vector system.
  • the transposon vector system comprises a vector comprising transposable elements and a polynucleotide contemplated herein; and a transposase.
  • the transposon vector system is a single transposase vector system, see, e.g., WO 2008/027384.
  • exemplary transposases include, but are not limited to: piggyBac, Sleeping Beauty, Mos1, Tc1/mariner, Tol2, mini-Tol2, Tc3, MuA, Himar I, Frog Prince, and derivatives thereof.
  • the piggyBac transposon and transposase are described, for example, in U.S. Patent 6,962,810, which is incorporated herein by reference in its entirety.
  • the Sleeping Beauty transposon and transposase are described, for example, in Izsvak et al., J. Mol.
  • the Tol2 transposon which was first isolated from the medaka fish Oryzias latipes and belongs to the hAT family of transposons is described in Kawakami et al. (2000).
  • Mini-Tol2 is a variant of Tol2 and is described in Balciunas et al. (2006).
  • the Tol2 and Mini-Tol2 transposons facilitate integration of a transgene into the genome of an organism when co-acting with the Tol2 transposase.
  • the Frog Prince transposon and transposase are described, for example, in Miskey et al., Nucleic Acids Res.31:6873-6881 (2003).
  • a polynucleotide sequence encoding the antibody or antigen- binding fragment thereof disclosed herein is operably linked to one or more control elements that allow expression of the polynucleotide in both prokaryotic and eukaryotic cells.
  • Control elements refer those non-translated regions of the vector which interact with host cellular proteins to carry out transcription and translation.
  • Non-limiting examples of control elements include origin of replication, selection cassettes, constitutive and inducible promoters, enhancers, translation initiation signals (Shine Dalgarno sequence or Kozak sequence) introns, transcription terminators, 5’ and 3’ untranslated regions. See e.g., Bitter et al.
  • the transcriptional control element may be functional in either a eukaryotic cell (e.g., a mammalian cell) or a prokaryotic cell (e.g., bacterial or archaeal cell).
  • a eukaryotic cell e.g., a mammalian cell
  • a prokaryotic cell e.g., bacterial or archaeal cell.
  • polynucleotides encoding the antibody or antigen-binding fragment thereof described herein are operably linked to a promoter and/or an enhancer.
  • promoter refers to a recognition site of a polynucleotide (DNA or RNA) to which an RNA polymerase binds.
  • RNA polymerase initiates and transcribes polynucleotides operably linked to the promoter.
  • promoters operative in mammalian cells comprise an AT-rich region located approximately 25 to 30 bases upstream from the site where transcription is initiated and/or another sequence found 70 to 80 bases upstream from the start of transcription, a CNCAAT region where N may be any nucleotide.
  • the term “enhancer” refers to a segment of DNA which contains sequences capable of providing enhanced transcription and in some instances can function independent of their orientation relative to another control sequence.
  • An enhancer can function cooperatively or additively with promoters and/or other enhancer elements.
  • Non-limiting examples of suitable eukaryotic promoters include those from cytomegalovirus (CMV) immediate early, herpes simplex virus (HSV) thymidine kinase, a viral simian virus 40 (SV40) (e.g., early and late SV40), a spleen focus forming virus (SFFV) promoter, long terminal repeats (LTRs) from retrovirus (e.g., a Moloney murine leukemia virus (MoMLV) LTR.
  • CMV cytomegalovirus
  • HSV herpes simplex virus
  • SFFV simian virus 40
  • LTRs long terminal repeats
  • retrovirus e.g., a Moloney murine leukemia virus (MoMLV) LTR.
  • HSV Rous sarcoma virus
  • H5 Rous sarcoma virus
  • HSV herpes simplex virus
  • H5 herpes simplex virus
  • H5 herpes simplex virus
  • EF1 ⁇ elongation factor 1 -alpha
  • EGR1 early growth response 1
  • FerH ferritin H
  • FerL ferritin L
  • GPDH Glyceraldehyde 3 -phosphate dehydrogenase
  • GPDH Glyceraldehyde 3 -phosphate dehydrogenase
  • EIF4A1 eukaryotic translation initiation factor 4A1
  • HSPA5 heat shock 70kDa protein 5
  • HSPA5 heat shock protein 90kDa beta
  • member 1 HP90B1 promoter
  • HSP70 heat shock protein 70kDa
  • p- KIN p-kinesin
  • a polynucleotide sequence encoding the antibody or antigenbinding fragment thereof described herein is operably linked to a constitutive promoter.
  • the polynucleotides encoding the antibody or antigen-binding fragment thereof described herein are constitutively and/or ubiquitously expressed in a cell.
  • a polynucleotide sequence encoding the antibody or antigenbinding fragment thereof described herein is operably linked to an inducible promoter.
  • polynucleotides encoding the antibody or antigen- binding fragment thereof described herein are conditionally expressed.
  • conditional expression may refer to any type of conditional expression including, but not limited to, inducible expression; repressible expression; expression in cells or tissues having a particular physiological, biological, or disease state (e.g., cell type or tissue specific expression) etc.
  • inducible promoters/systems include, but are not limited to, steroid-inducible promoters such as promoters for genes encoding glucocorticoid or estrogen receptors (inducible by treatment with the corresponding hormone), metallothionine promoter (inducible by treatment with various heavy metals), MX-I promoter (inducible by interferon), the “GeneSwitch” mifepristone-regulatable system (Sirin et al., 2003, Gene, 323:67), the cumate inducible gene switch (WO 2002/088346), tetracycline-dependent regulatory systems, etc.
  • steroid-inducible promoters such as promoters for genes encoding glucocorticoid or estrogen receptors (inducible by treatment with the corresponding hormone), metallothionine promoter (inducible by treatment with various heavy metals), MX-I promoter (inducible by interferon), the “GeneSwitch”
  • the vectors described herein further comprise a transcription termination signal. Elements directing the efficient termination and polyadenylation of the heterologous nucleic acid transcripts increases heterologous gene expression. Transcription termination signals are generally found downstream of the polyadenylation signal.
  • vectors comprise a polyadenylation sequence 3’ of a polynucleotide encoding a polypeptide to be expressed.
  • poly A site or “poly A sequence” as used herein denotes a DNA sequence which directs both the termination and polyadenylation of the nascent RNA transcript by RNA polymerase II.
  • Polyadenylation sequences can promote mRNA stability' by addition of a poly A tail to the 3’ end of the coding sequence and thus, contribute to increased translational efficiency.
  • Cleavage and polyadenylation is directed by a poly(A) sequence in the RNA.
  • the core poly(A) sequence for mammalian pre-mRNAs has two recognition elements flanking a cleavage-polyadenylation site.
  • an almost invariant AAUAAA hexamer lies 20-50 nucleotides upstream of a more variable element rich in U or GU residues.
  • Cleavage of the nascent transcript occurs between these two elements and is coupled to the addition of up to 250 adenosines to the 5’ cleavage product.
  • the core poly(A) sequence is an ideal polyA sequence (e.g., AATAAA, ATTAAA, AGTAAA).
  • the poly(A) sequence is an SV40 polyA sequence, a bovine growth hormone polyA sequence (BGHpA), a rabbit P-globin polyA sequence (rPgpA), variants thereof, or another suitable heterologous or endogenous polyA sequence known in the art.
  • the expression vector may also include nucleotide sequences encoding protein tags (e.g., 6xHis tag, hemagglutinin tag, green fluorescent protein, etc.) that are fused to the site-directed modifying polypeptide, thus resulting in a chimeric polypeptide.
  • Methods of introducing polynucleotides and recombinant vectors into a host cell are known in the art. Suitable methods include e.g., viral or bacteriophage infection, transfection, conjugation, protoplast fusion, lipofection, electroporation, calcium phosphate precipitation, polyethyleneimine (PEI)-mediated transfection, DEAE-dextran mediated transfection, liposome- mediated transfection, particle gun technology, calcium phosphate precipitation, direct micro injection, nanoparticle-mediated nucleic acid delivery (see, e.g., Panyam et al., Adv Drug Deliv Rev. 2012 Sep 13.
  • PKI polyethyleneimine
  • delivery via electroporation comprises mixing the cells with the polynucleotides encoding the antibody or antigen-binding fragment thereof in a cartridge, chamber, or cuvette and applying one or more electrical impulses of defined duration and amplitude.
  • cells are mixed with polynucleotides encoding the antibody or antigen-binding fragment thereof in a vessel connected to a device (e.g., a pump) which feeds the mixture into a cartridge, chamber, or cuvette wherein one or more electrical impulses of defined duration and amplitude are applied, after which the cells are delivered to a second vessel.
  • a device e.g., a pump
  • polynucleotide delivery systems suitable for use in particular embodiments contemplated include, but are not limited to, those provided by Amaxa Biosystems, Maxcyte, Inc., BTX Molecular Delivery Systems, Neon® Transfection Systems, and Copernicus Therapeutics Inc.
  • Lipofection reagents are sold commercially (e.g., TransfectamTM and LipofectinTM). Cationic and neutral lipids that are suitable for efficient lipofection of polynucleotides have been described in the literature. See e.g., Liu et al. (2003) Gene Therapy. 10:180–187; and Balazs et al. (2011) Journal of Drug Delivery.2011:1-12.
  • polynucleotides encoding the antibody or antigen-binding fragment thereof described herein are introduced to a cell in a non-viral delivery vehicle, such as a transposon, a nanoparticle (e.g., a lipid nanoparticle), a liposome, an exosome, an attenuated bacterium, or a virus-like particle.
  • a non-viral delivery vehicle such as a transposon, a nanoparticle (e.g., a lipid nanoparticle), a liposome, an exosome, an attenuated bacterium, or a virus-like particle.
  • the vehicle is an attenuated bacterium (e.g., naturally or artificially engineered to be invasive but attenuated to prevent pathogenesis including Listeria monocytogenes, certain Salmonella strains, Bifidobacterium longum, and modified Escherichia coli), bacteria having nutritional and tissue-specific tropism to target specific cells, and bacteria having modified surface proteins to alter target cell specificity.
  • the vehicle is a genetically modified bacteriophage (e.g., engineered phages having large packaging capacity, less immunogenicity, containing mammalian plasmid maintenance sequences and having incorporated targeting ligands).
  • the vehicle is a biological liposome.
  • the biological liposome is a phospholipid-based particle derived from human cells (e.g., erythrocyte ghosts, which are red blood cells broken down into spherical structures derived from the subject and wherein tissue targeting can be achieved by attachment of various tissue or cell-specific ligands), secretory exosomes, or subject-derived membrane-bound nanovesicles (30-100 nm) of endocytic origin (e.g., can be produced from various cell types and can therefore be taken up by cells without the need for targeting ligands).
  • human cells e.g., erythrocyte ghosts, which are red blood cells broken down into spherical structures derived from the subject and wherein tissue targeting can be achieved by attachment of various tissue or cell-specific ligands), secretory exosomes, or subject-derived membrane-bound nanovesicles (30-100 nm) of endocytic origin (e.g., can be produced from various cell types and can therefore be taken up by cells without
  • vectors comprising polynucleotides encoding the antibody or antigen-binding fragment thereof described herein are introduced to cells by viral delivery methods, e.g., by viral transduction.
  • viral delivery methods e.g., by viral transduction.
  • retroviruses provide a convenient platform for gene delivery systems.
  • the heterologous nucleic acid can be inserted into a vector and packaged in retroviral particles using techniques known in the art.
  • the recombinant virus can then be isolated and delivered to the engineered mammalian cell in vitro or ex vivo.
  • retroviral systems are known in the art.
  • adenovirus vectors are used.
  • adenovirus vectors are known in the art.
  • lentivirus vectors are used.
  • self-inactivating lentiviral vectors are used.
  • self-inactivating lentiviral vectors carrying the immunomodulator (such as immune checkpoint inhibitor) coding sequence and/or self-inactivating lentiviral vectors carrying chimeric antigen receptors can be packaged with protocols known in the art.
  • the resulting lentiviral vectors can be used to transduce a mammalian cell (such as primary human T cells) using methods known in the art.
  • Vectors derived from retroviruses such as lentivirus are suitable tools to achieve long-term gene transfer, because they allow long-term, stable integration of a transgene and its propagation in progeny cells. Lentiviral vectors also have low immunogenicity, and can transduce nonproliferating cells.
  • the vehicle is a mammalian virus-like particle.
  • modified viral particles can be generated (e.g., by purification of the “empty” particles followed by ex vivo assembly of the virus with the desired cargo).
  • this disclosure also provides (i) a nucleic acid molecule encoding a polypeptide chain of the antibody or antigen-binding fragment thereof described herein; (ii) a vector comprising the nucleic acid molecule as described; and (iii) a cultured host cell comprising the vector as described. Also provided is a method for producing a polypeptide, comprising: (a) obtaining the cultured host cell as described; (b) culturing the cultured host cell in a medium under conditions permitting expression of a polypeptide encoded by the vector and assembling of an antibody or fragment thereof; and (c) purifying the antibody or fragment from the cultured cell or the medium of the cell.
  • Polynucleotides disclosed herein can be at least about 5, at least about 10, at least about 15, at least about 20, at least about 25, at least about 30, at least about 40, at least about 50, at least about 100, at least about 200, at least about 300, at least about 400, at least about 500, at least about 1000, at least about 5000, at least about 10000, or at least about 15000 or more nucleotides in length, as well as all intermediate lengths.
  • intermediate lengths means any length between the quoted values, such as 6, 7, 8, 9, etc., 101, 102, 103, etc.; 151, 152, 153, etc.; 201, 202, 203, etc.
  • the present disclosure further relates to variants of the polynucleotides disclosed herein.
  • the polynucleotide variants can contain alterations in the coding regions, non-coding regions, or both.
  • a polynucleotide variant comprising a nucleotide sequence that is at least about 75 %, about 80 %, about 85 %, about 90 %, about 91 %, about 92 %, about 93 %, about 94 %, about 95 %, about 96 %, about 97 %, about 98%, or about 99 % identical to the nucleotide sequence of a polynucleotide disclosed herein.
  • a polynucleotide variant contains substitutions, additions, or deletions that alter the properties or activities of the encoded polypeptide.
  • a polynucleotide variant contains silent substitutions, additions, or deletions that does not alter the properties or activities of the encoded polypeptide.
  • a polynucleotide variant is produced to modulate or alter expression (or expression levels) of the encoded polypeptide.
  • a polynucleotide variant is produced to increase expression of the encoded polypeptide.
  • a polynucleotide variant is produced to decrease expression of the encoded polypeptide.
  • a polynucleotide variant has increased expression of the encoded polypeptide as compared to a parental polynucleotide sequence.
  • polynucleotide variant has decreased expression of the encoded polypeptide as compared to a parental polynucleotide sequence.
  • polynucleotides are codon-optimized.
  • the term “codon-optimized” refers to substituting codons in a polynucleotide encoding a polypeptide in order to increase the expression, stability and/or activity of the polypeptide.
  • Factors that influence codon optimization include, but are not limited to one or more of: (i) variation of codon biases between two or more organisms or genes or synthetically constructed bias tables, (ii) variation in the degree of codon bias within an organism, gene, or set of genes, (iii) systematic variation of codons including context, (iv) variation of codons according to their decoding tRNAs, (v) variation of codons according to GC %, either overall or in one position of the triplet, (vi) variation in degree of similarity to a reference sequence for example a naturally occurring sequence, (vii) variation in the codon frequency cutoff, (viii) structural properties of mRNAs transcribed from the DNA sequence, (ix) prior knowledge about the function of the DNA sequences upon which design of the codon substitution set is to be based, (x) systematic variation of codon sets for each amino acid, (xi) isolated removal of spurious translation initiation sites and/or (xii) elimination of fortuitous polyadeny
  • nucleotide sequences that encode a polypeptide, or fragment of variant thereof, as described herein. Some of these polynucleotides bear minimal homology to the nucleotide sequence of any native gene. Nonetheless, polynucleotides that vary due to differences in codon usage are specifically contemplated in particular embodiments, for example polynucleotides that are optimized for human and/or primate codon selection. Further, alleles of the genes comprising the polynucleotide sequences provided herein may also be used.
  • alleles are endogenous genes that are altered as a result of one or more mutations, such as deletions, additions and/or substitutions of nucleotides.
  • the polynucleotides contemplated herein may be combined with other DNA sequences, such as promoters and/or enhancers, untranslated regions (UTRs), signal sequences, Kozak sequences, polyadenylation signals, additional restriction enzyme sites, multiple cloning sites, internal ribosomal entry sites (IRES), recombinase recognition sites (e.g., LoxP, FRT, and Att sites), termination codons, transcriptional termination signals, and polynucleotides encoding self-cleaving polypeptides, epitope tags, as disclosed elsewhere herein or as known in the art, such that their overall length may vary considerably.
  • promoters and/or enhancers such as promoters and/or enhancers, untranslated regions (UTRs), signal sequences, Kozak sequences, polya
  • polynucleotide fragment of almost any length may be employed in particular embodiments, with the total length preferably being limited by the ease of preparation and use in the intended recombinant DNA protocol.
  • Polynucleotides can be prepared, isolated, purified, manipulated, and/or expressed using any of a variety of well-established techniques known and available in the art. 1.4. Methods of Production [00295] Antibodies may be produced using recombinant methods and compositions, e.g., as described in U.S. Pat. No. 4,816,567. In one embodiment, an isolated nucleic acid encoding an antibody described herein is provided.
  • nucleic acid may encode an amino acid sequence comprising the VL and/or an amino acid sequence comprising the VH of the antibody (e.g., the light and/or heavy chains of the antibody).
  • one or more vectors comprising such nucleic acid are provided.
  • a host cell comprising such nucleic acid is provided.
  • a host cell comprises (e.g., has been transformed with): (1) a vector comprising a nucleic acid that encodes an amino acid sequence comprising the VL of the antibody and an amino acid sequence comprising the VH of the antibody, or (2) a first vector comprising a nucleic acid that encodes an amino acid sequence comprising the VL of the antibody and a second vector comprising a nucleic acid that encodes an amino acid sequence comprising the VH of the antibody.
  • the host cell is eukaryotic, e.g., a Chinese Hamster Ovary (CHO) cell or lymphoid cell (e.g., Y0, NS0, Sp20 cell).
  • a method of making an antibody comprises culturing a host cell comprising a nucleic acid encoding the antibody, as provided above, under conditions suitable for expression of the antibody, and optionally recovering the antibody from the host cell (or host cell culture medium).
  • a nucleic acid encoding an antibody e.g., as described herein, is isolated and inserted into one or more vectors for further cloning and/or expression in a host cell.
  • Such nucleic acid may be readily isolated and sequenced using conventional procedures (e.g., by using oligonucleotide probes that are capable of binding specifically to genes encoding the heavy and light chains of the antibody).
  • Suitable host cells for cloning or expression of antibody-encoding vectors include prokaryotic or eukaryotic cells described herein.
  • antibodies may be produced in bacteria, in particular when glycosylation and Fc effector function are not needed.
  • For expression of antibody fragments and polypeptides in bacteria see, e.g., U.S. Pat. Nos.5,648,237, 5,789,199, and 5,840,523. (See also Charlton, Methods in Molecular Biology, Vol. 248 (B.K.C. Lo, ed., Humana Press, Totowa, N.J., 2003), pp. 245-254, describing expression of antibody fragments in E.
  • the antibody may be isolated from the bacterial cell paste in a soluble fraction and can be further purified.
  • eukaryotic microbes such as filamentous fungi or yeast are suitable cloning or expression hosts for antibody-encoding vectors, including fungi and yeast strains whose glycosylation pathways have been “humanized,” resulting in the production of an antibody with a partially or fully human glycosylation pattern. See Gerngross, Nat. Biotech. 22:1409-1414 (2004), and Li et al., Nat. Biotech. 24:210-215 (2006).
  • Suitable host cells for the expression of glycosylated antibodies are also derived from multicellular organisms (invertebrates and vertebrates). Examples of invertebrate cells include plant and insect cells. Numerous baculoviral strains have been identified, which may be used in conjunction with insect cells, particularly for transfection of Spodoptera frugiperda cells. [00300] Plant cell cultures can also be utilized as hosts. See, e.g., U.S. Pat. Nos. 5,959,177, 6,040,498, 6,420,548, 7,125,978, and 6,417,429 (describing PLANTIBODIES technology for producing antibodies in transgenic plants). [00301] Vertebrate cells may also be used as hosts.
  • mammalian cell lines that are adapted to grow in suspension may be useful.
  • useful mammalian host cell lines are monkey kidney CV1 line transformed by SV40 (COS-7); human embryonic kidney line (293 or 293 cells as described, e.g., in Graham et al., J. Gen Virol. 36:59 (1977)); baby hamster kidney cells (BHK); mouse sertoli cells (TM4 cells as described, e.g., in Mather, Biol. Reprod.
  • monkey kidney cells (CV1); African green monkey kidney cells (VERO-76); human cervical carcinoma cells (HELA); canine kidney cells (MDCK; buffalo rat liver cells (BRL 3A); human lung cells (W138); human liver cells (Hep G2); mouse mammary tumor (MMT 060562); TRI cells, as described, e.g., in Mather et al., Annals N.Y. Acad. Sci. 383:44-68 (1982); MRC 5 cells; and FS4 cells.
  • Other useful mammalian host cell lines include CHO cells, including DHFR- CHO cells (Urlaub et al., Proc. Natl. Acad. Sci.
  • lymphocytes In the hybridoma method (see, e.g., described in Kohler, et al., Nature, 1975, 256:495- 7), a mouse or other appropriate host animal, such as a hamster, is immunized as described above to elicit lymphocytes that produce or are capable of producing antibodies that will specifically bind to the protein used for immunization. Alternatively, lymphocytes may be immunized in vitro. After immunization, lymphocytes are isolated and then fused with a myeloma cell line using a suitable fusing agent, such as polyethylene glycol, to form a hybridoma cell (Goding, Monoclonal Antibodies: Principles and Practice 59-103 (1986)).
  • a suitable fusing agent such as polyethylene glycol
  • the hybridoma cells thus prepared are seeded and grown in a suitable culture medium, which, in some embodiments, contains one or more substances that inhibit the growth or survival of the unfused, parental myeloma cells (also referred to as fusion partner).
  • a suitable culture medium which, in some embodiments, contains one or more substances that inhibit the growth or survival of the unfused, parental myeloma cells (also referred to as fusion partner).
  • the parental myeloma cells lack the enzyme hypoxanthine guanine phosphoribosyl transferase (HGPRT or HPRT)
  • HGPRT hypoxanthine guanine phosphoribosyl transferase
  • HGPRT hypoxanthine guanine phosphoribosyl transferase
  • HAT medium thymidine
  • Exemplary parental myeloma cells are those that fuse efficiently, support stable high- level production of antibody by the selected antibody-producing cells, and are sensitive to a selective medium that selects against the unfused parental cells.
  • Exemplary myeloma cell lines are murine myeloma lines, such as SP-2 and derivatives, for example, X63-Ag8-653 cells available from the American Type Culture Collection (Manassas, VA), and those derived from MOPC-21 and MPC-11 mouse tumors available from the Salk Institute Cell Distribution Center (San Diego, CA).
  • Human myeloma and mouse-human heteromyeloma cell lines also have been described for the production of human monoclonal antibodies (Kozbor, Immunol.
  • Culture medium in which hybridoma cells are growing is assayed for production of monoclonal antibodies directed against the antigen.
  • the binding specificity of monoclonal antibodies produced by hybridoma cells is determined by immunoprecipitation or by an in vitro binding assay, such as RIA or ELISA.
  • the binding affinity of the monoclonal antibody can, for example, be determined by the Scatchard analysis described in Munson et al., Anal. Biochem., 1980, 107:220-39.
  • the clones may be subcloned by limiting dilution procedures and grown by standard methods (Goding, supra). Suitable culture media for this purpose include, for example, DMEM or RPMI-1640 medium.
  • the hybridoma cells may be grown in vivo as ascites tumors in an animal, for example, by i.p. injection of the cells into mice.
  • the monoclonal antibodies secreted by the subclones are suitably separated from the culture medium, ascites fluid, or serum by conventional antibody purification procedures such as, for example, affinity chromatography (e.g., using protein A or protein G-Sepharose) or ion- exchange chromatography, hydroxyapatite chromatography, gel electrophoresis, dialysis, etc.
  • affinity chromatography e.g., using protein A or protein G-Sepharose
  • ion- exchange chromatography e.g., using protein A or protein G-Sepharose
  • ion- exchange chromatography e.g., using protein A or protein G-Sepharose
  • hydroxyapatite chromatography hydroxyapatite chromatography
  • gel electrophoresis hydroxyapatite chromatography
  • dialysis etc.
  • synthetic antibody clones are selected by screening phage libraries containing phages that display various fragments of antibody variable region (Fv) fused
  • Variable domains can be displayed functionally on phage, either as single-chain Fv (scFv) fragments, in which VH and VL are covalently linked through a short, flexible peptide, or as Fab fragments, in which they are each fused to a constant domain and interact non-covalently, as described, for example, in Winter et al., 1994, Ann. Rev. Immunol.12:433-55.
  • scFv single-chain Fv
  • Repertoires of VH and VL genes can be separately cloned by PCR and recombined randomly in phage libraries, which can then be searched for antigen-binding clones as described in Winter et al, supra.
  • Libraries from immunized sources provide high-affinity antibodies to the antigen without the requirement of constructing hybridomas.
  • naive libraries can be cloned to provide a single source of human antibodies to a wide range of non-self and self-antigens without any immunization as described by Griffiths et al., EMBO J, 1993, 12:725-34.
  • naive libraries can also be made synthetically by cloning the unrearranged V-gene segments from stem cells, and using PCR primers containing random sequence to encode the highly variable CDR3 regions and to accomplish rearrangement in vitro as described, for example, by Hoogenboom and Winter, J. Mol. Biol., 1992, 227:381-88. [00312] Screening of the libraries can be accomplished by various techniques known in the art. For example, the antigen can be used to coat the wells of adsorption plates, expressed on host cells affixed to adsorption plates or used in cell sorting, conjugated to biotin for capture with streptavidin-coated beads, or used in any other method for panning display libraries.
  • phage display methods that can be used herein include those disclosed in Antibody Phage Display: Methods and Protocols (O’Brien and Aitken, eds., 2002); Brinkman, et al, J. Immunol.
  • DNA encoding the monoclonal antibodies is readily isolated from the hybridoma cells and the screened libraries. Such DNA can be sequenced using conventional procedures (e.g., by using oligonucleotide probes that are capable of binding specifically to genes encoding the heavy and light chains of murine antibodies). Once isolated, the DNA may be placed into expression vectors, which are then transfected into host cells, such as E.
  • an antibody molecule provided herein may be purified by any method known in the art for purification of an immunoglobulin molecule, for example, by chromatography (e.g., ion exchange, affinity, particularly by affinity for the specific antigen after Protein A, and sizing column chromatography), centrifugation, differential solubility, or by any other standard technique for the purification of proteins.
  • chromatography e.g., ion exchange, affinity, particularly by affinity for the specific antigen after Protein A, and sizing column chromatography
  • centrifugation e.g., ion exchange, affinity, particularly by affinity for the specific antigen after Protein A, and sizing column chromatography
  • differential solubility e.g., differential solubility, or by any other standard technique for the purification of proteins.
  • the antibodies provided herein can be fused to heterologous polypeptide sequences described herein or otherwise known in the art to facilitate purification.
  • the antibodies disclosed herein can comprise “chimeric” sequences in which a portion of the heavy and/or light chain is identical with or homologous to corresponding sequences in antibodies derived from a particular species or belonging to a particular antibody class or subclass, while the remainder of the chain(s) is identical with or homologous to corresponding sequences in antibodies derived from another species or belonging to another antibody class or subclass, so long as they exhibit the desired biological activity (see U S. Pat. No. 4,816,567; and Morrison, et al. , Proc. Natl. Acad. Sci. USA , 1984, 81:6851-55).
  • the antibodies disclosed herein can be humanized antibodies.
  • a humanized antibody can comprise human framework region and human constant region sequences. In some embodiments, one or more FR region residues of the human immunoglobulin are replaced by corresponding nonhuman residues. In some embodiments, humanized antibodies comprise residues that are not found in the recipient antibody or in the donor antibody. These modifications are made to further refine antibody performance. [00318] Humanized antibodies can be produced using a variety of techniques known in the art, including but not limited to, CDR-grafting (European Patent No. EP 239,400; International publication No. WO 91/09967; and U S. Patent Nos. 5,225,539, 5,530,101, and 5,585,089), veneering or resurfacing (European Patent Nos.
  • the humanized antibodies are constructed by CDR grafting, in which the amino acid sequences of the six CDRs of the parent non-human antibody (e.g., rodent) are grafted onto a human antibody framework.
  • CDR grafting in which the amino acid sequences of the six CDRs of the parent non-human antibody (e.g., rodent) are grafted onto a human antibody framework.
  • Padlan, et al. determined that only about one third of the residues in the CDRs actually contact the antigen, and termed these the “specificity determining residues,” or SDRs (Padlan, et al, FASEB J., 1995, 9: 133-9).
  • SDR grafting only the SDR residues are grafted onto the human antibody framework (see, e.g., Kashmiri, et al, Methods, 2005, 36:25-34).
  • variable domains both light and heavy
  • sequence of the variable domain of a non-human (e.g., rodent) antibody is screened against the entire library of known human variable-domain sequences.
  • the human sequence that is closest to that of the rodent may be selected as the human framework for the humanized antibody (Sims et al., J. Immunol., 1993, 151:2296-308; and Chothia et al., J. Mol. Biol., 1987, 196:901-17).
  • Another method uses a particular framework derived from the consensus sequence of all human antibodies of a particular subgroup of light or heavy chains.
  • the same framework may be used for several different humanized antibodies (Carter et al., Proc. Natl. Acad. Sci. USA, 1992, 89:4285-89; and Presta et al., J Immunol., 1993, 151:2623-32).
  • the framework is derived from the consensus sequences of the most abundant human subclasses, VL6 subgroup I (VL6I) and VH subgroup III (VHIII).
  • VL6I VL6 subgroup I
  • VHIII VH subgroup III
  • the method consists of comparison of the non-human sequence with the functional human germline gene repertoire. Those genes encoding the same or closely related canonical structures to the murine sequences are then selected. Next, within the genes sharing the canonical structures with the non-human antibody, those with highest homology within the CDRs are chosen as FR donors. Finally, the non-human CDRs are grafted onto these FRs (see, e.g., Tan et al., J. Immunol., 2002, 169:1119-25).
  • Another method for antibody humanization is based on a metric of antibody humanness termed Human String Content (HSC). This method compares the mouse sequence with the repertoire of human germline genes, and the differences are scored as HSC.
  • HSC Human String Content
  • the target sequence is then humanized by maximizing its HSC rather than using a global identity measure to generate multiple diverse humanized variants (Lazar et al.,MoI. Immunol., 2007, 44:1986-98).
  • empirical methods may be used to generate and select humanized antibodies. These methods include those that are based upon the generation of large libraries of humanized variants and selection of the best clones using enrichment technologies or high throughput screening techniques.
  • Antibody variants may be isolated from phage, ribosome, and yeast display libraries as well as by bacterial colony screening (see, e.g., Hoogenboom, Nat.
  • FR shuffling whole FRs are combined with the non-human CDRs instead of creating combinatorial libraries of selected residue variants (see, e.g., DalTAcqua et al., Methods, 2005, 36:43-60).
  • the libraries may be screened for binding in a two-step process, first humanizing VL, followed by VH. Alternatively, a one-step FR shuffling process may be used.
  • the “human engineering” method involves altering a non-human antibody or antibody fragment, such as a mouse or chimeric antibody or antibody fragment, by making specific changes to the amino acid sequence of the antibody so as to produce a modified antibody with reduced immunogenicity in a human that nonetheless retains the desirable binding properties of the original non-human antibodies.
  • the technique involves classifying amino acid residues of a non-human (e.g., mouse) antibody as “low risk,” “moderate risk,” or “high risk” residues. The classification is performed using a global risk/reward calculation that evaluates the predicted benefits of making particular substitution (e.g., for immunogenicity in humans) against the risk that the substitution will affect the resulting antibody’s folding.
  • the particular human amino acid residue to be substituted at a given position (e.g., low or moderate risk) of a non-human (e.g., mouse) antibody sequence can be selected by aligning an amino acid sequence from the non- human antibody’s variable regions with the corresponding region of a specific or consensus human antibody sequence.
  • the amino acid residues at low or moderate risk positions in the non-human sequence can be substituted for the corresponding residues in the human antibody sequence according to the alignment.
  • the antibodies disclosed herein can be composite human antibodies.
  • a composite human antibody can be generated using, for example, Composite Human AntibodyTM technology (Antitope Ltd., Cambridge, United Kingdom).
  • variable region sequences are designed from fragments of multiple human antibody variable region sequences in a manner that avoids T cell epitopes, thereby minimizing the immunogenicity of the resulting antibody.
  • Such antibodies can comprise human constant region sequences, e.g., human light chain and/or heavy chain constant regions.
  • the antibodies disclosed herein can be deimmunized antibodies.
  • a deimmunized antibody is an antibody in which T-cell epitopes have been removed. Methods for making deimmunized antibodies have been described (see, e.g., Jones, et al., Methods Mol Biol., 2009, 525:405-23; and De Groot, et al., Cell. Immunol., 2006, 244:148-153).
  • Deimmunized antibodies comprise T-cell epitope-depleted variable regions and human constant regions. Briefly, VH and VL of an antibody are cloned and T-cell epitopes are subsequently identified by testing overlapping peptides derived from the VH and VL of the antibody in a T cell proliferation assay.
  • T cell epitopes are identified via in silico methods to identify peptide binding to human MHC class II. Mutations are introduced in the VH and VL to abrogate binding to human MHC class II. Mutated VH and VL are then utilized to generate the deimmunized antibody.
  • antibodies be humanized with retention of their affinity for the antigen and other favorable biological properties.
  • humanized antibodies are prepared by a process of analysis of the parental sequences and various conceptual humanized products using three-dimensional models of the parental and humanized sequences. Three-dimensional immunoglobulin models are commonly available and are familiar to those skilled in the art.
  • Computer programs are available which illustrate and display probable three-dimensional conformational structures of selected candidate immunoglobulin sequences. These include, for example, WAM (Whitelegg and Rees, Protein Eng., 2000, 13:819-24), Modeller (Sali and Blundell, J. Mol. Biol., 1993, 234:779-815), and Swiss PDB Viewer (Guex and Peitsch, Electrophoresis, 1997, 18:2714-23). Inspection of these displays permits analysis of the likely role of the residues in the functioning of the candidate immunoglobulin sequence, e.g., the analysis of residues that influence the ability of the candidate immunoglobulin to bind its antigen.
  • WAM Whitelegg and Rees, Protein Eng., 2000, 13:819-24
  • Modeller Sali and Blundell, J. Mol. Biol., 1993, 234:779-815
  • Swiss PDB Viewer Guiex and Peitsch, Electrophoresis, 1997, 18:2714-23. Inspection of these displays permits analysis of
  • the antibodies disclosed herein can be fully human antibodies, which possesses an amino acid sequence corresponding to that of an antibody produced by a human. This definition of a human antibody specifically excludes a humanized antibody comprising non-human antigen- binding residues. Human antibodies can be produced using various techniques known in the art, including phage-display libraries (Hoogenboom and Winter, J. Mol. Biol, 1991, 227:381; Marks, et al., 1991, J. Mol.
  • Human antibodies can also be prepared by administering the antigen to a transgenic animal that has been modified to produce such antibodies in response to antigenic challenge, but whose endogenous loci have been disabled, e.g., mice (see, e.g., Jakobovits, Curr. Opin. Biotechnol., 1995, 6(5):561-66; Bruggemann and Taussing, Curr. Opin. Biotechnol., 1997, 8(4):455-58; and U.S. Pat. Nos. 6,075,181 and 6,150,584 regarding XENOMOUSETM technology). See also, for example, Li, et al., Proc. Natl. Acad. Sci.
  • the antibodies disclosed herein can be recombinant human antibodies, which are human antibodies prepared, expressed, created or isolated by recombinant means, such as antibodies expressed using a recombinant expression vector transfected into a host cell, antibodies isolated from a recombinant, combinatorial human antibody library, antibodies isolated from an animal (e.g., a mouse or cow) that is transgenic and/or transchromosomal for human immunoglobulin genes (see e.g., Taylor, L. D., et al., Nucl.
  • Such recombinant human antibodies can have variable and constant regions derived from human germline immunoglobulin sequences (See Kabat, E. A., et al. (1991) Sequences of Proteins of Immunological Interest Fifth Edition, U.S. Department of Health and Human Services, NIH Publication No.91-3242).
  • such recombinant human antibodies are subjected to in vitro mutagenesis (or, when an animal transgenic for human Ig sequences is used, in vivo somatic mutagenesis) and thus the amino acid sequences of the VH and VL regions of the recombinant antibodies are sequences that, while derived from and related to human germline VH and VL sequences, may not naturally exist within the human antibody germline repertoire in vivo.
  • Modifications, Variations, and Derivatives [00333] Amino acid sequence modification(s) of the antibody or antigen-binding fragment thereof provided herein are contemplated.
  • the antibody or antigen-binding fragment thereof may be desirable to improve the binding affinity between the antibody or antigen-binding fragment thereof and the S polypeptide of a SARS-CoV-2; it may also be desirable to improve other biological properties of the antibody or antigen-binding fragment thereof, including but not limited to specificity, thermostability, expression level, or solubility.
  • variants can be prepared.
  • the antibody or antigen-binding fragment thereof provided herein are chemically modified, for example, by the covalent attachment of any type of molecule to the antibody or antigen-binding fragment thereof.
  • Exemplary non-limiting modifications include glycosylation, acetylation, pegylation, phosphorylation, amidation, derivatization by known protecting/blocking groups, proteolytic cleavage, linkage to a cellular ligand or other protein, etc.
  • the antibody or antigen-binding fragment thereof may contain one or more non-classical amino acids.
  • variations may be a substitution, deletion, or insertion of one or more codons encoding the antibody or antigen-binding fragment thereof that results in a change in the amino acid sequence as compared with the original sequence.
  • the variation allowed may be determined by systematically making insertions, deletions, or substitutions of amino acids in the sequence and testing the resulting variants for activity exhibited by the full-length or mature native sequence.
  • Amino acid substitutions can be the result of replacing one amino acid with another amino acid having similar structural and/or chemical properties, such as the replacement of a leucine with a serine, e.g., conservative amino acid replacements.
  • Standard techniques known to those of skill in the art can be used to introduce mutations in the nucleotide sequence encoding a molecule provided herein, including, for example, site-directed mutagenesis and PCR-mediated mutagenesis which results in amino acid substitutions.
  • a “conservative amino acid substitution” is one in which the amino acid residue is replaced with an amino acid residue having a side chain with a similar charge.
  • Families of amino acid residues having side chains with similar charges have been defined in the art. These families include amino acids with basic side chains (e.g., lysine, arginine, histidine), acidic side chains (e.g., aspartic acid, glutamic acid), uncharged polar side chains (e.g., glycine, asparagine, glutamine, serine, threonine, tyrosine, cysteine), nonpolar side chains (e.g., alanine, valine, leucine, isoleucine, proline, phenylalanine, methionine, tryptophan), beta-branched side chains (e.g., threonine, valine, isoleucine) and aromatic side chains (e.g., tyrosine, phenylalanine, trypto
  • mutations can be introduced randomly along all or part of the coding sequence, such as by saturation mutagenesis, and the resultant mutants can be screened for biological activity to identify mutants that retain activity. Following mutagenesis, the encoded protein can be expressed and the activity of the protein can be determined.
  • Substantial modifications in the biological properties of the antibody are accomplished by selecting substitutions that differ significantly in their effect on maintaining (a) the structure of the polypeptide backbone in the area of the substitution, for example, as a sheet or helical conformation, (b) the charge or hydrophobicity of the molecule at the target site, or (c) the bulk of the side chain.
  • substitutions may be in the range of about 1 to 100 amino acids.
  • the substitution includes fewer than about 100 amino acid substitutions, fewer than about 95 amino acid substitutions, fewer than about 90 amino acid substitutions, fewer than about 85 amino acid substitutions, fewer than about 80 amino acid substitutions, fewer than about 75 amino acid substitutions, fewer than about 70 amino acid substitutions, fewer than about 65 amino acid substitutions, fewer than about 50 amino acid substitutions, fewer than about 45 amino acid substitutions, fewer than about 40 amino acid substitutions, fewer than about 35 amino acid substitutions, fewer than about 30 amino acid substitutions, fewer than about 25 amino acid substitutions, fewer than about 20 amino acid substitutions, fewer than about 15 amino acid substitutions, fewer than about 10 amino acid substitutions, fewer than about 5 amino acid substitutions, fewer than about 4 amino acid substitutions, fewer than about 3 amino acid substitutions, or fewer than about 2 amino acid substitutions relative to the original molecule.
  • Amino acid sequence insertions include amino- and/or carboxyl-terminal fusions ranging in length from 1 residue to polypeptides containing 100 or more residues, as well as intrasequence insertions of single or multiple amino acid residues.
  • terminal insertions include an antibody with an N-terminal methionyl residue.
  • Other insertional variants of the antibody molecule include the fusion to the N or C terminus of the antibody to an enzyme (e.g., for antibody-directed enzyme prodrug therapy) or a polypeptide which increases the serum half- life of the antibody.
  • the insertion is about 1 amino acid to about 100 amino acids. In some embodiments, the insertion is at least about 1 amino acid.
  • the insertion is at most about 100 amino acids. In some embodiments, the insertion is about 1 amino acid to about 5 amino acids, about 1 amino acid to about 10 amino acids, about 1 amino acid to about 20 amino acids, about 1 amino acid to about 30 amino acids, about 1 amino acid to about 40 amino acids, about 1 amino acid to about 50 amino acids, about 1 amino acid to about 60 amino acids, about 1 amino acid to about 70 amino acids, about 1 amino acid to about 80 amino acids, about 1 amino acid to about 90 amino acids, about 1 amino acid to about 100 amino acids, about 5 amino acids to about 10 amino acids, about 5 amino acids to about 20 amino acids, about 5 amino acids to about 30 amino acids, about 5 amino acids to about 40 amino acids, about 5 amino acids to about 50 amino acids, about 5 amino acids to about 60 amino acids, about 5 amino acids to about 70 amino acids, about 5 amino acids to about 80 amino acids, about 5 amino acids to about 90 amino acids, about 5 amino acids to about 100 amino acids, about 10 amino acids to about 20 amino acids, about 10 amino acids to about 30 amino acids,
  • the insertion is about 1 amino acid, about 5 amino acids, about 10 amino acids, about 20 amino acids, about 30 amino acids, about 40 amino acids, about 50 amino acids, about 60 amino acids, about 70 amino acids, about 80 amino acids, about 90 amino acids, or about 100 amino acids.
  • Amino acid sequence deletions include amino- and/or carboxyl-terminal deletions ranging in length from one residue to polypeptides containing a hundred or more residues, as well as intrasequence deletions of single or multiple amino acid residues.
  • the deletion is about 1 amino acid to about 100 amino acids. In some embodiments, the deletion is at least about 1 amino acid. In some embodiments, the deletion is at most about 100 amino acids.
  • the deletion is about 1 amino acid to about 5 amino acids, about 1 amino acid to about 10 amino acids, about 1 amino acid to about 20 amino acids, about 1 amino acid to about 30 amino acids, about 1 amino acid to about 40 amino acids, about 1 amino acid to about 50 amino acids, about 1 amino acid to about 60 amino acids, about 1 amino acid to about 70 amino acids, about 1 amino acid to about 80 amino acids, about 1 amino acid to about 90 amino acids, about 1 amino acid to about 100 amino acids, about 5 amino acids to about 10 amino acids, about 5 amino acids to about 20 amino acids, about 5 amino acids to about 30 amino acids, about 5 amino acids to about 40 amino acids, about 5 amino acids to about 50 amino acids, about 5 amino acids to about 60 amino acids, about 5 amino acids to about 70 amino acids, about 5 amino acids to about 80 amino acids, about 5 amino acids to about 90 amino acids, about 5 amino acids to about 100 amino acids, about 10 amino acids to about 20 amino acids, about 10 amino acids to about 30 amino acids, about 10 amino acids to about 40 amino acids, about 10 amino acids to about 50 amino acids, about
  • the deletion is about 1 amino acid, about 5 amino acids, about 10 amino acids, about 20 amino acids, about 30 amino acids, about 40 amino acids, about 50 amino acids, about 60 amino acids, about 70 amino acids, about 80 amino acids, about 90 amino acids, or about 100 amino acids.
  • a polypeptide variant comprising an amino acid sequence that is at least about 75 %, about 80 %, about 85 %, about 90 %, about 91 %, about 92 %, about 93 %, about 94 %, about 95 %, about 96 %, about 97 %, about 98%, or about 99 % identical to the amino acid sequence of a polypeptide disclosed herein.
  • a “molecule derived from an antibody” refers to a functional antigen-binding fragment of an antibody. It is a portion of an antibody heavy and/or light chain polypeptide that retains some or all of the binding activity of the antibody from which the fragment was derived.
  • functional fragments include single-chain Fvs (scFv), Fab fragments, F(ab’) fragments, F(ab)2 fragments, F(ab’)2 fragments, disulfide-linked Fvs (dsFv), Fd fragments, Fv fragments, diabody, triabody, tetrabody, and minibody.
  • Such functional antigen-binding fragment can be found in, for example, Harlow and Lane, Antibodies: A Laboratory Manual (1989); Mol. Biology and Biotechnology: A Comprehensive Desk Reference (Myers, ed., 1995); Huston, et al, 1993, Cell Biophysics 22:189- 224; Pliickthun and Skerra, 1989, Meth. Enzymol. 178:497-515; and Day, Advanced Immunochemistrv (2d ed.1990).
  • Compositions and Kits [00346]
  • the antibodies of this invention represent an excellent way for the development of antiviral therapies either alone or in antibody cocktails with additional anti-SARS-CoV-2 virus antibodies for the treatment of human SARS-CoV-2 infections in humans.
  • the present invention provides a pharmaceutical composition
  • a pharmaceutical composition comprising the antibodies of the present invention described herein formulated together with a pharmaceutically acceptable carrier.
  • the composition may optionally contain one or more additional pharmaceutically active ingredients, such as another antibody or a therapeutic agent.
  • the pharmaceutical comprises two or more of the antibody or antigen-binding fragment thereof described herein, such as any combinations of the antibody or antigen-binding fragment thereof comprising a heavy chain and a light chain that comprise the respective amino acid sequences described herein.
  • the present disclosure provides compositions comprising two or more monoclonal antibodies, or antigen- binding fragments thereof, described herein.
  • the present disclosure provides compositions comprising 2, 3, or 4 monoclonal antibodies, or antigen-binding fragments thereof, described herein.
  • the present disclosure provides a composition comprising one of the following combinations of monoclonal antibodies or antigen- binding fragments thereof:
  • the pharmaceutical compositions disclosed herein are for use in treating a SARS-CoV-2 infection in a subject.
  • the pharmaceutical compositions disclosed herein are for use in the prevention of a SARS-CoV-2 infection in a subject.
  • the pharmaceutical compositions of the invention also can be administered in a combination therapy with, for example, another immune-stimulatory agent, an antiviral agent, a vaccine, etc.
  • a composition comprises an antibody of this invention at a concentration of at least 1 mg/ml, 5 mg/ml, 10 mg/ml, 50 mg/ml, 100 mg/ml, 150 mg/ml, 200 mg/ml, 1-300 mg/ml, or 100-300 mg/ml.
  • the second therapeutic agent comprises an anti-inflammatory drug or an antiviral compound.
  • the antiviral compound comprises: a nucleoside analog, a peptoid, an oligopeptide, a polypeptide, a protease inhibitor, a 3C-like protease inhibitor, a papain-like protease inhibitor, or an inhibitor of an RNA dependent RNA polymerase.
  • the antiviral compound may include: acyclovir, ganciclovir, vidarabine, foscarnet, cidofovir, amantadine, ribavirin, trifluorothymidine, zidovudine, didanosine, zalcitabine, or an interferon.
  • the interferon is an interferon- ⁇ RU ⁇ DQ ⁇ interferon- ⁇
  • the pharmaceutical composition can comprise any number of excipients. Excipients that can be used include carriers, surface-active agents, thickening or emulsifying agents, solid binders, dispersion or suspension aids, solubilizers, colorants, flavoring agents, coatings, disintegrating agents, lubricants, sweeteners, preservatives, isotonic agents, and combinations thereof.
  • a pharmaceutical composition is suitable for intravenous, intramuscular, subcutaneous, parenteral, spinal, or epidermal administration (e.g., by injection or infusion).
  • the active compound can be coated in a material to protect it from the action of acids and other natural conditions that may inactivate it.
  • parenteral administration means modes of administration other than enteral and topical administration, usually by injection, and includes, without limitation, intravenous, intramuscular, intraarterial, intrathecal, intracapsular, intraorbital, intracardiac, intradermal, intraperitoneal, transtracheal, subcutaneous, subcuticular, intraarticular, subcapsular, subarachnoid, intraspinal, epidural and intrasternal injection and infusion.
  • an antibody of the present invention described herein can be administered via a non-parenteral route, such as a topical, epidermal, or mucosal route of administration, e.g., intranasally, orally, vaginally, rectally, sublingually, or topically.
  • a non-parenteral route such as a topical, epidermal, or mucosal route of administration, e.g., intranasally, orally, vaginally, rectally, sublingually, or topically.
  • the pharmaceutical compositions of the invention may be prepared in many forms that include tablets, hard or soft gelatin capsules, aqueous solutions, suspensions, liposomes, and other slow-release formulations, such as shaped polymeric gels.
  • An oral dosage form may be formulated such that the antibody is released into the intestine after passing through the stomach. Such formulations are described in U.S. Pat. No.6,306,434 and in the references contained therein.
  • Oral liquid pharmaceutical compositions may be in the form of, for example, aqueous or oily suspensions, solutions, emulsions, syrups, or elixirs, or may be presented as a dry product for constitution with water or other suitable vehicle before use.
  • Such liquid pharmaceutical compositions may contain conventional additives such as suspending agents, emulsifying agents, non-aqueous vehicles (which may include edible oils), or preservatives.
  • An antibody can be formulated for parenteral administration (e.g., by injection, for example, bolus injection or continuous infusion) and may be presented in unit dosage form in ampules, prefilled syringes, small volume infusion containers, or multi-dose containers with an added preservative.
  • compositions may take such forms as suspensions, solutions, or emulsions in oily or aqueous vehicles, and may contain formulatory agents such as suspending, stabilizing and/or dispersing agents.
  • Pharmaceutical compositions suitable for rectal administration can be prepared as unit dose suppositories. Suitable carriers include saline solution and other materials commonly used in the art. [00358]
  • an antibody can be conveniently delivered from an insufflator, nebulizer, a pressurized pack, or other convenient means of delivering an aerosol spray.
  • Pressurized packs may comprise a suitable propellant such as dichlorodifluoromethane, trichlorofluoromethane, dichlorotetrafluoroethane, carbon dioxide, or other suitable gas.
  • a suitable propellant such as dichlorodifluoromethane, trichlorofluoromethane, dichlorotetrafluoroethane, carbon dioxide, or other suitable gas.
  • the dosage unit may be determined by providing a valve to deliver a metered amount.
  • an antibody may take the form of a dry powder composition, for example, a powder mix of a modulator and a suitable powder base such as lactose or starch.
  • the powder composition may be presented in a unit dosage form in, for example, capsules or cartridges or, e.g., gelatin or blister packs from which the powder may be administered with the aid of an inhalator or insufflator.
  • an antibody may be administered via a liquid spray, such as via a plastic bottle atomizer.
  • Pharmaceutical compositions of the invention may also contain other ingredients such as flavorings, colorings, anti-microbial agents, or preservatives. It will be appreciated that the amount of an antibody required for use in treatment will vary not only with the particular carrier selected but also with the route of administration, the nature of the condition being treated, and the age and condition of the patient. Ultimately the attendant health care provider may determine a proper dosage.
  • a pharmaceutical composition may be formulated as a single unit dosage form.
  • the pharmaceutical composition of the present invention can be in the form of sterile aqueous solutions or dispersions. It can also be formulated in a microemulsion, liposome, or other ordered structure suitable to high drug concentration.
  • An antibody of the present invention described herein can be administered as a sustained release formulation, in which case less frequent administration is required. Dosage and frequency vary depending on the half-life of the antibody in the patient. In general, human antibodies show the longest half-life, followed by humanized antibodies, chimeric antibodies, and nonhuman antibodies. The dosage and frequency of administration can vary depending on whether the treatment is prophylactic or therapeutic.
  • a relatively low dosage is administered at relatively infrequent intervals over a long period of time. Some patients continue to receive treatment for the rest of their lives. In therapeutic applications, a relatively high dosage at relatively short intervals is sometimes required until progression of the disease is reduced or terminated, and preferably until the patient shows partial or complete amelioration of symptoms of disease. Thereafter, the patient can be administered a prophylactic regime.
  • the amount of active ingredient that can be combined with a carrier material to produce a single dosage form will vary depending upon the subject being treated and the particular mode of administration and will generally be that amount of the composition, which produces a therapeutic effect.
  • this amount will range from about 0.01% to about 99% of active ingredient, preferably from about 0.1% to about 70%, most preferably from about 1% to about 30% of active ingredient in combination with a pharmaceutically acceptable carrier.
  • Dosage regimens can be adjusted to provide the optimum desired response (e.g., a therapeutic response). For example, a single bolus can be administered, several divided doses can be administered over time, or the dose can be proportionally reduced or increased as indicated by the exigencies of the therapeutic situation. It is especially advantageous to formulate parenteral compositions in dosage unit form for ease of administration and uniformity of dosage.
  • Dosage unit form as used herein refers to physically discrete units suited as unitary dosages for the subjects to be treated; each unit contains a predetermined quantity of active compound calculated to produce the desired therapeutic effect in association with the required pharmaceutical carrier.
  • the antibody can be administered as a sustained release formulation, in which case less frequent administration is required.
  • the dosage ranges from about 0.0001 to 800 mg/kg, and more usually 0.01 to 5 mg/kg, of the host body weight.
  • dosages can be 0.3 mg/kg body weight, 1 mg/kg body weight, 3 mg/kg body weight, 5 mg/kg body weight or 10 mg/kg body weight or within the range of 1-10 mg/kg.
  • An exemplary treatment regime entails administration once per week, once every two weeks, once every three weeks, once every four weeks, once a month, once every 3 months or once every three to 6 months.
  • Preferred dosage regimens for an antibody of the invention include 1 mg/kg body weight or 3 mg/kg body weight via intravenous administration, with the antibody being given using one of the following dosing schedules: (i) every four weeks for six dosages, then every three months; (ii) every three weeks; (iii) 3 mg/kg body weight once followed by 1 mg/kg body weight every three weeks.
  • dosage is adjusted to achieve a plasma antibody concentration of about 1-1000 ug /ml, and in some methods, about 25-300 ⁇ g /ml.
  • a “therapeutically effective dosage” of an antibody of the invention preferably results in a decrease in seventy of disease symptoms, an increase in frequency and duration of disease symptom-free periods, or a prevention of impairment or disability due to the disease affliction.
  • a “therapeutically effective dosage” preferably inhibits SARS-CoV- 2 virus replication or uptake by host cells by at least about 20%, more preferably by at least about 40%, even more preferably by at least about 60%, and still more preferably by at least about 80% relative to untreated subjects.
  • a therapeutically effective amount of a therapeutic compound can neutralize SARS-CoV-2 virus, or otherwise ameliorate symptoms in a subject, which is typically a human or can be another mammal.
  • the pharmaceutical composition can be a controlled release formulation, including implants, transdermal patches, and microencapsulated delivery systems.
  • Biodegradable, biocompatible polymers can be used, such as ethylene-vinyl acetate, polyanhydrides, polyglycolic acid, collagen, poly orthoesters, and polylactic acid. See, e.g., Sustained and Controlled Release Drug Delivery Systems, J. R. Robinson, ed., Marcel Dekker, Inc., New York, 1978.
  • compositions can be administered via medical devices such as (1) needleless hypodermic injection devices (e.g,, US 5,399,163; 5,383,851; 5,312,335; 5,064,413; 4,941,880; 4,790,824; and 4,596,556); (2) micro-infusion pumps (US 4,487,603); (3) transdermal devices (US 4,486,194); (4) infusion apparati (US 4,447,233 and 4,447,224); and (5) osmotic devices (US 4,439,196 and 4,475,196); the disclosures of which are incorporated herein byreference.
  • needleless hypodermic injection devices e.g, US 5,399,163; 5,383,851; 5,312,335; 5,064,413; 4,941,880; 4,790,824; and 4,596,556
  • micro-infusion pumps e.g, US 5,487,603
  • transdermal devices e.g., dermal devices
  • the human monoclonal antibodies of the invention described herein can be formulated to ensure proper distribution in vivo.
  • the therapeutic compounds of the invention can be formulated in liposomes, which may additionally comprise targeting moieties to enhance selective transport to specific cells or organs. See, e.g., US 4,522,811; 5,374,548; 5,416,016; and 5,399,331; V.V. Ranade (1989) Clin. Pharmacol. 29:685; Urnezawa et al., (1988) Biochem. Biophys. Res. Cornmun. 153:1038; Bloeman et al., (1995) FEBS Lett.
  • the initial dose may be followed by administration of a second or a plurality of subsequent doses of the antibody or antigen-binding fragment thereof in an amount that can be approximately the same or less than that of the initial dose, wherein the subsequent doses are separated by at least 1 day to 3 days; at least one week, at least 2 weeks; at least 3 weeks; at least 4 weeks; at least 5 weeks; at least 6 weeks; at least 7 weeks; at least 8 weeks; at least 9 weeks; at least 10 weeks; at least 12 weeks; or at least 14 weeks.
  • Various delivery systems are known and can be used to administer the pharmaceutical composition of the invention, e.g., encapsulation in liposomes, microparticles, microcapsules, recombinant cells capable of expressing the mutant viruses, receptor-mediated endocytosis (see, e.g., Wu et al., (1987) J. Biol. Chem. 262:4429-4432).
  • Methods of introduction include, but are not limited to, intradermal, transdermal, intramuscular, intraperitoneal, intravenous, subcutaneous, intranasal, epidural, and oral routes.
  • the composition may be administered by any convenient route, for example by infusion or bolus injection, by absorption through epithelial or mucocutaneous linings (e.g., oral mucosa, rectal and intestinal mucosa, etc.) and may be administered together with other biologically active agents. Administration can be systemic or local.
  • the pharmaceutical composition can also be delivered in a vesicle, in particular, a liposome (see, for example, Langer (1990) Science 249: 1527-1533).
  • the use of nanoparticles to deliver the antibodies of the present invention is also contemplated herein. Antibody-conjugated nanoparticles may be used both for therapeutic and diagnostic applications.
  • Nanoparticles may be developed and conjugated to antibodies contained in pharmaceutical compositions to target cells. Nanoparticles for drug delivery have also been described in, for example, US 8257740, or US 8246995, each incorporated herein in its entirety.
  • the pharmaceutical composition can be delivered in a controlled release system. In one embodiment, a pump may be used. In another embodiment, polymeric materials can be used.
  • the injectable preparations may include dosage forms for intravenous, subcutaneous, intracutaneous, intracranial, intraperitoneal, intramuscular injections, drip infusions, etc. These injectable preparations may be prepared by methods publicly known. For example, the injectable preparations may be prepared, e.g., by dissolving, suspending, or emulsifying the antibody or its salt described herein in a sterile aqueous medium or an oily medium conventionally used for injections.
  • aqueous medium for injections there are, for example, physiological saline, an isotonic solution containing glucose and other auxiliary agents, etc., which may be used in combination with an appropriate solubilizing agent such as an alcohol (e.g., ethanol), a polyalcohol (e.g., propylene glycol, polyethylene glycol), a nonionic surfactant [e.g., polysorbate 80, HCO-50 (polyoxyethylene (50 mol) adduct of hydrogenated castor oil)], etc.
  • an alcohol e.g., ethanol
  • a polyalcohol e.g., propylene glycol, polyethylene glycol
  • a nonionic surfactant e.g., polysorbate 80, HCO-50 (polyoxyethylene (50 mol) adduct of hydrogenated castor oil
  • a pharmaceutical composition of the present invention can be delivered subcutaneously or intravenously with a standard needle and syringe.
  • a pen delivery device readily has applications in delivering a pharmaceutical composition of the present invention.
  • Such a pen delivery device can be reusable or disposable.
  • a reusable pen delivery device generally utilizes a replaceable cartridge that contains a pharmaceutical composition.
  • the empty cartridge can readily be discarded and replaced with a new cartridge that contains the pharmaceutical composition.
  • the pen delivery device can then be reused.
  • a disposable pen delivery device there is no replaceable cartridge. Rather, the disposable pen delivery device comes prefilled with the pharmaceutical composition held in a reservoir within the device. Once the reservoir is emptied of the pharmaceutical composition, the entire device is discarded.
  • Numerous reusable pens and autoinjector delivery devices have applications in the subcutaneous delivery of a pharmaceutical composition of the present invention.
  • Examples include, but certainly are not limited to AUTOPENTM (Owen Mumford, Inc., Woodstock, UK), DISETRONICTM pen (Disetronic Medical Systems, Burghdorf, Switzerland), HUMALOG MIX 75/25TM pen, HUMALOGTM pen, HUMALIN 70/30TM pen (Eli Lilly and Co., Indianapolis, IN), NOVOPENTM I, II and III (Novo Nordisk, Copenhagen, Denmark), NOVOPEN JUNIORTM (Novo Nordisk, Copenhagen, Denmark), BDTM pen (Becton Dickinson, Franklin Lakes, NJ), OPTIPENTM, OPTIPEN PROTM, OPTIPEN STARLETTM, and OPTICLIKTM (Sanofi-Aventis, Frankfurt, Germany), to name only a few.
  • Examples of disposable pen delivery devices having applications in subcutaneous delivery of a pharmaceutical composition of the present invention include, but certainly are not limited to, the SOLOSTARTM pen (Sanofi- Aventis), the FLEXPENTM (Novo Nordisk), and the KWIKPENTM (Eli Lilly), the SURECLICKTM Autoinjector (Amgen, Thousand Oaks, CA), the PENLETTM (Haselmeier, Stuttgart, Germany), the EPIPEN® (Dey, L.P.) and the HUMIRATM Pen (Abbott Labs, Abbott Park, IL), to name only a few.
  • the pharmaceutical compositions for oral or parenteral use described herein are prepared into dosage forms in a unit dose suited to fit a dose of the active ingredients.
  • Such dosage forms in a unit dose include, for example, tablets, pills, capsules, injections (ampoules), suppositories, etc.
  • the amount of the antibody contained is generally about 5 to about 500 mg per dosage form in a unit dose; especially in the form of injection, it is preferred that the antibody is contained in about 5 to about 300 mg and in about 10 to about 300 mg for the other dosage forms.
  • this disclosure provides a kit comprising a pharmaceutically acceptable dose unit of the antibody or antigen-binding fragment thereof or the pharmaceutical composition as described herein.
  • kits for the diagnosis, prognosis, or monitoring of treatment of SARS-CoV-2 in a subject comprising: the antibody or antigen-binding fragment thereof as described; and a least one detection reagent that binds specifically to the antibody or antigen-binding fragment thereof.
  • the kit also includes a container that contains the composition and optionally informational material.
  • the informational material can be descriptive, instructional, marketing, or other material that relates to the methods described herein and/or the use of the agents for therapeutic benefit.
  • the kit also includes an additional therapeutic agent, as described herein.
  • the kit includes a first container that contains the composition and a second container for the additional therapeutic agent.
  • the informational material of the kits is not limited in its form.
  • the informational material can include information about production of the composition, concentration, date of expiration, batch or production site information, and so forth.
  • the informational material relates to methods of administering the composition, e.g., in a suitable dose, dosage form, or mode of administration (e.g., a dose, dosage form, or mode of administration described herein), to treat a subject in need thereof.
  • the instructions provide a dosing regimen, dosing schedule, and/or route of administration of the composition or the additional therapeutic agent.
  • the information can be provided in a variety of formats, including printed text, computer-readable material, video recording, audio recording, or information that contains a link or address to substantive material.
  • the kit can include one or more containers for the composition.
  • the kit contains separate containers, dividers, or compartments for the composition and informational material.
  • the composition can be contained in a bottle or vial, and the informational material can be contained in a plastic sleeve or packet.
  • the separate elements of the kit are contained within a single, undivided container.
  • the composition is contained in a bottle or vial that has attached thereto the informational material in the form of a label.
  • the kit includes a plurality (e.g., a pack) of individual containers, each containing one or more unit dosage forms (e.g., a dosage form described herein) of the agents.
  • the kit optionally includes a device suitable for administration of the composition or other suitable delivery device.
  • the device can be provided pre-loaded with one or both of the agents or can be empty, but suitable for loading.
  • Such a kit may optionally contain a syringe to allow for injection of the antibody contained within the kit into an animal, such as a human. 1.6.
  • compositions thereof Provided herein are methods of treating a SARS-CoV-2 infection in a subject, comprising administering to the subject a therapeutically effective amount of the antibodies, antigen-binding fragments thereof, or pharmaceutical compositions thereof disclosed herein. Also provided herein are methods of preventing a SARS-CoV-2 infection in a subject, comprising administering to the subject a therapeutically effective amount of the antibodies, antigen-binding fragments thereof, or pharmaceutical compositions thereof disclosed herein. Also provided herein are methods of neutralizing a SARS-CoV-2 infection in a subject, comprising administering to the subject a therapeutically effective amount of the antibodies, antigen-binding fragments thereof, or pharmaceutical compositions thereof disclosed herein.
  • the method described herein may be used in the treatment and/or prevention of SARS-CoV-2.
  • SARS-CoV-2 variants include WHO alpha variant, WHO beta variant, WHO gamma variant, WHO delta variant, WHO epsilon variant, WHO Eta variant, WHO iota variant, WHO kappa variant, WHO omicron variant, WHO zeta variant, WHO mu variant, and B.1.617.3.
  • the subject may be a neonate, a juvenile, or an adult. In some embodiments, the subject is human.
  • the subject is non-human primates (e.g., monkeys, baboons, and chimpanzees), mice, rats, bovines, horses, household cats, tigers and other large cats, dogs, pigs, rabbits, goats, deer, sheep, ferrets, gerbils, guinea pigs, hamsters, bats, and birds (e.g., chickens, turkeys, and ducks).
  • a number of these household pets and farm animals are capable of carrying and transmitting SARS-CoV-2 viruses without themselves getting substantially sick or dying, thereby transmitting the disease to humans.
  • these animals are treated not because they are suffering from disease, but rather, because they can transmit virus to humans and cause human disease.
  • the antibodies, antigen-binding fragments thereof, or pharmaceutical compositions thereof are administered to a subject in order to prevent infection with a SARS-CoV-2 virus. In some embodiments, the antibodies, antigen-binding fragments thereof, or pharmaceutical compositions thereof disclosed herein are administered to a subject prior to exposure to or infection with SARS-CoV-2 viruses. In some embodiments, the antibodies, antigen-binding fragments thereof, or pharmaceutical compositions thereof disclosed herein are administered to a subject after the exposure to or infection with SARS-CoV-2 viruses.
  • the neutralizing of the SARS-CoV-2 virus can be done via (i) inhibiting SARS-CoV-2 virus binding to a target cell; (ii) inhibiting SARS-CoV-2 virus uptake by a target cell; (iii) inhibiting SARS- CoV-2 virus replication; and (iv) inhibiting SARS-CoV-2 virus particles release from infected cells.
  • One skilled in the art possesses the ability to perform any assay to assess neutralization of SARS-CoV-2 virus.
  • the neutralizing properties of antibodies may be assessed by a variety of tests, which all may assess the consequences of (i) inhibition of SARS-CoV-2 virus binding to a target cell; (ii) inhibition of SARS-CoV-2 virus uptake by a target cell; (iii) inhibition of SARS-CoV-2 virus replication; and (iv) inhibition of SARS-CoV-2 virus particles release from infected cells.
  • implementing different tests may lead to the observation of the same consequence, i.e., the loss of infectivity of the SARS-CoV-2 virus.
  • the present invention provides a method of neutralizing a SARS-CoV-2 virus in a subject comprising administering to the subject a therapeutically effective amount of the antibody of the present invention described herein.
  • the present disclosure provides methods of preventing infection with a SARS-CoV-2 virus in an immunocompromised subject.
  • Immunocompromised subjects include subjects that suffer from an immune deficiency (e.g., a primary or acquired immune deficiency) or autoimmune disease, subjects that have undergone or are currently undergoing treatment with one or more immunosuppressive drugs (e.g., chemotherapy, glucocorticoids, protease inhibitors, immune cell depleting monoclonal antibodies, etc.), subjects that have recently received an organ transplant or hematopoietic stem cell transplant, subjects that have received a CAR-T therapy, and subjects that have undergone or are currently undergoing radiation treatment.
  • Immunosuppressive drugs are known to those in the art. See e.g., Hussain Y, Khan H. Immunosuppressive Drugs. Encyclopedia of Infection and Immunity.
  • Vaccines including vaccines against SARS-CoV-2 infections, are less effective in immunocompromised subjects. Furthermore, some immunocompromised subjects may not be able to receive a SARS- CoV-2 vaccine.
  • the antibodies and antigen-binding fragments thereof provided herein therefore provide a therapeutic option for subjects who cannot receive a SARS-CoV-2 vaccine or who demonstrate reduced efficacy of a SARS-CoV-2 vaccine.
  • Another aspect of the present invention provides a method of treating a SARS-CoV-2- related disease. Such a method includes therapeutic (following SARS-CoV-2 infection) and prophylactic (prior to SARS-CoV-2 exposure, infection, or pathology).
  • therapeutic and prophylactic methods of treating an individual for a SARS-CoV-2 infection include treatment of an individual having or at risk of having a SARS-CoV-2 infection or pathology, treating an individual with a SARS-CoV-2 infection, and methods of protecting an individual from a SARS- CoV-2 infection, to decrease or reduce the probability of a SARS-CoV-2 infection in an individual, to decrease or reduce susceptibility of an individual to a SARS-CoV-2 infection, or to inhibit or prevent a SARS-CoV-2 infection in an individual, and to decrease, reduce, inhibit or suppress transmission of a SARS-CoV-2 from an infected individual to an uninfected individual.
  • Such methods include administering an antibody of the present invention or a composition comprising the antibody disclosed herein to therapeutically or prophylactically treat (vaccinate or immunize) an individual having or at risk of having a SARS-CoV-2 infection or pathology. Accordingly, methods can treat the SARS-CoV-2 infection or pathology, or provide the individual with protection from infection (e.g., prophylactic protection).
  • a method of treating a SARS-CoV-2-related disease comprises administering to an individual in need thereof an antibody or therapeutic composition disclosed herein in an amount sufficient to reduce one or more physiological conditions or symptoms associated with a SARS-CoV-2 infection or pathology, thereby treating the SARS-CoV-2 -related disease.
  • an antibody or therapeutic composition disclosed herein is used to treat a SARS-CoV-2-related disease.
  • Use of an antibody or therapeutic composition disclosed herein treats a SARS-CoV-2-related disease by reducing one or more physiological conditions or symptoms associated with a SARS-CoV-2 infection or pathology.
  • administration of an antibody or therapeutic composition disclosed herein is in an amount sufficient to reduce one or more physiological conditions or symptoms associated with a SARS- CoV-2 infection or pathology, thereby treating the SARS-CoV-2-based disease.
  • administering refers to the treatment of a disease in a mammal, e.g., in a human, including (a) inhibiting the disease, i.e., arresting disease development or preventing disease progression; (b) relieving the disease, i.e., causing regression of the disease state or relieving one or more symptoms of the disease; and (c) curing the disease, i.e., remission of one or more disease symptoms.
  • treatment results in an improvement or remediation of the symptoms of the disease.
  • treatment may refer to a short- term (e.g., temporary and/or acute) and/or a long-term (e.g., sustained) improvement or remediation in one or more disease symptoms.
  • the improvement is an observable or measurable improvement.
  • the improvement is an improvement in the general feeling of well-being of the subject.
  • administration of the pharmaceutical compositions disclosed herein may reduce one or more symptoms of the SARS-COV-2 infection, including but not limited to, death, incidence of emphysema, incidence of pneumonia, shortness of breath, racing heart, fever, cough, sore throat, congestion, muscle or body aches, headaches, fatigue, vomiting, diarrhea, loss of taste or smell, cognitive issues like “brain fog”, memory or attention problems, and Postural Orthostatic Tachycardia Syndrome (POTS).
  • PTS Postural Orthostatic Tachycardia Syndrome
  • the method of neutralizing SARS-CoV-2 in a subject comprises administering to a subject in need thereof a therapeutically effective amount of a first antibody or antigen-binding fragment thereof and a second antibody or antigen-binding fragment thereof of the antibody or antigen-binding fragment, as described herein, wherein the first antibody or antigen-binding fragment thereof and the second antibody or antigen-binding fragment thereof exhibit synergistic activity or a therapeutically effective amount of the pharmaceutical composition described herein.
  • the method of preventing or treating a SARS-CoV-2 infection comprising administering to a subject in need thereof a therapeutically effective amount of a first antibody or antigen-binding fragment thereof and a second antibody or antigen-binding fragment thereof of the antibody or antigen-binding fragment, as described herein, wherein the first antibody or antigen-binding fragment thereof and the second antibody or antigen-binding fragment thereof exhibit synergistic activity or a therapeutically effective amount of the pharmaceutical composition described herein.
  • the first antibody or antigen-binding fragment thereof is administered before, after, or concurrently with the second antibody or antigen-binding fragment thereof.
  • the first antibody or antigen-binding fragment thereof and the second antibody or antigen-binding fragment thereof can be any combinations of the antibody or antigen-binding fragment thereof comprising a heavy chain and a light chain that comprise the respective amino acid sequences described herein.
  • the first antibody or antigen-binding fragment thereof comprises a heavy chain variable region and a light chain variable region comprising a HCVR/LCVR amino acid sequence pair of SEQ ID NOs: 1-2, 3-4, 5-6, 7-8, 9-10, 11-12, 13-14, 15-16, 17-18, 19-20, 21-22, 23-24, 25-26, 27-28, 29-30, 31-32, 33-34, 35-36, 37-38, 39-40, 41-42, 43-44, 45-46, 47-48, 49-50, 51-52, 53-54, 55-56, 57-58, 59-60, 61-62, 63-64, 65-66, 67-68, 69-70, 71-72, 73-74, 75-76, 77-78, 79-80, 81-82, 83-84, 85-86, 87-88, 89-90, 91-92, 93-94, 95-96, 97-98, 99-
  • the first antibody or antigen-binding fragment thereof comprises a heavy chain variable region and a light chain variable region comprising a HCVR/LCVR amino acid sequence pair of SEQ ID NOs: 25-26, 191-192, 219-220, 395-396, or 397-398.
  • the second antibody or antigen-binding fragment thereof comprising a HCVR/LCVR amino acid sequence pair of SEQ ID NOs: 1-2, 3-4, 5-6, 7-8, 9-10, 11-12, 13-14, 15-16, 17-18, 19-20, 21-22, 23-24, 25-26, 27-28, 29-30, 31-32, 33-34, 35-36, 37-38, 39-40, 41-42, 43-44, 45-46, 47-48, 49-50, 51-52, 53-54, 55-56, 57-58, 59-60, 61-62, 63-64, 65-66, 67-68, 69-70, 71-72, 73-74, 75-76, 77-78, 79-80, 81-82, 83-84, 85-86, 87-88, 89-90, 91-92, 93-94, 95-96, 97-98, 99-100, 101-102, 103-104, 105-
  • the second antibody or antigen-binding fragment thereof comprises a HCVR/LCVR amino acid sequence pair of SEQ ID NOs: 25-26, 191-192, 219-220, 395-396, or 397-398, wherein the first antibody or antigen-binding fragment thereof is different from the second antibody or antigen-binding fragment thereof.
  • the second therapeutic agent comprises an anti-inflammatory drug or an antiviral compound.
  • the antiviral compound comprises: a nucleoside analog, a peptoid, an oligopeptide, a polypeptide, a protease inhibitor, a 3C-like protease inhibitor, a papain-like protease inhibitor, or an inhibitor of an RNA dependent RNA polymerase.
  • the antiviral compound may include: acyclovir, ganciclovir, vidarabine, foscarnet, cidofovir, amantadine, ribavirin, trifluorothymidine, zidovudine, didanosine, zalcitabine, or an interferon.
  • the interferon is an interferon- ⁇ RU ⁇ DQ ⁇ interferon- ⁇
  • the antibody or antigen-binding fragment thereof is administered before, after, or concurrently with the second therapeutic agent or therapy.
  • the antibody or antigen-binding fragment thereof is administered to the subject intravenously, subcutaneously, or intraperitoneally.
  • the antibody or antigen-binding fragment thereof is administered prophylactically or therapeutically.
  • the antibodies described herein can be used together with one or more of other anti- SARS-CoV-2 virus antibodies to neutralize SARS-CoV-2 virus, thereby treating SARS-CoV-2 infections.
  • administration route is intraarterial, intracranial, intradermal, intraduodenal, intrammamary, intrameningeal, intraperitoneal, intrathecal, intratumoral, intravenous, intravitreal, ophthalmic, parenteral, spinal, subcutaneous, ureteral, urethral, vaginal, or intrauterine.
  • administration route is local or systemic.
  • the effective amount of the antibodies, antigen-binding fragments thereof, or pharmaceutical compositions thereof administered to a particular subject will depend on a variety of factors, several of which will differ from patient to patient including the disorder being treated and the severity of the disorder; activity of the specific agent(s) employed; the age, body weight, general health, sex and diet of the patient; the timing of administration, route of administration; the duration of the treatment; drugs used in combination; the judgment of the prescribing physician; and like factors known in the medical arts. Dosage amount and interval can be adjusted individually to provide plasma levels of the compound(s) which are sufficient to maintain therapeutic or prophylactic effect.
  • Dosage amounts of the pharmaceutical compositions disclosed herein can typically be in the range of from about 0.0001 mg/kg/day to about 1000 mg/kg/day, but can be higher or lower, depending upon, among other factors, the activity of the compound, its bioavailability, and various factors discussed above. In some embodiments, the dose is from about 0.0001 mg/kg to about 1000 mg/kg of body weight per day. In some embodiments, the dose is from about 0.001 mg/kg to about 1000 mg/kg of body weight per day.
  • the dose is from about 0.01 mg/kg to about 1000 mg/kg of body weight per day. In some embodiments, the dose is from about 0.1 mg/kg to about 100 mg/kg of body weight per day. In some embodiments, the dose is from about 0.5 mg/kg to about 50 mg/kg of body weight per day. In some embodiments, the dose is from about 1 mg/kg to about 25 mg/kg of body weight per day. In some embodiments, the dose is from about 5 mg/kg to about 15 mg/kg of body weight per day.
  • the dose is about 1 mg/kg, about 5 mg/kg, about 10 mg/kg, about 15 mg/kg, about 20 mg/kg, about 25 mg/kg, about 30 mg/kg, about 35 mg/kg, about 40 mg/kg, about 45 mg/kg, about 50 mg/kg, about 55 mg/kg, about 60 mg/kg, about 65 mg/kg, about 70 mg/kg, about 75 mg/kg, about 80 mg/kg, about 85 mg/kg, about 90 mg/kg, about 95 mg/kg, or about 100 mg/kg.
  • the number of administrations of treatment to a subject may vary.
  • introducing the antibodies, antigen-binding fragments thereof, or pharmaceutical compositions thereof into the subject may be a one-time event.
  • such treatment may require an on-going series of repeated treatments (e.g., once per day, once per week, or multiple times per day or week).
  • multiple administrations of the pharmaceutical compositions may be required before an effect is observed.
  • the exact protocols depend upon the disease or condition, the stage of the disease and parameters of the individual subject being treated.
  • Combination Therapies [00407]
  • the antibodies, antigen-binding fragments thereof, or pharmaceutical compositions thereof disclosed herein are administered in combination with one or more additional therapeutic composition(s).
  • the additional therapeutic composition is an anti-viral drug.
  • the additional therapeutic composition is a viral entry inhibitor.
  • the additional therapeutic composition is a viral attachment inhibitor.
  • the antibodies, antigen-binding fragments thereof, or pharmaceutical compositions thereof disclosed herein and the additional therapeutic composition(s) are administered simultaneously. In some embodiments, the antibodies, antigen-binding fragments thereof, or pharmaceutical compositions thereof disclosed herein is administered before the additional therapeutic composition(s). In some embodiments, the antibodies, antigen-binding fragments thereof, or pharmaceutical compositions thereof disclosed herein is administered after the additional therapeutic composition(s).
  • Combination therapies may include an anti- SARS-CoV-2 antibody of the invention and any additional therapeutic agent that may be advantageously combined with an antibody of the invention or with a biologically active fragment of an antibody of the invention.
  • the antibodies of the present invention may be combined synergistically with one or more drugs or therapy used to treat a disease or disorder associated with a viral infection, such as a SARS-CoV-2 infection.
  • the antibodies of the invention may be combined with a second therapeutic agent to ameliorate one or more symptoms of said disease.
  • the antibodies of the invention may be combined with a second antibody to provide synergistic activity in ameliorating one or more symptoms of said disease.
  • the first antibody or antigen-binding fragment thereof is administered before, after, or concurrently with the second antibody or antigen-binding fragment thereof.
  • the antibody described herein can be used in various detection methods for use in, e.g., monitoring the progression of a SARS-CoV-2 infection; monitoring patient response to treatment for such an infection, etc.
  • the present disclosure provides methods of detecting a neuraminidase polypeptide in a biological sample obtained from an individual. The methods generally involve: a) contacting the biological sample with a subject anti- neuraminidase antibody; and b) detecting binding, if any, of the antibody to an epitope present in the sample.
  • the antibody comprises a detectable label.
  • the level of neuraminidase polypeptide detected in the biological sample can provide an indication of the stage, degree, or severity of a SARS-CoV-2 infection.
  • the level of the neuraminidase polypeptide detected in the biological sample can provide an indication of the individual's response to treatment for a SARS-CoV-2 infection.
  • the second therapeutic agent is another antibody to a SARS- COV-2 protein or a fragment thereof. It is contemplated herein to use a combination (“cocktail”) of antibodies with broad neutralization or inhibitory activity against SARS-COV-2.
  • non-competing antibodies may be combined and administered to a subject in need thereof.
  • the antibodies comprising the combination bind to distinct non- overlapping epitopes on the protein.
  • the second antibody may possess a longer half-life in human serum.
  • the term “in combination with” means that additional therapeutically active component(s) may be administered prior to, concurrent with, or after the administration of the anti-SARS-COV-2 antibody of the present invention.
  • the term “in combination with” also includes sequential or concomitant administration of an anti-SARS-COV-2 antibody and a second therapeutic agent.
  • the additional therapeutically active component(s) may be administered to a subject prior to administration of an anti-SARS-COV-2 antibody of the present invention.
  • a first component may be deemed to be administered “prior to” a second component if the first component is administered 1 week before, 72 hours before, 60 hours before, 48 hours before, 36 hours before, 24 hours before, 12 hours before, 6 hours before, 5 hours before, 4 hours before, 3 hours before, 2 hours before, 1 hour before, 30 minutes before, 15 minutes before, 10 minutes before, 5 minutes before, or less than 1 minute before administration of the second component.
  • the additional therapeutically active component(s) may be administered to a subject after administration of an anti-SARS-COV-2 antibody of the present invention.
  • a first component may be deemed to be administered “after” a second component if the first component is administered 1 minute after, 5 minutes after, 10 minutes after, 15 minutes after, 30 minutes after, 1 hour after, 2 hours after, 3 hours after, 4 hours after, 5 hours after, 6 hours after, 12 hours after, 24 hours after, 36 hours after, 48 hours after, 60 hours after, 72 hours after administration of the second component.
  • the additional therapeutically active component(s) may be administered to a subject concurrent with administration of an anti- SARS-COV-2 antibody of the present invention.
  • Constant administration includes, e.g., administration of an anti-SARS-COV-2 antibody and an additional therapeutically active component to a subject in a single dosage form, or in separate dosage forms administered to the subject within about 30 minutes or less of each other. If administered in separate dosage forms, each dosage form may be administered via the same route (e.g., both the anti-SARS-COV-2 antibody and the additional therapeutically active component may be administered intravenously, etc.); alternatively, each dosage form may be administered via a different route (e.g., the anti-SARS-COV-2 antibody may be administered intravenously, and the additional therapeutically active component may be administered orally).
  • each dosage form may be administered via the same route (e.g., both the anti-SARS-COV-2 antibody and the additional therapeutically active component may be administered intravenously, etc.); alternatively, each dosage form may be administered via a different route (e.g., the anti-SARS-COV-2 antibody may be administered intravenously, and the additional therapeutically active component may be administered
  • administering the components in a single dosage form, in separate dosage forms by the same route, or in separate dosage forms by different routes are all considered “concurrent administration,” for purposes of the present disclosure.
  • administration of an anti-SARS-COV-2 antibody “prior to,” “concurrent with,” or “after” (as those terms are defined hereinabove) administration of an additional therapeutically active component is considered administration of an anti-SARS-COV-2 antibody “in combination with” an additional therapeutically active component.
  • the present invention includes pharmaceutical compositions in which an anti-SARS- COV-2 antibody of the present invention is co-formulated with one or more of the additional therapeutically active component(s) as described elsewhere herein.
  • a single dose of an anti-SARS-COV-2 antibody of the invention may be administered to a subject in need thereof.
  • multiple doses of an anti-SARS-COV-2 antibody may be administered to a subject over a defined time course.
  • the methods according to this aspect of the invention comprise sequentially administering to a subject multiple doses of an anti-SARS-COV-2 antibody of the invention.
  • sequentially administering means that each dose of anti- SARS-COV-2 antibody is administered to the subject at a different point in time, e.g., on different days separated by a predetermined interval (e.g., hours, days, weeks, or months).
  • the present invention includes methods that comprise sequentially administering to the patient a single initial dose of an anti- SARS-COV-2 antibody, followed by one or more secondary doses of the anti-SARS-COV-2 antibody, and optionally followed by one or more tertiary doses of the anti-SARS-COV-2 antibody.
  • the terms “initial dose,” “secondary doses,” and “tertiary doses,” refer to the temporal sequence of administration of the anti-SARS-COV-2 antibody of the invention.
  • the “initial dose” is the dose, which is administered at the beginning of the treatment regimen (also referred to as the “baseline dose”);
  • the “secondary doses” are the doses, which are administered after the initial dose;
  • the “tertiary doses” are the doses which are administered after the secondary doses.
  • the initial, secondary, and tertiary doses may all contain the same amount of anti-SARS-COV-2 antibody, but generally may differ from one another in terms of frequency of administration.
  • the amount of anti-SARS-COV-2 antibody contained in the initial, secondary and/or tertiary doses varies from one another (e.g., adjusted up or down as appropriate) during the course of treatment.
  • two or more (e.g., 2, 3, 4, or 5) doses are administered at the beginning of the treatment regimen as “loading doses” followed by subsequent doses that are administered on a less frequent basis (e.g., “maintenance doses”).
  • each secondary and/or tertiary dose is administered 1 to 48 hours (e.g., 1, 1 1 ⁇ 2, 2, 21 ⁇ 2, 3, 31 ⁇ 2, 4, 41 ⁇ 2, 5, 51 ⁇ 2, 6, 61 ⁇ 2, 7, 71 ⁇ 2, 8, 81 ⁇ 2, 9, 91 ⁇ 2, 10, 101 ⁇ 2, 11, 11 1 ⁇ 2, 12, 121 ⁇ 2, 13, 131 ⁇ 2, 14, 141 ⁇ 2, 15, 151 ⁇ 2, 16, 161 ⁇ 2, 17, 171 ⁇ 2, 18, 181 ⁇ 2, 19, 191 ⁇ 2, 20, 201 ⁇ 2, 21, 21 1 ⁇ 2, 22, 221 ⁇ 2, 23, 23 1 ⁇ 2, 24, 241 ⁇ 2, 25, 25 1 ⁇ 2, 26, 261 ⁇ 2, or more) after the immediately preceding dose.
  • 1 to 48 hours e.g., 1, 1 1 ⁇ 2, 2, 21 ⁇ 2, 3, 31 ⁇ 2, 4, 41 ⁇ 2, 5, 51 ⁇ 2, 6, 61 ⁇ 2, 7, 71 ⁇ 2, 8, 81 ⁇ 2, 9, 91 ⁇ 2, 10, 101 ⁇ 2, 11, 11 1 ⁇ 2, 12, 121 ⁇ 2, 13, 131 ⁇ 2, 14, 141 ⁇ 2, 15, 151 ⁇ 2, 16, 161 ⁇ 2, 17, 171 ⁇ 2, 18, 181 ⁇
  • the immediately preceding dose means, in a sequence of multiple administrations, the dose of anti-SARS-COV-2 antibody, which is administered to a patient prior to the administration of the very next dose in the sequence with no intervening doses.
  • the methods may comprise administering to a patient any number of secondary and/or tertiary doses of an anti-SARS-COV-2 antibody.
  • any number of secondary and/or tertiary doses of an anti-SARS-COV-2 antibody may comprise administering to a patient any number of secondary and/or tertiary doses of an anti-SARS-COV-2 antibody.
  • only a single secondary dose is administered to the patient.
  • two or more (e.g., 2, 3, 4, 5, 6, 7, 8, or more) secondary doses are administered to the patient.
  • only a single tertiary dose is administered to the patient.
  • two or more (e.g., 2, 3, 4, 5, 6, 7, 8, or more) tertiary doses are administered to the patient.
  • the frequency at which the secondary and/or tertiary doses are administered to a patient can vary over the course of the treatment regimen. The frequency of administration may also be adjusted during the course of treatment by a physician depending on the needs of the individual patient following clinical examination. Diagnostic Uses of the Antibodies [00421]
  • the anti-SARS-COV-2 antibodies of the present invention may be used to detect and/or measure SARS-COV-2 in a sample, e.g., for diagnostic purposes.
  • Some embodiments contemplate the use of one or more antibodies of the present invention in assays to detect a SARS-COV-2- associated disease or disorder.
  • Exemplary diagnostic assays for SARS-COV-2 may comprise, e.g., contacting a sample obtained from a patient with an anti-SARS-COV-2 antibody of this disclosure, wherein the anti-SARS-COV-2 antibody is labeled with a detectable label or reporter molecule or used as a capture ligand to selectively isolate SARS-COV-2 from patient samples.
  • an unlabeled anti-SARS-COV-2 antibody can be used in diagnostic applications in combination with a secondary antibody, which is itself detectably labeled.
  • the detectable label or reporter molecule can be a radioisotope, such as H, C, P, S, or I: a fluorescent or chemiluminescent moiety such as fluorescein isothiocyanate, or rhodamine; or an enzyme such as alkaline phosphatase, [3- galactosidase, horseradish peroxidase, or luciferase.
  • a radioisotope such as H, C, P, S, or I
  • a fluorescent or chemiluminescent moiety such as fluorescein isothiocyanate, or rhodamine
  • an enzyme such as alkaline phosphatase, [3- galactosidase, horseradish peroxidase, or luciferase.
  • Specific exemplary assays that can be used to detect or measure SARS-COV-2 in a sample include enzyme- linked immunosorbent assay (ELISA), radioimmunoassay (RIA),
  • this disclosure further provides a method for detecting the presence of SARS CoV-2 in a sample comprising the steps of: (i) contacting a sample with the antibody or antigen-binding fragment thereof described herein; and (ii) determining binding of the antibody or antigen-binding fragment to one or more SARS CoV-2 antigens, wherein binding of the antibody to the one or more SARS CoV-2 antigens is indicative of the presence of SARS CoV-2 in the sample.
  • the SARS-CoV-2 antigen comprises the receptor binding domain (RBD) of the S polypeptide.
  • the SARS-CoV-2 antigen comprises the N-terminal domain (NTD) of the S polypeptide.
  • the SARS-CoV-2 antigen comprises the supersite b-hairpin (residues 152-158), the
  • the SARS-CoV-2 antigen comprises residues 27-32, 57-60, 210-218, and/or 286-303 of the S polypeptide.
  • the SARS-CoV-2 antigen comprises residues 600-606 of the S polypeptide.
  • the antibody or antigen-binding fragment thereof is conjugated to a label.
  • the step of detecting comprises contacting a secondary' antibody with the antibody or antigen-binding fragment thereof and wherein the secondary/ antibody comprises a label.
  • the label includes a fluorescent label, a chemiluminescent label, a radiolabel, and an enzyme.
  • the step of detecting comprises detecting fluorescence or chemiluminescence. In some embodiments, the step of detecting comprises a competitive binding assay or ELISA.
  • the method further comprises binding the sample to a solid support.
  • the solid support includes microparticles, microbeads, magnetic beads, and an affinity purification column.
  • Samples that can be used in SARS-COV-2 diagnostic assays according to the present invention include any tissue or fluid sample obtainable from a patient, which contains detectable quantities of either SARS-COV-2 protein, or fragments thereof, under normal or pathological conditions. Generally, levels of SARS-COV-2 protein in a particular sample obtained from a healthy patient (e.g., a patient not afflicted with a disease associated with SARS-COV-2) will be measured to initially establish a baseline, or standard, level of SARS-COV-2.
  • the antibodies specific for SARS-COV-2 protein may contain no additional labels or moieties, or they may contain an N-terminal or C-terminal label or moiety.
  • the label or moiety is biotin.
  • the location of a label may determine the orientation of the peptide relative to the surface upon which the peptide is bound.
  • Embodiment 1 An isolated anti-SARS-CoV-2 antibody or antigen-binding fragment thereof that binds specifically to a SARS-CoV-2 antigen.
  • Embodiment 1 The antibody or antigen-binding fragment thereof of Embodiment 1, wherein the SARS-CoV-2 antigen comprises a spike polypeptide, and preferably wherein the SARS-CoV-2 antigen comprises a spike polypeptide of a human or an animal SARS-CoV-2.
  • Embodiment 3 The antibody or antigen-binding fragment thereof of any one of the preceding Embodiments, wherein the SARS-CoV-2 antigen comprises the receptor binding domain (RBD) or the N-terminal domain (NTD) of the spike polypeptide.
  • the SARS-CoV-2 antigen comprises the supersite b-hairpin (residues 152- ⁇ WKH ⁇ -strand (residue 97-102), N4-loop (residues 178-188), and/or N-linked glycans at positions N122 and N149 of the spike polypeptide; or (b) wherein the SARS-CoV-2 antigen comprises residues 27-32, 57-60, 210-218, and/or 286-303 of the spike polypeptide, and preferably wherein the SARS-CoV-2 antigen comprises residues 600-606 of the spike polypeptide.
  • Embodiment 5 The antibody or antigen-binding fragment thereof of any one of the preceding Embodiments, comprising: a. three heavy chain complementarity determining regions (HCDRs) (HCDR1, HCDR2, and HCDR3) of a heavy chain variable region (HCVR) having the amino acid sequence of SEQ ID NO: 25; and three light chain CDRs (LCDR1, LCDR2, and LCDR3) of a light chain variable region (LCVR) having the amino acid sequence of SEQ ID NO: 26; b.
  • HCDRs heavy chain complementarity determining regions
  • LCVR heavy chain variable region having the amino acid sequence of SEQ ID NO: 25
  • LCDR1, LCDR2, and LCDR3 three light chain CDRs
  • Embodiment 6 The antibody or antigen-binding fragment thereof of any one of the preceding Embodiments, comprising: a.
  • a HCVR comprising an amino acid sequence with at least 75% identity to the amino acid sequence of SEQ ID NO: 25 or comprising the amino acid sequence of SEQ ID NO: 25; and a LCVR comprising an amino acid sequence with at least 75% identity to the amino acid sequence of SEQ ID NO: 26 or comprising the amino acid sequence of SEQ ID NO: 26; b. a HCVR comprising an amino acid sequence with at least 75% identity to the amino acid sequence of SEQ ID NO: 191 or comprising the amino acid sequence of SEQ ID NO: 191; and a LCVR comprising an amino acid sequence with at least 75% identity to the amino acid sequence of SEQ ID NO: 192 or comprising the amino acid sequence of SEQ ID NO: 192; c.
  • a HCVR comprising an amino acid sequence with at least 75% identity to the amino acid sequence of SEQ ID NO: 219 or comprising the amino acid sequence of SEQ ID NO: 219; and a LCVR comprising an amino acid sequence with at least 75% identity to the amino acid sequence of SEQ ID NO: 220 or comprising the amino acid sequence of SEQ ID NO: 220; d.
  • a HCVR comprising an amino acid sequence with at least 75% identity to the amino acid sequence of SEQ ID NO: 395 or comprising the amino acid sequence of SEQ ID NO: 395; and a LCVR comprising an amino acid sequence with at least 75% identity to the amino acid sequence of SEQ ID NO; 396 or comprising the amino acid sequence of SEQ ID NO: 396; or e. a HCVR comprising an amino acid sequence with at least 75% identity to the amino acid sequence of SEQ ID NO: 397 or comprising the amino acid sequence of SEQ ID NO: 397; and a LCVR comprising an amino acid sequence with at least 75% identity to the amino acid sequence of SEQ ID NO; 398 or comprising the amino acid sequence of SEQ ID NO: 398.
  • Embodiment 7 The antibody or antigen-binding fragment thereof of any one of the preceding Embodiments, comprising a HCVR and a LCVR that comprise a HCVR/LCVR amino acid sequence pair of SEQ ID NOs: 25-26, 191-192, 219-220, 395-396, or 397-398.
  • Embodiment 9 The antibody or antigen-binding fragment thereof of any one of the preceding Embodiments, wherein the antibody or antigen-binding fragment thereof is capable of neutralizing a plurality of SARS-CoV-2 strains.
  • Embodiment 11 The antibody or antigen-binding fragment thereof of Embodiment 10, wherein the multivalent antibody is a bivalent or bispecific antibody.
  • Embodiment 13 The antibody or antigen-binding fragment thereof of any one of the preceding Embodiments, wherein the antibody is a monoclonal antibody.
  • Embodiment 14 The antibody or antigen-binding fragment thereof of any one of the preceding Embodiments, wherein the antibody or antigen-binding fragment thereof is a chimeric antibody, a humanized antibody, or humanized monoclonal antibody.
  • Embodiment 16 The antibody or antigen-binding fragment thereof of any one of the preceding Embodiments, wherein the antibody or antigen-binding fragment thereof is detectably labeled or conjugated to a toxin, a therapeutic agent, a polymer, a receptor, an enzyme, or a receptor ligand, preferably wherein the polymer is polyethylene glycol (PEG).
  • a pharmaceutical composition comprising the antibody or antigen- binding fragment thereof of any one of the preceding Embodiments and optionally a pharmaceutically acceptable carrier or excipient.
  • Embodiment 18 The pharmaceutical composition of Embodiment 16, wherein the pharmaceutical comprises two or more of the antibody or antigen-binding fragment thereof of any one of Embodiments 1 to 16.
  • Embodiment 19 The pharmaceutical composition of Embodiment 18, wherein the two or more of the antibody or antigen-binding fragment thereof comprise: a. a first antibody or antigen-binding fragment thereof comprising a HCVR/LCVR amino acid sequence pair of SEQ ID NOs: 25-26, 191-192, 219-220, 395-396, or 397-398, b.
  • Embodiment 20 The pharmaceutical composition of Embodiment 18, wherein the two or more of the antibody or antigen-binding fragment thereof comprise: a. a first antibody or antigen-binding fragment thereof comprising a HCVR/LCVR amino acid sequence pair with at least 75% identity to the amino acid sequence of SEQ ID NOs: 25-26, 191-192, 219-220, 395-396, or 397-398, b. wherein the first antibody or antigen-binding fragment thereof is different from the second antibody or antigen-binding fragment thereof.
  • Embodiment 21 Embodiment 21.
  • Embodiment 22 The pharmaceutical composition of Embodiment 21, wherein: (a) the antiviral compound comprises: a nucleoside analog, a peptoid, an oligopeptide, a polypeptide, a protease inhibitor, a 3C-like protease inhibitor, a papain-like protease inhibitor, or an inhibitor of an RNA dependent RNA polymerase; or (b) the antiviral compound is selected from: acyclovir, ganciclovir, vidarabine, foscarnet, cidofovir, amantadine, ribavirin, trifluorothymidine, zidovudine, didanosine, zalcitabine, and an interferon, and preferably wherein the interferon is an interferon- ⁇ or an interferon- ⁇ or an interferon
  • Embodiment 24 A nucleic acid molecule encoding a polypeptide chain of the antibody or antigen-binding fragment thereof of any one of Embodiments 1 to 16.
  • Embodiment 25 A vector comprising the nucleic acid molecule of Embodiment 24.
  • Embodiment 26 A cultured host cell comprising the vector of Embodiment 25.
  • Embodiment 27 A cultured host cell comprising the vector of Embodiment 27.
  • Embodiment 28 A kit comprising a pharmaceutically acceptable dose unit of the antibody or antigen-binding fragment thereof of any one of Embodiments 1 to 16 or the pharmaceutical composition of any one of Embodiments 17 to 22. [00459] Embodiment 29.
  • kits for the diagnosis, prognosis or monitoring treatment of SARS- CoV-2 in a subject comprising: the antibody or antigen-binding fragment thereof of any one of Embodiments 1 to 16; and a least one detection reagent that binds specifically to the antibody or antigen-binding fragment thereof.
  • Embodiment 30 A method of neutralizing SARS-CoV-2 in a subject, comprising administering to a subject in need thereof a therapeutically effective amount of the antibody or antigen-binding fragment thereof of any one of Embodiments 1 to 16 or a therapeutically effective amount of the pharmaceutical composition of any one of Embodiments 17 to 22.
  • Embodiment 31 Embodiment 31.
  • a method of preventing or treating a SARS-CoV-2 infection comprising administering to a subject in need thereof a therapeutically effective amount of the antibody or antigen-binding fragment thereof of any one of Embodiments 1 to 16 or a therapeutically effective amount of the pharmaceutical composition of any one of Embodiments 17 to 22.
  • a therapeutically effective amount of the antibody or antigen-binding fragment thereof of any one of Embodiments 1 to 16 or a therapeutically effective amount of the pharmaceutical composition of any one of Embodiments 17 to 22 [00462] Embodiment 32.
  • a method of neutralizing SARS-CoV-2 in a subject comprising administering to a subject in need thereof a therapeutically effective amount of a first antibody or antigen-binding fragment thereof and a second antibody or antigen-binding fragment thereof of any one of Embodiments 1 to 16 or a therapeutically effective amount of the pharmaceutical composition of any one of Embodiments 17 to 22, wherein the first antibody or antigen-binding fragment thereof and the second antibody or antigen-binding fragment thereof exhibit synergistic activity.
  • a method of preventing or treating a SARS-CoV-2 infection comprising administering to a subject in need thereof a therapeutically effective amount of a first antibody or antigen-binding fragment thereof of any one of Embodiments 1 to 16 and a second antibody or antigen-binding fragment thereof of any one of Embodiments 1 to 16 or a therapeutically effective amount of the pharmaceutical composition of any one of Embodiments 17 to 22, wherein the first antibody or antigen-binding fragment thereof and the second antibody or antigen-binding fragment thereof exhibit synergistic activity.
  • Embodiment 34 The method of Embodiment 30 or 31, further comprising administering to the subject a therapeutically effective amount of a second therapeutic agent or therapy.
  • Embodiment 35 Embodiment 35.
  • Embodiment 34 wherein the antibody or antigen- binding fragment thereof comprises a heavy chain variable region and a light chain variable region comprising a HCVR/LCVR amino acid sequence pair of SEQ ID NOs: 25-26, 191-192, 219-220, 395-396, or 397-398, wherein the antibody or antigen-binding fragment thereof is different from the second therapeutic agent or therapy.
  • Embodiment 36 Embodiment 36.
  • Embodiment 34 wherein the antibody or antigen- binding fragment thereof comprises a heavy chain variable region and a light chain variable region comprising a HCVR/LCVR amino acid sequence pair with at least 75% identity to the amino acid sequence of SEQ ID NOs: 25-26, 191-192, 219-220, 395-396, or 397-398, wherein the antibody or antigen-binding fragment thereof is different from the second therapeutic agent or therapy.
  • Embodiment 37 The method of Embodiment 34 or 35, wherein the antibody or antigen- binding fragment thereof is administered before, after, or concurrently with the second therapeutic agent or therapy.
  • Embodiment 38 Embodiment 38.
  • Embodiment 39 The method of Embodiment 38, wherein: (a) the antiviral compound comprises: a nucleoside analog, a peptoid, an oligopeptide, a polypeptide, a protease inhibitor, a 3C-like protease inhibitor, a papain-like protease inhibitor, or an inhibitor of an RNA dependent RNA polymerase; or (b) the antiviral compound is selected from: acyclovir, ganciclovir, vidarabine, foscarnet, cidofovir, amantadine, ribavirin, trifluorothymidine, zidovudine, didanosine, zalcitabine, and an interferon, and preferably wherein the interferon is an interferon- ⁇ RU ⁇ DQ ⁇ interferon- ⁇ [00
  • Embodiment 41 comprises a heavy chain variable region and a light chain variable region comprising a HCVR/LCVR amino acid sequence pair of SEQ ID NOs: 25-26, 191- 192, 219-220, 395-396, or 397-398, wherein the first antibody or antigen-binding fragment thereof is different from the second antibody or antigen-binding fragment thereof.
  • Embodiment 32 or 33 wherein the first antibody or antigen-binding fragment thereof comprises a heavy chain variable region and a light chain variable region comprising a HCVR/LCVR amino acid sequence pair with at least 75% identity to the amino acid sequence of SEQ ID NOs: 25-26, 191-192, 219-220, 395-396, or 397-398, wherein the first antibody or antigen-binding fragment thereof is different from the second antibody or antigen-binding fragment thereof.
  • Embodiment 42 The method of any one of Embodiments 32,33 and 40, wherein the first antibody or antigen-binding fragment thereof is administered before, after, or concurrently with the second antibody or antigen-binding fragment thereof.
  • Embodiment 43 The method of any one of Embodiments 30 to 42, wherein the antibody or antigen-binding fragment thereof is administered to the subject intravenously, subcutaneously, or intraperitoneally.
  • Embodiment 44 The method of Embodiment 32 or 33, wherein the antibody or antigen- binding fragment thereof is administered prophylactically or therapeutically.
  • Embodiment 45 A method for detecting the presence of SARS CoV-2 in a sample comprising the steps of: a. contacting a sample with the antibody or antigen-binding fragment thereof of any one of Embodiments 1 to 16; and b.
  • Embodiment 46 The method of Embodiment 45, wherein the SARS-CoV-2 antigen comprises a spike polypeptide of a human or an animal SARS-CoV-2, and preferably wherein the SARS-CoV-2 antigen comprises the receptor binding domain (RBD) or the N-terminal domain (NTD) of the S polypeptide.
  • Embodiment 47 The method of Embodiment 45, wherein the SARS-CoV-2 antigen comprises a spike polypeptide of a human or an animal SARS-CoV-2, and preferably wherein the SARS-CoV-2 antigen comprises the receptor binding domain (RBD) or the N-terminal domain (NTD) of the S polypeptide.
  • Embodiment 48 The method of any one of Embodiments 45 to 47, further comprising contacting a secondary antibody with the antibody or antigen-binding fragment thereof, wherein the secondary antibody comprises a label.
  • Embodiment 49 The method of Embodiment 48, further comprising detecting fluorescence or chemiluminescence of the label, preferably wherein the step of detecting comprises a competitive binding assay or ELISA.
  • Embodiment 50 The method of any one of Embodiments 45 to 49, wherein the sample is a blood sample.
  • Embodiment 51 The method of any one of Embodiment 45 to 50, further comprising binding the sample to a solid support, preferably wherein the solid support is selected from microparticles, microbeads, magnetic beads, and an affinity purification column.
  • PBMCs Peripheral Blood Mononuclear Cells obtained from samples collected were further purified as previously reported by gradient centrifugation and stored in liquid nitrogen in the presence of Fetal Calf Serum (FCS) and Dimethylsulfoxide (DMSO) (Gaebler et al., 2021; Robbiani et al., 2020). Heparinized serum and plasma samples were aliquoted and stored at -20°C or less. Prior to experiments, aliquots of plasma samples were heat-inactivated (56°C for 1 hour) and then stored at 4°C.
  • FCS Fetal Calf Serum
  • DMSO Dimethylsulfoxide
  • Enzyme-Linked Immunosorbent Assays (ELISAs)(Amanat et al., 2020; Grifoni. et al,, 2020) were performed to evaluate antibodies binding to SARS-CoV-2 wild-type (Wuhan-Hu- 1) RBD, and variants of concern Delta (B.1 ,617.2) RBD, and Omicron (BA.1) spike protein by coating of high-binding 96-half-well plates (Corning 3690) with 50 ⁇ l per well of a 1 ug/ml indicated protein solution in Phosphate-buffered Saline (PBS) overnight at 4°C.
  • ELISAs Enzyme-Linked Immunosorbent Assays
  • EC50 ELISA half-maximal concentration
  • Omicron BA.l A67V, A69-70, T95I, G142D, A143-145, A211, L212I, ins214EPE, G339D, S371L, S373P, S375F, K417N, N440K, G446S, S477N, T478K, E484A, Q493K, G496S, Q498R, N501Y, Y505H, T547K, D614G, H655Y, H679K, P681H, N764K, D796Y, N856K, Q954H, N969H, N969K, L981F
  • Omicron BA.2 T19I, L24S, de!25-27, G142D, V213G, G339D, S371F, S373P, S375F, T376A, D405N, R408S, K417N, N440K, S477N, T478K, E484A, Q493R, Q498R, N501Y, Y5O5H, D614G, H655Y, N679K, P681H, N764K, D796Y, Q954H, N969K [00493] Omicron BA.4/5: T19I, L24S, del25-27, del69-70, G142D, V213G, G339D, S371F, S373P, S375F, T376A, D405N, R408S, K417N, N440K, L452R, S477N, T478K, E48
  • SARS-CoV-2 pseudotyped particles were generated as previously described (Robbiani et al., 2020; Schmidt et al., 2020a). Briefly, 293T (CRL-11268) cells were obtained from ATCC, and the cells were transfected with pNL4- ⁇ (QY-nanoluc and pSARS-CoV-2-S ⁇ . Particles were harvested 48 hours post-transfection, filtered, and stored at -80°C.
  • Pseudotyped virus neutralization assay Pre-pandemic negative control plasma from healthy donors, plasma from individuals who received mRNA vaccines and had Delta or Omicron BA.1 breakthrough infection, or monoclonal antibodies were five-fold serially diluted and incubated with SARS-CoV-2 pseudotyped virus for 1 hour at 37 °C. The mixture was subsequently incubated with 293TAce2 cells (Robbiani et al., 2020) (for all WT neutralization assays) or HT1080/Ace2 cl14 cells (for all variant neutralization assays) for 48 hours, after which cells were washed with PBS and lysed with Luciferase Cell Culture Lysis 5 ⁇ reagent (Promega).
  • Nanoluc Luciferase activity in lysates was measured using the Nano-Glo Luciferase Assay System (Promega) with the ClarioStar Microplate Multimode Reader (BMG).
  • BMG ClarioStar Microplate Multimode Reader
  • the relative luminescence units were normalized to those derived from cells infected with SARS-CoV-2 pseudotyped virus(Wang et al., 2021c) in the absence of plasma or monoclonal antibodies.
  • Biotinylation of viral protein for use in flow cytometry [00497] Purified and Avi-tagged SARS-CoV-2 WT and Delta RBD were biotinylated using the Biotin-Protein Ligase-BIRA kit according to manufacturer’s instructions (Avidity) as described before (Robbiani et al., 2020). Ovalbumin (Sigma, A5503-1G) was biotinylated using the EZ-Link Sulfo-NHS-LC-Biotinylation kit according to the manufacturer’s instructions (Thermo Scientific).
  • Biotinylated ovalbumin was conjugated to streptavidin-BV711 for single-cell sorts (BD biosciences, 563262) or to streptavidin-BB515 for phenotyping panel (BD, 564453).
  • WT RBD was conjugated to streptavidin-PE (BD Biosciences, 554061) and streptavidin-AF647 (Biolegend, 405237) for single-cell sorts, or streptavidin- BV421 (Biolegend, 405225) and streptavidin-BV711 (BD biosciences, 563262) for phenotyping.
  • Delta RBD was conjugated to streptavidin-PE (BD Biosciences, 554061) and Omicron BA. 1 RBD (ACROBiosystems, SPD-C82E4) was conjugated to streptavidin-AF647 (Biolegend, 405237).
  • PBMC peripheral blood mononuclear cells
  • the enriched B cells were incubated in Fluorescence- Activated Cell-sorting (FACS) buffer (1 x PBS, 2% FCS, 1 mM ethylenediaminetetraacetic acid (EDTA)) with the following anti-human antibodies (all at 1 :200 dilution): anti-CD20-PECy7 (BD Biosciences, 335793), anti-CD3-APC-eFluro 780 (Invitrogen, 47-0037-41), anti-CD8-APC-eFluor 780 (Invitrogen, 47-0086-42), anti-CDl 6-APC-eFluor 780 (Invitrogen, 47-0168-41), anti-CDl 4-APC- eFluor 780 (Invitrogen, 47-0149-42), as well as Zombie NIR (BioLegend, 423105) and fluorophore-labeled RBD and ovalbumin (Ova) for 30 min on ice.
  • B cells were also stained with the following anti-human antibodies (all at 1:200 dilution): anti-IgD-BV650 (BD, 740594), anti- CD27-BV786 (BD biosciences, 563327), anti-CDl 9-BV605 (Biolegend, 302244), anti-CD71 - PerCP-Cy5.5 (Biolegend, 334114), anti-IgG-PECF594 (BD, 562538), anti-IgM-AF700 (Biolegend, 314538), anti-IgA-Viogreen (Miltenyi Biotec, 130-113-481).
  • anti-human antibodies all at 1:200 dilution
  • Antibody sequencing, cloning, and expression [00499] Antibodies were identified and sequenced as described previously (Robbiani et aL, 2020; Wang et aL, 2021a). In brief, RNA from single cells was reverse-transcribed (SuperScript III Reverse Transcriptase, Invitrogen, 18080-044), and the cDNA was stored at ---20 °C or used for subsequent amplification of the variable IGH, IGL, and IGK genes by nested PCR and Sanger sequencing. Sequence analysis was performed using Mac Vector. Amplicons from the first PCR reaction were used as templates for sequence- and ligation-independent cloning into antibody expression vectors. Recombinant monoclonal antibodies were produced and purified as previously described (Robbiani et al., 2020).
  • the heavy and light chain fragments were amplified using primers that reached into the framework region of the antibodies.
  • the kappa primer introduces non-native ammo acids into the kappa chain.
  • the heavy chain was truncated at the C-termmus for the purpose of antibody production, being one amino acid shorter than what usually is present in IgG heavy chain constant regions.
  • Biolayer interferometry assays were performed as previously described (Robbiani et al,, 2020). In brief, the Octet Red instrument (ForteBio) was used at 30 °C with shaking at 1 ,000 r.p.m.
  • Epitope binding assays were performed with protein A biosensor (ForteBio 18-5010), following the manufacturer’s protocol “classical sandwich assay” as follows: (1 ) Sensor check: sensors immersed 30 sec in buffer alone (buffer ForteBio 18-1105), (2) Capture 1st Ab: sensors immersed 10 mm with Abl at 10 ⁇ g/mL, (3) Baseline: sensors immersed 30 sec in buffer alone, (4) Blocking: sensors immersed 5 min with IgG isotype control at 10 ⁇ g/mL. (5) Baseline: sensors immersed 30 sec in buffer alone, (6) Antigen association: sensors immersed 5 min with RBD at 10 ⁇ g/mL. (7) Baseline: sensors immersed 30 sec in buffer alone.
  • Association Ab2 sensors immersed 5 min with Ab2 at 10 ⁇ g/mL. Curve fitting was performed using the Fortebio Octet Data analysis software (ForteBio). Affinity measurements of anti-SARS-CoV-2 IgGs binding were corrected by subtracting the signal obtained from traces performed with IgGs in the absence of WT RBD.
  • the kinetic analysis using protein A biosensor was performed as follows: (1) baseline: 60 sec immersion in buffer. (2) loading: 200 sec immersion in a solution with IgGs 10 ⁇ g/ml. (3) baseline: 200 sec immersion in buffer. (4) Association: 300 sec immersion in solution with WT RBD at 20, 10 or 5 ⁇ g/ml (5) dissociation: 600 sec immersion in buffer.
  • Curve fitting was performed using a fast 1:1 binding model and the Data analysis software (ForteBio). Mean KD values were determined by averaging all binding curves that matched the theoretical fit with an R2 value ⁇ 0.8.
  • Computational analyses of antibody sequences [00502] Antibody sequences were trimmed based on quality and annotated using Igblastn v.1.14. with IMGT domain delineation system. Annotation was performed systematically using Change- O toolkit v.0.4.540 (Gupta et al., 2015). Clonality of heavy and light chains was determined using DefineClones.py implemented by Change-O v0.4.5 (Gupta et al., 2015).
  • the script calculates the Hamming distance between each sequence in the data set and its nearest neighbor. Distances are subsequently normalized and to account for differences in junction sequence length, and clonality is determined based on a cut-off threshold of 0.15. Heavy and light chains derived from the same cell were subsequently paired, and clonotypes were assigned based on their V and J genes using in-house R and Perl scripts. All scripts and the data used to process antibody sequences are publicly available on GitHub (github.com/stratust/igpipeline/tree/igpipeline2_timepoint_v2).
  • the two- sided binomial test was used to check whether the number of sequences belonging to a specific IGHV or IGLV gene in the repertoire is different according to the frequency of the same IgV gene in the database. Adjusted p-values were calculated using the false discovery rate (FDR) correction. Significant differences are denoted with stars.
  • FDR false discovery rate
  • Nucleotide somatic hypermutation and Complementarity-Determining Region (CDR3) length were determined using in-house R and Perl scripts. For somatic hypermutations, IGHV and IGLV nucleotide sequences were aligned against their closest germlines using Igblastn, and the number of differences was considered to correspond to nucleotide mutations.
  • Anti-WT-RBD IgG titers were significantly increased after Delta breakthrough infection in individuals who received 2 doses of mRNA vaccine (Delta BT), compared to vaccinated individuals who did not experience infection (5m-Vax2) (p ⁇ 0.0001, Vax2 (Cho et al., 2021) vs. Delta, FIG. 1b and Table 1).
  • Plasma neutralizing activity in 49 participants was measured using HIV-1 pseudotyped with the WT SARS-CoV-2 spike protein (Cho et al., 2021; Wang et al., 2021c) (FIG.1c and Table 1).
  • NT50 geometric mean half-maximal neutralizing titer
  • Plasma neutralizing activity was also assessed against SARS-CoV-2 Delta, Omicron BA.1, BA.2, and BA.4/5 variants using viruses pseudotyped with appropriate variant spike proteins.
  • Memory B cells [00510] mRNA vaccines elicit memory B cells (MBCs) that can contribute to durable immune protection from serious disease by mediating rapid and anamnestic antibody response (Muecksch et al., 2022; Victora and Nussenzweig, 2022).
  • RBD-specific MBCs were enumerated using Alexa Fluor 647 (AF647)- and phycoerythrin (PE)-labeled WT RBD of the SARS-CoV-2 spike protein by flow cytometry (FIG. 2a, FIG. 5).
  • FIG.2d Clonally expanded WT-RBD-specific B cells represented 9% of all memory B cells after Delta breakthrough infection and 28% of the repertoire after Omicron BA.1 breakthrough infection (FIG. 2d). Similar to mRNA vaccinees (Cho et al., 2021; Muecksch et al., 2022; Wang et al., 2021c), several sets of VH genes including VH3-30 and VH3-53 were over-represented in Delta- or Omicron BA.1 breakthrough infection (FIGs. 6a-f).
  • VH3-49, VH4-38, and VH1-24 were exclusively over-represented after Delta breakthrough infection (FIG. 6a), while VH1-69, VH1-58, VH4-61, and VH4-38 were specifically over-represented after Omicron BA.1 breakthrough infection (FIG. 6d).
  • VH1-69, VH1-58, VH4-61, and VH4-38 were specifically over-represented after Omicron BA.1 breakthrough infection (FIG. 6d).
  • Anti-RBD antibodies elicited by mRNA vaccination target 4 structurally defined classes of epitopes on the SARS-CoV-2 RBD (Barnes et al., 2020a; Cho et al., 2021; Muecksch et al., 2022; Yuan et al., 2020).
  • SARS-CoV-2 RBD structurally defined classes of epitopes on the SARS-CoV-2 RBD (Barnes et al., 2020a; Cho et al., 2021; Muecksch et al., 2022; Yuan et al., 2020).
  • BLI competition experiments were performed.
  • a preformed antibody-RBD-complex was exposed to a second antibody recognizing one of four classes of structurally defined antigenic sites (C105 as Class 1; C144 as Class 2, C135 as Class 3, and C2172 as Class 4 (Barnes et al., 2020a)).
  • IC50s lower than 1000 ng/mL (neutralizing) against WT
  • Vax3 antibodies and Omicron BA.1 breakthrough antibodies were enriched for those neutralizing BA.4/5 with IC50 values of less than 1000 ng/ml with 37% and 38% of all tested antibodies neutralizing BA.4/5, respectively, while only 17% and 27% of Vax2 and Delta breakthrough antibodies, respectively neutralized BA.4/5 (FIG. 3f).
  • IC50 values less than 1000 ng/ml
  • Vax2 and Delta breakthrough antibodies respectively neutralized BA.4/5
  • FIG. 3f Vax3 antibodies and Omicron BA.1 breakthrough antibodies
  • the memory antibodies that develop after a 3rd or 4th antigenic exposure by Delta or Omicron BA.1 infection were examine, respectively.
  • a 3rd exposure to antigen by Delta breakthrough increases the number of memory B cells that produce antibodies with comparable potency and breadth to a 3rd mRNA vaccine dose.
  • a 4th antigenic exposure with Omicron BA.1 infection increased variant specific plasma antibody and memory B cell responses.
  • the 4th exposure did not increase the overall frequency of memory B cells or their general potency or breadth compared to a 3rd mRNA vaccine dose.
  • a 3rd antigenic exposure by Delta infection elicits strain-specific memory responses and increases in the overall potency and breadth of the memory B cells.
  • Omicron BA.1 the effects of a 4th antigenic exposure with Omicron BA.1 is limited to increased strain specific memory with little effect on the potency or breadth of memory B cell antibodies.
  • Omicron and its subvariants are reported to be more transmissible than any prior VoC and have spurred a resurgence of new cases worldwide (Mallapaty, 2022). While early reports suggested that Omicron may cause less severe illness, recent studies show variant-specific symptoms but similar virulence (Whitaker et al., 2022), and increased resistance to approved vaccine regimens (Nealon and Cowling, 2022).
  • a 3rd mRNA vaccine dose boosts plasma antibody responses to SARS-CoV-2 variants, including Omicron BA.1, and increases the number, potency, and breadth of the antibodies found in the memory B cell compartment (Goel et al., 2022; Muecksch et al., 2022). Although the antibodies in plasma are generally not sufficient to prevent breakthrough infection boosted individuals are protected against serious disease upon breakthrough infection (Kuhlmann et al., 2022; Nemet et al., 2022). The present study shows that a 3rd exposure to antigen in the form of Delta breakthrough infection produces similar effects on the overall size of the memory compartment to a 3rd mRNA vaccine dose, and specifically boosts strain-specific responses.
  • Zahradnik, P. Supasa C. Liu, H.M.E. Duyvesteyn, H.M. Ginn, A.J. Mentzer, A. Tuekprakhon, R. Nutalai, B. Wang, A. Dijokaite, S. Khan, O. Avinoam, M. Bahar, D. Skelly, S. Adele, S.A. Johnson, A. Amini, T.G. Ritter, C. Mason, C. Dold, D. Pan, S. Assadi, A. Bellass, N. Omo-Dare, D. Koeckerling, A. Flaxman, D. Jenkin, P.K. Aley, M. Voysey, S.A.
  • cAb-Rep a database of curated antibody repertoires for exploring antibody diversity and predicting antibody prevalence. Frontiers in immunology 2365. [00538] Gupta, N.T., J.A. Vander Heiden, M. Uduman, D. Gadala-Maria, G. Yaari, and S.H. Kleinstein. 2015. Change-O: a toolkit for analyzing large-scale B cell immunoglobulin repertoire sequencing data. Bioinformatics 31:3356-3358. [00539] Hachmann, N.P., J. Miller, A.-r.Y. Collier, J.D. Ventura, J. Yu, M. Rowe, E.A. Bondzie, O.
  • SARS-CoV-2 Omicron triggers cross-reactive neutralization and Fc effector functions in previously vaccinated, but not unvaccinated, individuals.

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Abstract

This disclosure provides novel neutralizing and potent anti-SARS-CoV-2 antibodies, related nucleic acids, related cells, related kits, related compositions, and related methods or uses.

Description

ANTI-SARS-COV-2 ANTIBODIES AND METHODS OF USE THEREOF CROSS-REFERENCE TO RELATED APPLICATIONS [0001] This application claims priority to U.S. Provisional Application No. 63/371,285, filed August 12, 2022, which is hereby incorporated by reference in its entirety. REFERENCE TO AN ELECTRONIC SEQUENCE LISTING [0002] The contents of the electronic sequence listing (070413.20758_SeqListing.xml; Size: 1,420,000 bytes; and Date of Creation: August 9, 2023) is herein incorporated by reference in its entirety. FIELD OF THE INVENTION [0003] The present invention relates to antibodies directed to epitopes of SARS-CoV-2 Coronavirus 2 (“SARS-CoV-2”). The present invention further relates to the preparation and use of neutralizing antibodies directed to the SARS-CoV-2 spike (S) glycoproteins for the prevention and treatment of SARS-CoV-2 infection. BACKGROUND [0004] Severe Acute Respiratory Syndrome Coronavirus (SARS-CoV-2) emerged in late 2019, causing a global pandemic with more than 500 million infections and over 6 million deaths reported to date. Over the course of the pandemic, SARS-COV-2 continued to evolve, resulting in substantial genetic distance between circulating variants and the initial viral sequence on which vaccines are based. Several of these circulating variants have been designated variants of concern (VoC) and have led to successive waves of infection, most notably by VoCs Alpha (Supasa et al., 2021), Delta (Liu et al., 2021), Omicron (Dejnirattisai et al., 2022). [0005] Higher rates of re-infection and vaccine-breakthrough infection with the Delta and Omicron variants highlighted the potential for immune escape from neutralizing antibody responses resulting in reduced vaccine efficacy against SARS-CoV-2 infection(Cao et al.; Cele et al., 2022; Dejnirattisai et al., 2022; Gaebler et al., 2022; Hachmann et al., 2022; Kuhlmann et al., 2022; Liu et al., 2021). With the emergence of Omicron BA.1 and related lineages, infection has surged worldwide, and these new variants account for over 95% of recent COVID-19 cases. To date, BA.2.12.1 variant (a BA.2 lineage) contributes 59% of new cases in the United States, while BA.4 and BA.5 caused a fifth wave of COVID-19 infection in South Africa. Nevertheless, vaccine- elicited immunity continues to provide robust protection against severe disease, even in the face of viral variants(Andrews et al., 2022; Madhi et al., 2022; Wolter et al., 2022; World Health, 2022). [0006] Previous studies have shown that Delta or Omicron breakthrough infection boosts plasma neutralizing activity against both the Wuhan-Hu-1 strain and the infecting variant, which might suggest recall responses of cross-reactive vaccine-induced memory B cells (Kaku et al., 2022; Quandt et al., 2022; Richardson et al., 2022; Seaman et al., 2022; Servellita et al., 2022). However, far less is known about the memory antibody responses after breakthrough infection. SUMMARY [0007] In some embodiments, the present disclosure provides an antibody or antigen-binding fragment thereof that binds to the S polypeptide of a Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) wherein the antibody or antigen-binding fragment thereof comprises: three heavy chain complementarity determining regions (HCDRs) (HCDR1, HCDR2, and HCDR3) of a heavy chain variable region (HCVR) having the amino acid sequence of SEQ ID NO: 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31, 33, 35, 37, 39, 41, 43, 45, 47, 49, 51, 53, 55, 57, 59, 61, 63, 65, 67, 69, 71, 73, 75, 77, 79, 81, 83, 85, 87, 89, 91, 93, 95, 97, 99, 101, 103, 105, 107, 109, 111, 113, 115, 117, 119, 121, 123, 125, 127, 129, 131, 133, 135, 137, 139, 141, 143, 145, 147, 149, 151, 153, 155, 157, 159, 161, 163, 165, 167, 169, 171, 173, 175, 177, 179, 181, 183, 185, 187, 189, 191, 193, 195, 197, 199, 201, 203, 205, 207, 209, 211, 213, 215, 217, 219, 221, 223, 225, 227, 229, 231, 233, 235, 237, 239, 241, 243, 245, 247, 249, 251, 253, 255, 257, 259, 261, 263, 265, 267, 269, 271, 273, 275, 277, 279, 281, 283, 285, 287, 289, 291, 293, 295, 297, 299, 301, 303, 305, 307, 309, 311, 313, 315, 317, 319, 321, 323, 325, 327, 329, 331, 333, 335, 337, 339, 341, 343, 345, 347, 349, 351, 353, 355, 357, 359, 361, 363, 365, 367, 369, 371, 373, 375, 377, 379, 381, 383, 385, 387, 389, 391, 393, 395, 397, 399, 401, 403, 405, 407, 409, 411, 413, 415, 417, 419, 421, 423, 425, 427, 429, 431, 433, 435, 437, 439, 441, 443, 445, 447, 449, 451, 453, 455, 457, 459, 461, 463, 465, 467, 469, 471, 473, 475, 477, 479, 481, 483, 485, 487, 489, 491, 493, 495, 497, 499, 501, 503, 505, 507, 509, 511, 513, 515, 517, 519, 521, 523, 525, 527, 529, 531, 533, 535, 537, 539, 541, 543, 545, 547, 549, 551, 553, 555, 557, 559, 561, 563, 565, 567, 569, 571, 573, 575, 577, 579, 581, 583, 585, 587, 589, 591, 593, 595, 597, 599, 601, 603, 605, 607, 609, 611, 613, 615, 617, 619, 621, 623, 625, 627, 629, 631, 633, 635, 637, 639, 641, 643, 645, 647, 649, 651, 653, 655, 657, 659, 661, 663, 665, 667, 669, 671, 673, or 675; and three light chain CDRs (LCDR1, LCDR2, and LCDR3) of a light chain variable region (LCVR) having the amino acid sequence of SEQ ID NO: 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 44, 46, 48, 50, 52, 54, 56, 58, 60, 62, 64, 66, 68, 70, 72, 74, 76, 78, 80, 82, 84, 86, 88, 90, 92, 94, 96, 98, 100, 102, 104, 106, 108, 110, 112, 114, 116, 118, 120, 122, 124, 126, 128, 130, 132, 134, 136, 138, 140, 142, 144, 146, 148, 150, 152, 154, 156, 158, 160, 162, 164, 166, 168, 170, 172, 174, 176, 178, 180, 182, 184, 186, 188, 190, 192, 194, 196, 198, 200, 202, 204, 206, 208, 210, 212, 214, 216, 218, 220, 222, 224, 226, 228, 230, 232, 234, 236, 238, 240, 242, 244, 246, 248, 250, 252, 254, 256, 258, 260, 262, 264, 266, 268, 270, 272, 274, 276, 278, 280, 282, 284, 286, 288, 290, 292, 294, 296, 298, 300, 302, 304, 306, 308, 310, 312, 314, 316, 318, 320, 322, 324, 326, 328, 330, 332, 334, 336, 338, 340, 342, 344, 346, 348, 350, 352, 354, 356, 358, 360, 362, 364, 366, 368, 370, 372, 374, 376, 378, 380, 382, 384, 386, 388, 390, 392, 394, 396, 398, 400, 402, 404, 406, 408, 410, 412, 414, 416, 418, 420, 422, 424, 426, 428, 430, 432, 434, 436, 438, 440, 442, 444, 446, 448, 450, 452, 454, 456, 458, 460, 462, 464, 466, 468, 470, 472, 474, 476, 478, 480, 482, 484, 486, 488, 490, 492, 494, 496, 498, 500, 502, 504, 506, 508, 510, 512, 514, 516, 518, 520, 522, 524, 526, 528, 530, 532, 534, 536, 538, 540, 542, 544, 546, 548, 550, 552, 554, 556, 558, 560, 562, 564, 566, 568, 570, 572, 574, 576, 578, 580, 582, 584, 586, 588, 590, 592, 594, 596, 598, 600, 602, 604, 606, 608, 610, 612, 614, 616, 618, 620, 622, 624, 626, 628, 630, 632, 634, 636, 638, 640, 642, 644, 646, 648, 650, 652, 654, 656, 658, 660, 662, 664, 666, 668, 670, 672, 674, or 676. [0008] In some embodiments, the antibody or antigen-binding fragment thereof comprises: a heavy chain variable region comprising an amino acid sequence with at least 75% identity to the amino acid sequence of SEQ ID NO: 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31, 33, 35, 37, 39, 41, 43, 45, 47, 49, 51, 53, 55, 57, 59, 61, 63, 65, 67, 69, 71, 73, 75, 77, 79, 81, 83, 85, 87, 89, 91, 93, 95, 97, 99, 101, 103, 105, 107, 109, 111, 113, 115, 117, 119, 121, 123, 125, 127, 129, 131, 133, 135, 137, 139, 141, 143, 145, 147, 149, 151, 153, 155, 157, 159, 161, 163, 165, 167, 169, 171, 173, 175, 177, 179, 181, 183, 185, 187, 189, 191, 193, 195, 197, 199, 201, 203, 205, 207, 209, 211, 213, 215, 217, 219, 221, 223, 225, 227, 229, 231, 233, 235, 237, 239, 241, 243, 245, 247, 249, 251, 253, 255, 257, 259, 261, 263, 265, 267, 269, 271, 273, 275, 277, 279, 281, 283, 285, 287, 289, 291, 293, 295, 297, 299, 301, 303, 305, 307, 309, 311, 313, 315, 317, 319, 321, 323, 325, 327, 329, 331, 333, 335, 337, 339, 341, 343, 345, 347, 349, 351, 353, 355, 357, 359, 361, 363, 365, 367, 369, 371, 373, 375, 377, 379, 381, 383, 385, 387, 389, 391, 393, 395, 397, 399, 401, 403, 405, 407, 409, 411, 413, 415, 417, 419, 421, 423, 425, 427, 429, 431, 433, 435, 437, 439, 441, 443, 445, 447, 449, 451, 453, 455, 457, 459, 461, 463, 465, 467, 469, 471, 473, 475, 477, 479, 481, 483, 485, 487, 489, 491, 493, 495, 497, 499, 501, 503, 505, 507, 509, 511, 513, 515, 517, 519, 521, 523, 525, 527, 529, 531, 533, 535, 537, 539, 541, 543, 545, 547, 549, 551, 553, 555, 557, 559, 561, 563, 565, 567, 569, 571, 573, 575, 577, 579, 581, 583, 585, 587, 589, 591, 593, 595, 597, 599, 601, 603, 605, 607, 609, 611, 613, 615, 617, 619, 621, 623, 625, 627, 629, 631, 633, 635, 637, 639, 641, 643, 645, 647, 649, 651, 653, 655, 657, 659, 661, 663, 665, 667, 669, 671, 673, or 675; or comprising the amino acid sequence of SEQ ID NO: 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31, 33, 35, 37, 39, 41, 43, 45, 47, 49, 51, 53, 55, 57, 59, 61, 63, 65, 67, 69, 71, 73, 75, 77, 79, 81, 83, 85, 87, 89, 91, 93, 95, 97, 99, 101, 103, 105, 107, 109, 111, 113, 115, 117, 119, 121, 123, 125, 127, 129, 131, 133, 135, 137, 139, 141, 143, 145, 147, 149, 151, 153, 155, 157, 159, 161, 163, 165, 167, 169, 171, 173, 175, 177, 179, 181, 183, 185, 187, 189, 191, 193, 195, 197, 199, 201, 203, 205, 207, 209, 211, 213, 215, 217, 219, 221, 223, 225, 227, 229, 231, 233, 235, 237, 239, 241, 243, 245, 247, 249, 251, 253, 255, 257, 259, 261, 263, 265, 267, 269, 271, 273, 275, 277, 279, 281, 283, 285, 287, 289, 291, 293, 295, 297, 299, 301, 303, 305, 307, 309, 311, 313, 315, 317, 319, 321, 323, 325, 327, 329, 331, 333, 335, 337, 339, 341, 343, 345, 347, 349, 351, 353, 355, 357, 359, 361, 363, 365, 367, 369, 371, 373, 375, 377, 379, 381, 383, 385, 387, 389, 391, 393, 395, 397, 399, 401, 403, 405, 407, 409, 411, 413, 415, 417, 419, 421, 423, 425, 427, 429, 431, 433, 435, 437, 439, 441, 443, 445, 447, 449, 451, 453, 455, 457, 459, 461, 463, 465, 467, 469, 471, 473, 475, 477, 479, 481, 483, 485, 487, 489, 491, 493, 495, 497, 499, 501, 503, 505, 507, 509, 511, 513, 515, 517, 519, 521, 523, 525, 527, 529, 531, 533, 535, 537, 539, 541, 543, 545, 547, 549, 551, 553, 555, 557, 559, 561, 563, 565, 567, 569, 571, 573, 575, 577, 579, 581, 583, 585, 587, 589, 591, 593, 595, 597, 599, 601, 603, 605, 607, 609, 611, 613, 615, 617, 619, 621, 623, 625, 627, 629, 631, 633, 635, 637, 639, 641, 643, 645, 647, 649, 651, 653, 655, 657, 659, 661, 663, 665, 667, 669, 671, 673, or 675; and/or a light chain variable region comprising an amino acid sequence with at least 75% identity to the amino acid sequence of SEQ ID NO: 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 44, 46, 48, 50, 52, 54, 56, 58, 60, 62, 64, 66, 68, 70, 72, 74, 76, 78, 80, 82, 84, 86, 88, 90, 92, 94, 96, 98, 100, 102, 104, 106, 108, 110, 112, 114, 116, 118, 120, 122, 124, 126, 128, 130, 132, 134, 136, 138, 140, 142, 144, 146, 148, 150, 152, 154, 156, 158, 160, 162, 164, 166, 168, 170, 172, 174, 176, 178, 180, 182, 184, 186, 188, 190, 192, 194, 196, 198, 200, 202, 204, 206, 208, 210, 212, 214, 216, 218, 220, 222, 224, 226, 228, 230, 232, 234, 236, 238, 240, 242, 244, 246, 248, 250, 252, 254, 256, 258, 260, 262, 264, 266, 268, 270, 272, 274, 276, 278, 280, 282, 284, 286, 288, 290, 292, 294, 296, 298, 300, 302, 304, 306, 308, 310, 312, 314, 316, 318, 320, 322, 324, 326, 328, 330, 332, 334, 336, 338, 340, 342, 344, 346, 348, 350, 352, 354, 356, 358, 360, 362, 364, 366, 368, 370, 372, 374, 376, 378, 380, 382, 384, 386, 388, 390, 392, 394, 396, 398, 400, 402, 404, 406, 408, 410, 412, 414, 416, 418, 420, 422, 424, 426, 428, 430, 432, 434, 436, 438, 440, 442, 444, 446, 448, 450, 452, 454, 456, 458, 460, 462, 464, 466, 468, 470, 472, 474, 476, 478, 480, 482, 484, 486, 488, 490, 492, 494, 496, 498, 500, 502, 504, 506, 508, 510, 512, 514, 516, 518, 520, 522, 524, 526, 528, 530, 532, 534, 536, 538, 540, 542, 544, 546, 548, 550, 552, 554, 556, 558, 560, 562, 564, 566, 568, 570, 572, 574, 576, 578, 580, 582, 584, 586, 588, 590, 592, 594, 596, 598, 600, 602, 604, 606, 608, 610, 612, 614, 616, 618, 620, 622, 624, 626, 628, 630, 632, 634, 636, 638, 640, 642, 644, 646, 648, 650, 652, 654, 656, 658, 660, 662, 664, 666, 668, 670, 672, 674, or 676; or comprising the amino acid sequence of SEQ ID NO: 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 44, 46, 48, 50, 52, 54, 56, 58, 60, 62, 64, 66, 68, 70, 72, 74, 76, 78, 80, 82, 84, 86, 88, 90, 92, 94, 96, 98, 100, 102, 104, 106, 108, 110, 112, 114, 116, 118, 120, 122, 124, 126, 128, 130, 132, 134, 136, 138, 140, 142, 144, 146, 148, 150, 152, 154, 156, 158, 160, 162, 164, 166, 168, 170, 172, 174, 176, 178, 180, 182, 184, 186, 188, 190, 192, 194, 196, 198, 200, 202, 204, 206, 208, 210, 212, 214, 216, 218, 220, 222, 224, 226, 228, 230, 232, 234, 236, 238, 240, 242, 244, 246, 248, 250, 252, 254, 256, 258, 260, 262, 264, 266, 268, 270, 272, 274, 276, 278, 280, 282, 284, 286, 288, 290, 292, 294, 296, 298, 300, 302, 304, 306, 308, 310, 312, 314, 316, 318, 320, 322, 324, 326, 328, 330, 332, 334, 336, 338, 340, 342, 344, 346, 348, 350, 352, 354, 356, 358, 360, 362, 364, 366, 368, 370, 372, 374, 376, 378, 380, 382, 384, 386, 388, 390, 392, 394, 396, 398, 400, 402, 404, 406, 408, 410, 412, 414, 416, 418, 420, 422, 424, 426, 428, 430, 432, 434, 436, 438, 440, 442, 444, 446, 448, 450, 452, 454, 456, 458, 460, 462, 464, 466, 468, 470, 472, 474, 476, 478, 480, 482, 484, 486, 488, 490, 492, 494, 496, 498, 500, 502, 504, 506, 508, 510, 512, 514, 516, 518, 520, 522, 524, 526, 528, 530, 532, 534, 536, 538, 540, 542, 544, 546, 548, 550, 552, 554, 556, 558, 560, 562, 564, 566, 568, 570, 572, 574, 576, 578, 580, 582, 584, 586, 588, 590, 592, 594, 596, 598, 600, 602, 604, 606, 608, 610, 612, 614, 616, 618, 620, 622, 624, 626, 628, 630, 632, 634, 636, 638, 640, 642, 644, 646, 648, 650, 652, 654, 656, 658, 660, 662, 664, 666, 668, 670, 672, 674, or 676. [0009] In some embodiments, the antibody or antigen-binding fragment thereof comprises a HCVR and a LCVR that comprise a HCVR/LCVR amino acid sequence pair of SEQ ID NOs: 1- 2, 3-4, 5-6, 7-8, 9-10, 11-12, 13-14, 15-16, 17-18, 19-20, 21-22, 23-24, 25-26, 27-28, 29-30, 31- 32, 33-34, 35-36, 37-38, 39-40, 41-42, 43-44, 45-46, 47-48, 49-50, 51-52, 53-54, 55-56, 57-58, 59-60, 61-62, 63-64, 65-66, 67-68, 69-70, 71-72, 73-74, 75-76, 77-78, 79-80, 81-82, 83-84, 85-86, 87-88, 89-90, 91-92, 93-94, 95-96, 97-98, 99-100, 101-102, 103-104, 105-106, 107-108, 109-110, 111-112, 113-114, 115-116, 117-118, 119-120, 121-122, 123-124, 125-126, 127-128, 129-130, 131-132, 133-134, 135-136, 137-138, 139-140, 141-142, 143-144, 145-146, 147-148, 149-150, 151-152, 153-154, 155-156, 157-158, 159-160, 161-162, 163-164, 165-166, 167-168, 169-170, 171-172, 173-174, 175-176, 177-178, 179-180, 181-182, 183-184, 185-186, 187-188, 189-190, 191-192, 193-194, 195-196, 197-198, 199-200, 201-202, 203-204, 205-206, 207-208, 209-210, 211-212, 213-214, 215-216, 217-218, 219-220, 221-222, 223-224, 225-226, 227-228, 229-230, 231-232, 233-234, 235-236, 237-238, 239-240, 241-242, 243-244, 245-246, 247-248, 249-250, 251-252, 253-254, 255-256, 257-258, 259-260, 261-262, 263-264, 265-266, 267-268, 269-270, 271-272, 273-274, 275-276, 277-278, 279-280, 281-282, 283-284, 285-286, 287-288, 289-290, 291-292, 293-294, 295-296, 297-298, 299-300, 301-302, 303-304, 305-306, 307-308, 309-310, 311-312, 313-314, 315-316, 317-318, 319-320, 321-322, 323-324, 325-326, 327-328, 329-330, 331-332, 333-334, 335-336, 337-338, 339-340, 341-342, 343-344, 345-346, 347-348, 349-350, 351-352, 353-354, 355-356, 357-358, 359-360, 361-362, 363-364, 365-366, 367-368, 369-370, 371-372, 373-374, 375-376, 377-378, 379-380, 381-382, 383-384, 385-386, 387-388, 389-390, 391-392, 393-394, 395-396, 397-398, 399-400, 401-402, 403-404, 405-406, 407-408, 409-410, 411-412, 413-414, 415-416, 417-418, 419-420, 421-422, 423-424, 425-426, 427-428, 429-430, 431-432, 433-434, 435-436, 437-438, 439-440, 441-442, 443-444, 445-446, 447-448, 449-450, 451-452, 453-454, 455-456, 457-458, 459-460, 461-462, 463-464, 465-466, 467-468, 469-470, 471-472, 473-474, 475-476, 477-478, 479-480, 481-482, 483-484, 485-486, 487-488, 489-490, 491-492, 493-494, 495-496, 497-498, 499-500, 501-502, 503-504, 505-506, 507-508, 509-510, 511-512, 513-514, 515-516, 517-518, 519-520, 521-522, 523-524, 525-526, 527-528, 529-530, 531-532, 533-534, 535-536, 537-538, 539-540, 541-542, 543-544, 545-546, 547-548, 549-550, 551-552, 553-554, 555-556, 557-558, 559-560, 561-562, 563-564, 565-566, 567-568, 569-570, 571-572, 573-574, 575-576, 577-578, 579-580, 581-582, 583-584, 585-586, 587-588, 589-590, 591-592, 593-594, 595-596, 597-598, 599-600, 601-602, 603-604, 605-606, 607-608, 609-610, 611-612, 613-614, 615-616, 617-618, 619-620, 621-622, 623-624, 625-626, 627-628, 629-630, 631-632, 633-634, 635-636, 637-638, 639-640, 641-642, 643-644, 645-646, 647-648, 649-650, 651-652, 653-654, 655-656, 657-658, 659-660, 661-662, 663-664, 665-666, 667-668, 669-670, 671-672, 673-674, or 675-676. [0010] In some embodiments, the antibody or antigen-binding fragment thereof comprises a HCVR and a LCVR that comprise a HCVR/LCVR amino acid sequence pair with at least 75% identity to the amino acid sequence of SEQ ID NOs: 1-2, 3-4, 5-6, 7-8, 9-10, 11-12, 13-14, 15-16, 17-18, 19-20, 21-22, 23-24, 25-26, 27-28, 29-30, 31-32, 33-34, 35-36, 37-38, 39-40, 41-42, 43-44, 45-46, 47-48, 49-50, 51-52, 53-54, 55-56, 57-58, 59-60, 61-62, 63-64, 65-66, 67-68, 69-70, 71-72, 73-74, 75-76, 77-78, 79-80, 81-82, 83-84, 85-86, 87-88, 89-90, 91-92, 93-94, 95-96, 97-98, 99- 100, 101-102, 103-104, 105-106, 107-108, 109-110, 111-112, 113-114, 115-116, 117-118, 119- 120, 121-122, 123-124, 125-126, 127-128, 129-130, 131-132, 133-134, 135-136, 137-138, 139- 140, 141-142, 143-144, 145-146, 147-148, 149-150, 151-152, 153-154, 155-156, 157-158, 159- 160, 161-162, 163-164, 165-166, 167-168, 169-170, 171-172, 173-174, 175-176, 177-178, 179- 180, 181-182, 183-184, 185-186, 187-188, 189-190, 191-192, 193-194, 195-196, 197-198, 199- 200, 201-202, 203-204, 205-206, 207-208, 209-210, 211-212, 213-214, 215-216, 217-218, 219- 220, 221-222, 223-224, 225-226, 227-228, 229-230, 231-232, 233-234, 235-236, 237-238, 239- 240, 241-242, 243-244, 245-246, 247-248, 249-250, 251-252, 253-254, 255-256, 257-258, 259- 260, 261-262, 263-264, 265-266, 267-268, 269-270, 271-272, 273-274, 275-276, 277-278, 279- 280, 281-282, 283-284, 285-286, 287-288, 289-290, 291-292, 293-294, 295-296, 297-298, 299- 300, 301-302, 303-304, 305-306, 307-308, 309-310, 311-312, 313-314, 315-316, 317-318, 319- 320, 321-322, 323-324, 325-326, 327-328, 329-330, 331-332, 333-334, 335-336, 337-338, 339- 340, 341-342, 343-344, 345-346, 347-348, 349-350, 351-352, 353-354, 355-356, 357-358, 359- 360, 361-362, 363-364, 365-366, 367-368, 369-370, 371-372, 373-374, 375-376, 377-378, 379- 380, 381-382, 383-384, 385-386, 387-388, 389-390, 391-392, 393-394, 395-396, 397-398, 399- 400, 401-402, 403-404, 405-406, 407-408, 409-410, 411-412, 413-414, 415-416, 417-418, 419- 420, 421-422, 423-424, 425-426, 427-428, 429-430, 431-432, 433-434, 435-436, 437-438, 439- 440, 441-442, 443-444, 445-446, 447-448, 449-450, 451-452, 453-454, 455-456, 457-458, 459- 460, 461-462, 463-464, 465-466, 467-468, 469-470, 471-472, 473-474, 475-476, 477-478, 479- 480, 481-482, 483-484, 485-486, 487-488, 489-490, 491-492, 493-494, 495-496, 497-498, 499- 500, 501-502, 503-504, 505-506, 507-508, 509-510, 511-512, 513-514, 515-516, 517-518, 519- 520, 521-522, 523-524, 525-526, 527-528, 529-530, 531-532, 533-534, 535-536, 537-538, 539- 540, 541-542, 543-544, 545-546, 547-548, 549-550, 551-552, 553-554, 555-556, 557-558, 559- 560, 561-562, 563-564, 565-566, 567-568, 569-570, 571-572, 573-574, 575-576, 577-578, 579- 580, 581-582, 583-584, 585-586, 587-588, 589-590, 591-592, 593-594, 595-596, 597-598, 599- 600, 601-602, 603-604, 605-606, 607-608, 609-610, 611-612, 613-614, 615-616, 617-618, 619- 620, 621-622, 623-624, 625-626, 627-628, 629-630, 631-632, 633-634, 635-636, 637-638, 639- 640, 641-642, 643-644, 645-646, 647-648, 649-650, 651-652, 653-654, 655-656, 657-658, 659- 660, 661-662, 663-664, 665-666, 667-668, 669-670, 671-672, 673-674, or 675-676. [0011] In some embodiments, the present disclosure provides an antibody or antigen-binding fragment thereof that binds to the S polypeptide of a Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), wherein the antibody or antigen-binding fragment thereof comprises: (a) a variable heavy chain CDR1 (HCDR1) comprising an amino acid sequence selected from SEQ ID NO: 1354, 1360, 1366, 1372, and 1378; (b) a variable heavy chain CDR2 (HCDR2) comprising an amino acid sequence of SEQ ID NO: 1355, 1361, 1367, 1373, and 1379; (c) a variable heavy chain CDR3 (HCDR3) comprising an amino acid sequence selected from SEQ ID NO: 1356, 1362, 1368, 1374, and 1380; (d) a variable light chain CDR1 (LCDR1) comprising an amino acid sequence selected from SEQ ID NO: 1357, 1363, 1369, 1375, and 1381; (e) a variable light chain CDR2 (LCDR2) comprising an amino acid sequence selected from SEQ ID NO: 1358, 1364, 1370, 1376, and 1382; and (f) a variable light chain CDR3 (LCDR3) comprising an amino acid sequence selected from SEQ ID NO: 1359, 1365, 1371, 1377, and 1383. [0012] In some embodiments, the antibody or antigen-binding fragment thereof comprises: a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1354, a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1355, a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1356, a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1357, a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1358, and a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1359. [0013] In some embodiments, the antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising an amino acid sequence that is at least about 75%, at least 85%, at least 90%, at least 95%, at least 99%, or 100% identical to SEQ ID NO: 25, and a light chain variable domain (VL) comprising an amino acid sequence that is at least about 75%, at least 85%, at least 90%, at least 95%, at least 99%, or 100% identical to SEQ ID NO: 26. [0014] In some embodiments, the antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising or consisting of the amino acid sequence of SEQ ID NO: 25, and a light chain variable domain (VL) comprising or consisting of the amino acid sequence of SEQ ID NO: 26. [0015] In some embodiments, the antibody or antigen-binding fragment thereof comprises a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1360, a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1361, a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1362, a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1363, a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1364, and a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1365. [0016] In some embodiments, the antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising an amino acid sequence that is at least about 75%, at least 85%, at least 90%, at least 95%, at least 99%, or 100% identical to SEQ ID NO: 395, and a light chain variable domain (VL) comprising an amino acid sequence that is at least about 75%, at least 85%, at least 90%, at least 95%, at least 99%, or 100% identical to SEQ ID NO: 396. In some embodiments, the antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising or consisting of the amino acid sequence of SEQ ID NO: 395, and a light chain variable domain (VL) comprising or consisting of the amino acid sequence of SEQ ID NO: 396. [0017] In some embodiments, the antibody or antigen-binding fragment thereof comprises a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1366, a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1367, a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1368, a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1369, a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1370, and a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1371. [0018] In some embodiments, the antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising an amino acid sequence that is at least about 75%, at least 85%, at least 90%, at least 95%, at least 99%, or 100% identical to SEQ ID NO: 397, and a light chain variable domain (VL) comprising an amino acid sequence that is at least about 75%, at least 85%, at least 90%, at least 95%, at least 99%, or 100% identical to SEQ ID NO: 398. In some embodiments, the antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising or consisting of the amino acid sequence of SEQ ID NO: 397, and a light chain variable domain (VL) comprising or consisting of the amino acid sequence of SEQ ID NO: 398. [0019] In some embodiments, the antibody or antigen-binding fragment thereof comprises a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1372, a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1373, a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1374, a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1375, a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1376, and a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1377. [0020] In some embodiments, the antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising an amino acid sequence that is at least about 75%, at least 85%, at least 90%, at least 95%, at least 99%, or 100% identical to SEQ ID NO: 191, and a light chain variable domain (VL) comprising an amino acid sequence that is at least about 75%, at least 85%, at least 90%, at least 95%, at least 99%, or 100% identical to SEQ ID NO: 192. In some embodiments, the antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising or consisting of the amino acid sequence of SEQ ID NO: 191, and a light chain variable domain (VL) comprising or consisting of the amino acid sequence of SEQ ID NO: 192. [0021] In some embodiments, the antibody or antigen-binding fragment thereof comprises a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1378, a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1379, a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1380, a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1381, a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1382, and a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1383. In some embodiments, the antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising an amino acid sequence that is at least about 75%, at least 85%, at least 90%, at least 95%, at least 99%, or 100% identical to SEQ ID NO: 219, and a light chain variable domain (VL) comprising an amino acid sequence that is at least about 75%, at least 85%, at least 90%, at least 95%, at least 99%, or 100% identical to SEQ ID NO: 220. [0022] In some embodiments, the antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising or consisting of the amino acid sequence of SEQ ID NO: 219, and a light chain variable domain (VL) comprising or consisting of the amino acid sequence of SEQ ID NO: 220. [0023] In some embodiments, the antibody or antigen-binding fragment thereof is a fully human antibody or antigen-binding fragment thereof. [0024] In some embodiments, the antibody or antigen-binding fragment thereof binds to an epitope within the receptor binding domain (RBD) or the N-terminal domain (NTD) of the S polypeptide. In some embodiments, the antibody or antigen-binding fragment thereof binds to an epitope selected from the following: (a) the supersite b-hairpin (residues 152-158 of SEQ ID NO: 1353); (b) WKH^ȕ^-strand (residue 97-102 of SEQ ID NO: 1353); (c) N4-loop (residues 178-188 of SEQ ID NO: 1353); (d) N-linked glycans at positions N122 and N149 of the spike polypeptide of SEQ ID NO: 1353; (e) residues 27-32, 57-60, 210-218, and/or 286-303 of the spike polypeptide of SEQ ID NO: 1353; and/or (f) residues 600-606 of the spike polypeptide of SEQ ID NO: 1353. [0025] In some embodiments, the present disclosure provides an antibody or antigen-binding fragment thereof, wherein the antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising an amino acid sequence that is at least about 75%, at least 85%, at least 90%, at least 95%, at least 99%, or 100% identical to a SEQ ID NO selected from column 7 of Table 2, and a light chain variable domain (VL) comprising an amino acid sequence that is at least about 75%, at least 85%, at least 90%, at least 95%, at least 99%, or 100% identical to a SEQ ID NO selected from column 11 of Table 2, wherein the VH SEQ ID NO and the VL SEQ ID NO are selected from the same row. In some embodiments, the antibody or antigen- binding fragment thereof comprises a heavy chain variable domain (VH) comprising or consisting of a SEQ ID NO selected from column 7 of Table 2, and a light chain variable domain (VL) comprising or consisting of a SEQ ID NO selected from column 11 of Table 2, wherein the VH SEQ ID No and the VL SEQ ID NO are selected from the same row. [0026] In some embodiments, the antibody or antigen-binding fragment thereof neutralizes SARS-CoV-2. In some embodiments, the antibody or antigen-binding fragment thereof inhibits the fusion of SARS-CoV-2 and a host cell membrane. In some embodiments, the antibody or antigen-binding fragment thereof is cytotoxic to a SARS-CoV-2 infected host cell. [0027] In some embodiments, the antibody or antigen-binding fragment thereof is a multivalent antibody comprising (a) a first target binding site that specifically binds to an epitope within the spike polypeptide, and (b) a second target binding site that binds to an epitope on a different epitope on the spike polypeptide or a different molecule. [0028] In some embodiments, the antibody or antigen-binding fragment thereof further comprises a variant Fc constant region. [0029] In some embodiments, the antibody or antigen-binding fragment thereof is a monoclonal antibody, a chimeric antibody, a humanized antibody, or humanized antibody. In some embodiments, the antibody or antigen-binding fragment thereof is a single-chain antibody, Fab, or Fab2 fragment. [0030] In some embodiments, the antibody or antigen-binding fragment thereof is detectably labeled or conjugated to a toxin, a therapeutic agent, a polymer, a receptor, an enzyme, or a receptor ligand, preferably wherein the polymer is polyethylene glycol (PEG). [0031] In some embodiments, the present disclosure provides a polynucleotide encoding the antibody or antigen-binding fragment thereof described herein. In some embodiments, the present disclosure provides a vector comprising a polynucleotide described herein. In some embodiments, the present disclosure provides a cultured host cell comprising a vector described herein. [0032] In some embodiments, the present disclosure provides a pharmaceutical composition comprising an antibody or antigen-binding fragment thereof, polynucleotide, or vector described herein. [0033] In some embodiments, the present disclosure provides a pharmaceutical composition comprising two or more antibodies or antigen-binding fragments thereof, wherein the two or more antibodies or antigen-binding fragments thereof are selected from: a HCVR/LCVR amino acid sequence pair of SEQ ID NOs: 1-2, 3-4, 5-6, 7-8, 9-10, 11-12, 13-14, 15-16, 17-18, 19-20, 21-22, 23-24, 25-26, 27-28, 29-30, 31-32, 33-34, 35-36, 37-38, 39-40, 41-42, 43-44, 45-46, 47-48, 49-50, 51-52, 53-54, 55-56, 57-58, 59-60, 61-62, 63-64, 65-66, 67-68, 69-70, 71-72, 73-74, 75-76, 77-78, 79-80, 81-82, 83-84, 85-86, 87-88, 89-90, 91-92, 93-94, 95-96, 97-98, 99-100, 101-102, 103-104, 105-106, 107-108, 109-110, 111-112, 113-114, 115-116, 117-118, 119-120, 121-122, 123-124, 125-126, 127-128, 129-130, 131-132, 133-134, 135-136, 137-138, 139-140, 141-142, 143-144, 145-146, 147-148, 149-150, 151-152, 153-154, 155-156, 157-158, 159-160, 161-162, 163-164, 165-166, 167-168, 169-170, 171-172, 173-174, 175-176, 177-178, 179-180, 181-182, 183-184, 185-186, 187-188, 189-190, 191-192, 193-194, 195-196, 197-198, 199-200, 201-202, 203-204, 205-206, 207-208, 209-210, 211-212, 213-214, 215-216, 217-218, 219-220, 221-222, 223-224, 225-226, 227-228, 229-230, 231-232, 233-234, 235-236, 237-238, 239-240, 241-242, 243-244, 245-246, 247-248, 249-250, 251-252, 253-254, 255-256, 257-258, 259-260, 261-262, 263-264, 265-266, 267-268, 269-270, 271-272, 273-274, 275-276, 277-278, 279-280, 281-282, 283-284, 285-286, 287-288, 289-290, 291-292, 293-294, 295-296, 297-298, 299-300, 301-302, 303-304, 305-306, 307-308, 309-310, 311-312, 313-314, 315-316, 317-318, 319-320, 321-322, 323-324, 325-326, 327-328, 329-330, 331-332, 333-334, 335-336, 337-338, 339-340, 341-342, 343-344, 345-346, 347-348, 349-350, 351-352, 353-354, 355-356, 357-358, 359-360, 361-362, 363-364, 365-366, 367-368, 369-370, 371-372, 373-374, 375-376, 377-378, 379-380, 381-382, 383-384, 385-386, 387-388, 389-390, 391-392, 393-394, 395-396, 397-398, 399-400, 401-402, 403-404, 405-406, 407-408, 409-410, 411-412, 413-414, 415-416, 417-418, 419-420, 421-422, 423-424, 425-426, 427-428, 429-430, 431-432, 433-434, 435-436, 437-438, 439-440, 441-442, 443-444, 445-446, 447-448, 449-450, 451-452, 453-454, 455-456, 457-458, 459-460, 461-462, 463-464, 465-466, 467-468, 469-470, 471-472, 473-474, 475-476, 477-478, 479-480, 481-482, 483-484, 485-486, 487-488, 489-490, 491-492, 493-494, 495-496, 497-498, 499-500, 501-502, 503-504, 505-506, 507-508, 509-510, 511-512, 513-514, 515-516, 517-518, 519-520, 521-522, 523-524, 525-526, 527-528, 529-530, 531-532, 533-534, 535-536, 537-538, 539-540, 541-542, 543-544, 545-546, 547-548, 549-550, 551-552, 553-554, 555-556, 557-558, 559-560, 561-562, 563-564, 565-566, 567-568, 569-570, 571-572, 573-574, 575-576, 577-578, 579-580, 581-582, 583-584, 585-586, 587-588, 589-590, 591-592, 593-594, 595-596, 597-598, 599-600, 601-602, 603-604, 605-606, 607-608, 609-610, 611-612, 613-614, 615-616, 617-618, 619-620, 621-622, 623-624, 625-626, 627-628, 629-630, 631-632, 633-634, 635-636, 637-638, 639-640, 641-642, 643-644, 645-646, 647-648, 649-650, 651-652, 653-654, 655-656, 657-658, 659-660, 661-662, 663-664, 665-666, 667-668, 669-670, 671-672, 673-674, or 675-676; and a second antibody or antigen- binding fragment thereof comprising a HCVR/LCVR amino acid sequence pair of SEQ ID NOs: 1-2, 3-4, 5-6, 7-8, 9-10, 11-12, 13-14, 15-16, 17-18, 19-20, 21-22, 23-24, 25-26, 27-28, 29-30, 31- 32, 33-34, 35-36, 37-38, 39-40, 41-42, 43-44, 45-46, 47-48, 49-50, 51-52, 53-54, 55-56, 57-58, 59-60, 61-62, 63-64, 65-66, 67-68, 69-70, 71-72, 73-74, 75-76, 77-78, 79-80, 81-82, 83-84, 85-86, 87-88, 89-90, 91-92, 93-94, 95-96, 97-98, 99-100, 101-102, 103-104, 105-106, 107-108, 109-110, 111-112, 113-114, 115-116, 117-118, 119-120, 121-122, 123-124, 125-126, 127-128, 129-130, 131-132, 133-134, 135-136, 137-138, 139-140, 141-142, 143-144, 145-146, 147-148, 149-150, 151-152, 153-154, 155-156, 157-158, 159-160, 161-162, 163-164, 165-166, 167-168, 169-170, 171-172, 173-174, 175-176, 177-178, 179-180, 181-182, 183-184, 185-186, 187-188, 189-190, 191-192, 193-194, 195-196, 197-198, 199-200, 201-202, 203-204, 205-206, 207-208, 209-210, 211-212, 213-214, 215-216, 217-218, 219-220, 221-222, 223-224, 225-226, 227-228, 229-230, 231-232, 233-234, 235-236, 237-238, 239-240, 241-242, 243-244, 245-246, 247-248, 249-250, 251-252, 253-254, 255-256, 257-258, 259-260, 261-262, 263-264, 265-266, 267-268, 269-270, 271-272, 273-274, 275-276, 277-278, 279-280, 281-282, 283-284, 285-286, 287-288, 289-290, 291-292, 293-294, 295-296, 297-298, 299-300, 301-302, 303-304, 305-306, 307-308, 309-310, 311-312, 313-314, 315-316, 317-318, 319-320, 321-322, 323-324, 325-326, 327-328, 329-330, 331-332, 333-334, 335-336, 337-338, 339-340, 341-342, 343-344, 345-346, 347-348, 349-350, 351-352, 353-354, 355-356, 357-358, 359-360, 361-362, 363-364, 365-366, 367-368, 369-370, 371-372, 373-374, 375-376, 377-378, 379-380, 381-382, 383-384, 385-386, 387-388, 389-390, 391-392, 393-394, 395-396, 397-398, 399-400, 401-402, 403-404, 405-406, 407-408, 409-410, 411-412, 413-414, 415-416, 417-418, 419-420, 421-422, 423-424, 425-426, 427-428, 429-430, 431-432, 433-434, 435-436, 437-438, 439-440, 441-442, 443-444, 445-446, 447-448, 449-450, 451-452, 453-454, 455-456, 457-458, 459-460, 461-462, 463-464, 465-466, 467-468, 469-470, 471-472, 473-474, 475-476, 477-478, 479-480, 481-482, 483-484, 485-486, 487-488, 489-490, 491-492, 493-494, 495-496, 497-498, 499-500, 501-502, 503-504, 505-506, 507-508, 509-510, 511-512, 513-514, 515-516, 517-518, 519-520, 521-522, 523-524, 525-526, 527-528, 529-530, 531-532, 533-534, 535-536, 537-538, 539-540, 541-542, 543-544, 545-546, 547-548, 549-550, 551-552, 553-554, 555-556, 557-558, 559-560, 561-562, 563-564, 565-566, 567-568, 569-570, 571-572, 573-574, 575-576, 577-578, 579-580, 581-582, 583-584, 585-586, 587-588, 589-590, 591-592, 593-594, 595-596, 597-598, 599-600, 601-602, 603-604, 605-606, 607-608, 609-610, 611-612, 613-614, 615-616, 617-618, 619-620, 621-622, 623-624, 625-626, 627-628, 629-630, 631-632, 633-634, 635-636, 637-638, 639-640, 641-642, 643-644, 645-646, 647-648, 649-650, 651-652, 653-654, 655-656, 657-658, 659-660, 661-662, 663-664, 665-666, 667-668, 669-670, 671-672, 673-674, or 675-676, wherein the first antibody or antigen-binding fragment thereof is different from the second antibody or antigen-binding fragment thereof. [0034] In some embodiments, the two or more of the antibody or antigen-binding fragment thereof comprise: a first antibody or antigen-binding fragment thereof comprising a HCVR/LCVR amino acid sequence pair with at least 75% identity to the amino acid sequence of SEQ ID NOs: 1-2, 3-4, 5-6, 7-8, 9-10, 11-12, 13-14, 15-16, 17-18, 19-20, 21-22, 23-24, 25-26, 27-28, 29-30, 31- 32, 33-34, 35-36, 37-38, 39-40, 41-42, 43-44, 45-46, 47-48, 49-50, 51-52, 53-54, 55-56, 57-58, 59-60, 61-62, 63-64, 65-66, 67-68, 69-70, 71-72, 73-74, 75-76, 77-78, 79-80, 81-82, 83-84, 85-86, 87-88, 89-90, 91-92, 93-94, 95-96, 97-98, 99-100, 101-102, 103-104, 105-106, 107-108, 109-110, 111-112, 113-114, 115-116, 117-118, 119-120, 121-122, 123-124, 125-126, 127-128, 129-130, 131-132, 133-134, 135-136, 137-138, 139-140, 141-142, 143-144, 145-146, 147-148, 149-150, 151-152, 153-154, 155-156, 157-158, 159-160, 161-162, 163-164, 165-166, 167-168, 169-170, 171-172, 173-174, 175-176, 177-178, 179-180, 181-182, 183-184, 185-186, 187-188, 189-190, 191-192, 193-194, 195-196, 197-198, 199-200, 201-202, 203-204, 205-206, 207-208, 209-210, 211-212, 213-214, 215-216, 217-218, 219-220, 221-222, 223-224, 225-226, 227-228, 229-230, 231-232, 233-234, 235-236, 237-238, 239-240, 241-242, 243-244, 245-246, 247-248, 249-250, 251-252, 253-254, 255-256, 257-258, 259-260, 261-262, 263-264, 265-266, 267-268, 269-270, 271-272, 273-274, 275-276, 277-278, 279-280, 281-282, 283-284, 285-286, 287-288, 289-290, 291-292, 293-294, 295-296, 297-298, 299-300, 301-302, 303-304, 305-306, 307-308, 309-310, 311-312, 313-314, 315-316, 317-318, 319-320, 321-322, 323-324, 325-326, 327-328, 329-330, 331-332, 333-334, 335-336, 337-338, 339-340, 341-342, 343-344, 345-346, 347-348, 349-350, 351-352, 353-354, 355-356, 357-358, 359-360, 361-362, 363-364, 365-366, 367-368, 369-370, 371-372, 373-374, 375-376, 377-378, 379-380, 381-382, 383-384, 385-386, 387-388, 389-390, 391-392, 393-394, 395-396, 397-398, 399-400, 401-402, 403-404, 405-406, 407-408, 409-410, 411-412, 413-414, 415-416, 417-418, 419-420, 421-422, 423-424, 425-426, 427-428, 429-430, 431-432, 433-434, 435-436, 437-438, 439-440, 441-442, 443-444, 445-446, 447-448, 449-450, 451-452, 453-454, 455-456, 457-458, 459-460, 461-462, 463-464, 465-466, 467-468, 469-470, 471-472, 473-474, 475-476, 477-478, 479-480, 481-482, 483-484, 485-486, 487-488, 489-490, 491-492, 493-494, 495-496, 497-498, 499-500, 501-502, 503-504, 505-506, 507-508, 509-510, 511-512, 513-514, 515-516, 517-518, 519-520, 521-522, 523-524, 525-526, 527-528, 529-530, 531-532, 533-534, 535-536, 537-538, 539-540, 541-542, 543-544, 545-546, 547-548, 549-550, 551-552, 553-554, 555-556, 557-558, 559-560, 561-562, 563-564, 565-566, 567-568, 569-570, 571-572, 573-574, 575-576, 577-578, 579-580, 581-582, 583-584, 585-586, 587-588, 589-590, 591-592, 593-594, 595-596, 597-598, 599-600, 601-602, 603-604, 605-606, 607-608, 609-610, 611-612, 613-614, 615-616, 617-618, 619-620, 621-622, 623-624, 625-626, 627-628, 629-630, 631-632, 633-634, 635-636, 637-638, 639-640, 641-642, 643-644, 645-646, 647-648, 649-650, 651-652, 653-654, 655-656, 657-658, 659-660, 661-662, 663-664, 665-666, 667-668, 669-670, 671-672, 673-674, or 675-676; and a second antibody or antigen-binding fragment thereof comprising a HCVR/LCVR amino acid sequence pair with at least 75% identity to the amino acid sequence of SEQ ID NOs: 1-2, 3-4, 5-6, 7-8, 9-10, 11-12, 13-14, 15-16, 17-18, 19-20, 21-22, 23- 24, 25-26, 27-28, 29-30, 31-32, 33-34, 35-36, 37-38, 39-40, 41-42, 43-44, 45-46, 47-48, 49-50, 51-52, 53-54, 55-56, 57-58, 59-60, 61-62, 63-64, 65-66, 67-68, 69-70, 71-72, 73-74, 75-76, 77-78, 79-80, 81-82, 83-84, 85-86, 87-88, 89-90, 91-92, 93-94, 95-96, 97-98, 99-100, 101-102, 103-104, 105-106, 107-108, 109-110, 111-112, 113-114, 115-116, 117-118, 119-120, 121-122, 123-124, 125-126, 127-128, 129-130, 131-132, 133-134, 135-136, 137-138, 139-140, 141-142, 143-144, 145-146, 147-148, 149-150, 151-152, 153-154, 155-156, 157-158, 159-160, 161-162, 163-164, 165-166, 167-168, 169-170, 171-172, 173-174, 175-176, 177-178, 179-180, 181-182, 183-184, 185-186, 187-188, 189-190, 191-192, 193-194, 195-196, 197-198, 199-200, 201-202, 203-204, 205-206, 207-208, 209-210, 211-212, 213-214, 215-216, 217-218, 219-220, 221-222, 223-224, 225-226, 227-228, 229-230, 231-232, 233-234, 235-236, 237-238, 239-240, 241-242, 243-244, 245-246, 247-248, 249-250, 251-252, 253-254, 255-256, 257-258, 259-260, 261-262, 263-264, 265-266, 267-268, 269-270, 271-272, 273-274, 275-276, 277-278, 279-280, 281-282, 283-284, 285-286, 287-288, 289-290, 291-292, 293-294, 295-296, 297-298, 299-300, 301-302, 303-304, 305-306, 307-308, 309-310, 311-312, 313-314, 315-316, 317-318, 319-320, 321-322, 323-324, 325-326, 327-328, 329-330, 331-332, 333-334, 335-336, 337-338, 339-340, 341-342, 343-344, 345-346, 347-348, 349-350, 351-352, 353-354, 355-356, 357-358, 359-360, 361-362, 363-364, 365-366, 367-368, 369-370, 371-372, 373-374, 375-376, 377-378, 379-380, 381-382, 383-384, 385-386, 387-388, 389-390, 391-392, 393-394, 395-396, 397-398, 399-400, 401-402, 403-404, 405-406, 407-408, 409-410, 411-412, 413-414, 415-416, 417-418, 419-420, 421-422, 423-424, 425-426, 427-428, 429-430, 431-432, 433-434, 435-436, 437-438, 439-440, 441-442, 443-444, 445-446, 447-448, 449-450, 451-452, 453-454, 455-456, 457-458, 459-460, 461-462, 463-464, 465-466, 467-468, 469-470, 471-472, 473-474, 475-476, 477-478, 479-480, 481-482, 483-484, 485-486, 487-488, 489-490, 491-492, 493-494, 495-496, 497-498, 499-500, 501-502, 503-504, 505-506, 507-508, 509-510, 511-512, 513-514, 515-516, 517-518, 519-520, 521-522, 523-524, 525-526, 527-528, 529-530, 531-532, 533-534, 535-536, 537-538, 539-540, 541-542, 543-544, 545-546, 547-548, 549-550, 551-552, 553-554, 555-556, 557-558, 559-560, 561-562, 563-564, 565-566, 567-568, 569-570, 571-572, 573-574, 575-576, 577-578, 579-580, 581-582, 583-584, 585-586, 587-588, 589-590, 591-592, 593-594, 595-596, 597-598, 599-600, 601-602, 603-604, 605-606, 607-608, 609-610, 611-612, 613-614, 615-616, 617-618, 619-620, 621-622, 623-624, 625-626, 627-628, 629-630, 631-632, 633-634, 635-636, 637-638, 639-640, 641-642, 643-644, 645-646, 647-648, 649-650, 651-652, 653-654, 655-656, 657-658, 659-660, 661-662, 663-664, 665-666, 667-668, 669-670, 671-672, 673-674, or 675-676, wherein the first antibody or antigen- binding fragment thereof is different from the second antibody or antigen-binding fragment thereof. [0035] In some embodiments, the present disclosure provides a pharmaceutical composition comprising two or more antibodies or antigen-binding fragments thereof, wherein the two or more antibodies or antigen-binding fragments thereof are selected from: (a) an antibody or antigen- binding fragment thereof comprising: a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1354, a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1355, a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1356, a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1357, a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1358, and a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1359; (b) an antibody or antigen-binding fragment thereof comprising: a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1360, a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1361, a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1362, a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1363, a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1364, and a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1365; (c) an antibody or antigen-binding fragment thereof comprising: a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1366, a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1367, a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1368, a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1369, a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1370, and a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1371; (d) an antibody or antigen-binding fragment thereof comprising: a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1372, a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1373, a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1374, a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1375, a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1376, and a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1377; and (e) an antibody or antigen-binding fragment thereof comprising: a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1378, a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1379, a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1380, a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1381, a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1382, and a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1383. [0036] In some embodiments, the two or more monoclonal antibodies or antigen-binding fragments thereof are selected from: (a) an antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising or consisting of the amino acid sequence of SEQ ID NO: 25, and a light chain variable domain (VL) comprising or consisting of the amino acid sequence of SEQ ID NO: 26; (b) an antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising or consisting of the amino acid sequence of SEQ ID NO: 395, and a light chain variable domain (VL) comprising or consisting of the amino acid sequence of SEQ ID NO: 396; (c) an antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising or consisting of the amino acid sequence of SEQ ID NO: 397, and a light chain variable domain (VL) comprising or consisting of the amino acid sequence of SEQ ID NO: 398; (d) an antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising or consisting of the amino acid sequence of SEQ ID NO: 191, and a light chain variable domain (VL) comprising or consisting of the amino acid sequence of SEQ ID NO: 192; and (e) an antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising or consisting of the amino acid sequence of SEQ ID NO: 219, and a light chain variable domain (VL) comprising or consisting of the amino acid sequence of SEQ ID NO: 220. [0037] In some embodiments, the pharmaceutical composition further comprises a pharmaceutically acceptable carrier. [0038] In some embodiments, the present disclosure provides a pharmaceutical composition for use in treating a SARS-CoV-2 infection in a subject. [0039] In some embodiments, the present disclosure provides a method of treating a SARS-CoV- 2 infection in a subject, comprising administering to the subject therapeutically effective amount of an antibody or antigen-binding fragment thereof or a pharmaceutical composition thereof. In some embodiments, the present disclosure provides a method of neutralizing SARS-CoV-2 in a subject, comprising administering to a subject in need thereof a therapeutically effective amount of an antibody or antigen-binding fragment thereof or a pharmaceutical composition thereof. In some embodiments, the present disclosure provides a method of preventing a SARS-CoV-2 infection in a subject, comprising administering to a subject in need thereof a therapeutically effective amount of an antibody or antigen-binding fragment thereof or pharmaceutical composition thereof. [0040] In some embodiments, the subject is immunocompromised. [0041] In some embodiments, the method further comprises administering a second therapeutic agent, preferably wherein the second therapeutic agent comprises an anti-inflammatory drug or an antiviral compound. In some embodiments, (a) the antiviral compound comprises: a nucleoside analog, a peptoid, an oligopeptide, a polypeptide, a protease inhibitor, a 3C-like protease inhibitor, a papain-like protease inhibitor, or an inhibitor of an RNA dependent RNA polymerase; or (b) the antiviral compound is selected from: acyclovir, ganciclovir, vidarabine, foscarnet, cidofovir, amantadine, ribavirin, trifluorothymidine, zidovudine, didanosine, zalcitabine, and an interferon, and preferably wherein the interferon is an interferon-Į^RU^DQ^LQWHUIHURQ-ȕ^^In some embodiments, the second therapeutic agent is administered before, after, or concurrently with the antibody or pharmaceutical composition thereof. [0042] In some embodiments, the pharmaceutical composition is administered to the subject after the exposure to SARS-CoV-2. [0043] In some embodiments, the present disclosure provides a kit for detecting a SARS-CoV- 2 infection in a subject, comprising an antibody or antigen-binding fragment thereof described herein. [0044] In some embodiments, the present disclosure provides a method of detecting the presence of a SARS-CoV-2 in a sample, comprising (1) contacting the sample with an antibody or antigen- binding fragment thereof described herein, and (2) detecting the presence of an antibody-antigen complex, wherein the presence of the antibody-antigen complex indicates the presence of SARS- CoV-2. In some embodiments, the antibody or antigen-binding fragment thereof is conjugated to a label, preferably wherein the label is selected from a fluorescent label, a chemiluminescent label, a radiolabel, and an enzyme. In some embodiments, the method further comprises contacting a secondary antibody with the antibody or antigen-binding fragment thereof, wherein the secondary antibody comprises a label. In some embodiments, the method further comprises detecting fluorescence or chemiluminescence of the label, preferably wherein the step of detecting comprises a competitive binding assay or ELISA. In some embodiments, the method further comprises binding the sample to a solid support, preferably wherein the solid support is selected from microparticles, microbeads, magnetic beads, and an affinity purification column. In some embodiments, the sample is a blood sample, a nasal swab, or a throat swab. [0045] In some embodiments, the present disclosure provides a method of preparing an antibody, or antigen-binding fragment thereof, comprising: (a) obtaining a cultured host cell described herein; (b) culturing the cultured host cell in a medium under conditions permitting expression of a polypeptide encoded by the vector and assembling of an antibody or fragment thereof; and (c) purifying the antibody or fragment from the cultured cell or the medium of the cell. [0046] In some embodiments, the present disclosure provides a kit comprising a pharmaceutically acceptable dose unit of an antibody or antigen-binding fragment thereof or pharmaceutical composition described herein. [0047] In some embodiments, the present disclosure provides a kit for the diagnosis, prognosis or monitoring treatment of SARS-CoV-2 in a subject, comprising: an antibody or antigen-binding fragment thereof described herein; and a least one detection reagent that binds specifically to the antibody or antigen-binding fragment thereof described herein. [0048] In some embodiments, the present disclosure provides a method of neutralizing SARS- CoV-2 in a subject, comprising administering to a subject in need thereof a therapeutically effective amount of two or more antibodies or antigen-binding fragments thereof described herein. [0049] In some embodiments, the present disclosure provides a method of treating a SARS-CoV- 2 in a subject in need thereof, comprising administering to a subject in need thereof a therapeutically effective amount of two or more antibodies or antigen-binding fragments thereof described herein. [0050] In some embodiments, the present disclosure provides a method of preventing a SARS- CoV-2 in a subject in need thereof, comprising administering to a subject in need thereof a therapeutically effective amount of two or more antibodies or antigen-binding fragments thereof described herein. [0051] In some embodiments, the two or more antibodies or antigen-binding fragments thereof are selected from: (a) an antibody or antigen-binding fragment thereof comprising: a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1354, a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1355, a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1356, a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1357, a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1358, and a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1359; (b) an antibody or antigen-binding fragment thereof comprising: a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1360, a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1361, a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1362, a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1363, a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1364, and a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1365; (c) an antibody or antigen-binding fragment thereof comprising: a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1366, a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1367, a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1368, a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1369, a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1370, and a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1371; (d) an antibody or antigen-binding fragment thereof comprising: a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1372, a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1373, a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1374, a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1375, a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1376, and a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1377; and (e) an antibody or antigen-binding fragment thereof comprising: a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1378, a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1379, a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1380, a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1381, a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1382, and a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1383. [0052] In some embodiments, the two or more monoclonal antibodies or antigen- binding fragments thereof are selected from: (a) an antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising or consisting of the amino acid sequence of SEQ ID NO: 25, and a light chain variable domain (VL) comprising or consisting of the amino acid sequence of SEQ ID NO: 26; (b) an antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising or consisting of the amino acid sequence of SEQ ID NO: 395, and a light chain variable domain (VL) comprising or consisting of the amino acid sequence of SEQ ID NO: 396; (c) an antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising or consisting of the amino acid sequence of SEQ ID NO: 397, and a light chain variable domain (VL) comprising or consisting of the amino acid sequence of SEQ ID NO: 398; (d) an antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising or consisting of the amino acid sequence of SEQ ID NO: 191, and a light chain variable domain (VL) comprising or consisting of the amino acid sequence of SEQ ID NO: 192; and (e) an antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising or consisting of the amino acid sequence of SEQ ID NO: 219, and a light chain variable domain (VL) comprising or consisting of the amino acid sequence of SEQ ID NO: 220. [0053] In some embodiments, the first antibody or antigen-binding fragment thereof and the second antibody or antigen-binding fragment thereof exhibit synergistic activity. [0054] In some embodiments, the method further comprises administering to the subject a therapeutically effective amount of a second therapeutic agent or therapy. In some embodiments, the second therapeutic agent comprises an anti-inflammatory drug or an antiviral compound. In some embodiments, (a) the antiviral compound comprises: a nucleoside analog, a peptoid, an oligopeptide, a polypeptide, a protease inhibitor, a 3C-like protease inhibitor, a papain-like protease inhibitor, or an inhibitor of an RNA dependent RNA polymerase; or (b) the antiviral compound is selected from: acyclovir ganciclovir, vidarabine, foscarnet, cidofovir, amantadine, ribavirin, trifluorothymidine, zidovudine, didanosine, zalcitabine, and an interferon, and preferably wherein the interferon is an interferon-Į^RU^DQ^LQWHUIHURQ-ȕ^ [0055] In some embodiments, the first antibody or antigen-binding fragment thereof is administered before, after, or concurrently with the second antibody or antigen-binding fragment thereof. In some embodiments, the antibody or antigen-binding fragment thereof is administered to the subject intravenously, subcutaneously, or intraperitoneally. [0056] The foregoing summary is not intended to define every aspect of the disclosure, and additional aspects are described in other sections, such as the following detailed description. The entire document is intended to be related as a unified disclosure, and it should be understood that all combinations of features described herein are contemplated, even if the combination of features are not found together in the same sentence, or paragraph, or section of this document. Other features and advantages of the invention will become apparent from the following detailed description. It should be understood, however, that the detailed description and the specific examples, while indicating specific embodiments of the disclosure, are given by way of illustration only, because various changes and modifications within the spirit and scope of the disclosure will become apparent to those skilled in the art from this detailed description. BRIEF DESCRIPTION OF THE DRAWINGS [0057] FIGs. 1a, 1b, and 1c show plasma ELISAs and neutralizing activity of the antibodies. FIG. 1a is a diagram showing blood donation schedules for vaccinated-only individuals 5 months after the 2nd dose (Vax2, top) (Cho et al., 2021), Delta breakthrough infection after Vax2 (Delta BT, 2nd from top), and vaccinated-only individuals 1 month after the 3rd dose (Vax3, 2nd from bottom)(Muecksch et al., 2022) and Omicron breakthrough infection after Vax3 (Omicron BT, bottom). FIG.1b is a graph showing area under the curve (AUG) for plasma IgG antibody binding to wild-type SARS-CoV-2 (WT) RBD after Vax2 (Cho et al., 2021), Delta BT for n=24 samples, Vax3(Muecksch et al., 2022) and Omicron BA.1 BT for n=26 samples. FIG. 1c shows plasma neutralizing activity against indicated SARS-CoV-2 variants after Vax2 (Cho et al., 2021) for n=18 samples, Delta BT for n=24 samples, Vax3(Muecksch et al., 2022) for n=18 samples and Omicron BA.1 BT for n=26 samples. WT and Omicron BA.1 NT50 values are derived from two previous reports (Gaebler et al., 2022; Schmidt et al., 2022). See Example 1 for a list of all substitutions/deletions/insertions in the spike variants. All experiments were performed at least in duplicate. The bars and values in FIGs.1a, 1b, and 1c represent geometric mean values. Statistical significance in FIG.1b and 1c was determined by a two-tailed Kruskal-Wallis test with subsequent Dunn’s multiple comparisons. [0058] FIGs.2a, 2b, 2c, 2d, and 2e show characterization of anti-SARS-CoV-2 RBD memory B cells after breakthrough infection. FIG. 2a shows representative flow cytometry plots indicating PE-WT-RBD and AlexaFluor-647-WT-RBD binding memory B cells from 4 individuals after Delta breakthrough infection following Vax2 (Delta BT), 2 individuals 1 month after Vax3, and 6 individuals after Omicron BA.1 breakthrough infection following Vax3 (Omicron BT). The number of WT RBD-specific B cells is indicated in FIG. 2b, 5 months after Vax2 (Cho et al., 2021), Delta BT (n=24), 1 month after Vax3 (Muecksch et al., 2022) and Omicron BT (n=29). c, Graphs showing the percentage of WT-, Delta-, and Omicron BA.1-RBD cross-binding B cells determined by flow cytometer in vaccinees (Vax3) and breakthrough individuals (Delta BT) or (Omicron BT) (See also in FIG. 5b). FIG. 2d depicts pie charts showing the distribution of IgG antibody sequences obtained from WT-specific memory B cells from: 2 individuals assayed sequentially 1 month after the 3rd mRNA dose (Vax3) an Omicron infection (left); 4 individuals after Delta breakthrough (Delta), and 4 individuals after Omicron breakthrough (Omicron). The number inside the circle indicates the number of sequences analyzed for the individual denoted above the circle. Pie slice size is proportional to the number of clonally related sequences. The black outline and associated numbers indicate the percentage of clonal sequences detected at each time point. Patterned slices indicate persisting clones (same IGHV and IGLV genes, with highly similar CDR3s) found at more than one timepoint within the same individual. Grey slices indicate clones unique to the timepoint. White slices indicate sequences isolated only once per time point. FIG. 2e shows number of nucleotide somatic hypermutations (SHM) in IGHV + IGLV in WT- RBD-specific sequences after Delta or Omicron breakthrough infection, compared to 5 months after Vax2, and 1 month after Vax3. The bars and numbers in FIG. 2b and FIG. 2c represent geometric mean, and in FIG. 2e, represent median values. FIG. 2e shows the results of statistical analysis in FIG. 2b and FIG. 2c that were determined by a two-tailed Kruskal-Wallis test with subsequent Dunn’s multiple-comparisons test and in FIG.2e by two-tailed Mann-Whitney test. [0059] FIGs. 3a, 3b, 3c, 3d, 3e, and 3f show characterization of anti-SARS-CoV-2 RBD monoclonal antibodies. FIG. 3a are graphs showing half-maximal effective concentration (EC50) of n=342 monoclonal antibodies measured by ELISA against WT-RBD, Delta-RBD, and Omicron BA.1-spike protein. Antibodies were obtained memory B cells after Delta breakthrough (Delta BT), after mRNA Vax3, and Omicron breakthrough (Omicron BT). FIG. 3b is a graph showing anti-SARS-CoV-2 neutralizing activity of monoclonal antibodies measured by a SARS-CoV-2 pseudotype virus neutralization assay using WT SARS-CoV-2 pseudovirus. IC50 values for all antibodies, including the 288 reported and tested herein, and 350 previously reported (Cho et al., 2021; Muecksch et al., 2022). FIGs. 3c-d are graphs showing IC50s of monoclonal antibodies against WT, Delta-RBD, and Omicron BA.1 SARS-CoV-2 pseudoviruses. Each dot represents one antibody, where 333 total antibodies were tested, including the 288 reported herein, and 455m- Vax2 antibodies previously reported (Cho et al., 2021; Muecksch et al., 2022). Values represent geometric mean values. In addition, 105 antibodies distributed over all four cohorts were also tested against Omicron BA.4/5 pseudovirus. FIG. 3e shows ring plots depicting fraction of neutralizing (IC50<1000ng/ml) antibodies against WT, Delta-RBD, and Omicron BA.1 SARS- CoV-2 pseudoviruses, and non-neutralizing (IC50>1000 ng/ml) antibodies from each time point. FIG. 3f shows ring plots depicting fraction of mAbs that are neutralizing (IC501-1000 ng/mL, white), or non-neutralizing (IC50>1000 ng/mL, black) against Omicron BA.4/5. Number in inner circles indicates number of antibodies tested. The deletions/substitutions corresponding to viral variants used in Figs. 3c-f were incorporated into a spike protein that also includes the R683G substitution, which disrupts the furin cleavage site and increases particle infectivity. Neutralizing activity against mutant pseudoviruses was compared to a wild-type (WT) SARS-CoV-2 spike sequence (NC_045512), carrying R683G where appropriate. The bars and values in FIGs. 3a, 3b, and 3d represent geometric mean values. Statistical significance in FIGs.3a and 3b was determined by a two-tailed Kruskal-Wallis test with subsequent Dunn’s multiple comparisons, in FIG.3c was determined by a two-tailed Wilcoxon test and in FIG. 3d was determined by a two-tailed Mann- Whitney test. [0060] FIGs. 4a, 4b, and 4c show the results of plasma ELISA. FIGs. 4a-b are graphs showing area under the curve (AUC) for plasma IgG binding to FIG.4a, SARS-CoV-2 Delta-RBD and FIG. 4b, Omicron-Spike for vaccinated individuals after Vax2(Cho et al., 2021), Delta breakthrough (Delta BT, n=24), and vaccinated individuals after Vax3 (Muecksch et al., 2022) and Omicron breakthrough infection after Vax3 (Omicron BT, n=26). All experiments were performed at least in duplicate and repeated twice. Bars and values represent geometric mean values. Statistical significance in FIG. 4a and FIG. 4b was determined by a two-tailed Kruskal-Wallis test with subsequent Dunn’s multiple comparisons. [0061] FIGs. 5a, 5b, 5c, 5d, 5e, and 5f show the results of flow cytometry. FIGs. 5a-b show gating strategy for phenotyping. Gating was on lymphocytes singlets that were CD20+ and CD3- CD8-CD16-Ova-. Anti-IgG, IgM, and IgA antibodies were used for B cell phenotype analysis. Antigen-specific cells were detected based on binding to WT RBD-PE+ and RBD-AF647+, or to Delta -RBD and Omicron BA.1-RBD. FIGs.5c-e are graphs showing the frequency of IgM, IgG, and IgA isotype expression in FIG. 5c, WT RBD+ MBCs, FIG. 5d, WT+Delta+ RBD binding MBCs, FIG. 5e, WT+Omicron BA.1+ RBD binding MBCs cells. FIG. 5f shows gating strategy for single-cell sorting for CD20+ B cells for WT RBD-PE and RBD-AF647. Statistical significance in FIG. 5c was determined by a two-tailed Kruskal-Wallis test with subsequent Dunn’s multiple comparisons. [0062] FIGs. 6a, 6b, 6c, 6d, 6e, 6f, 6g show frequency distribution of human V genes and antibody gene somatic hypermutations analysis. FIGs. 6a-c shows comparison of the frequency distribution of human V genes for heavy chain and light chains of anti-RBD antibodies from this study and from a database of shared clonotypes of human B cell receptor generated by Cinque Soto et al. (Soto et al., 2019). The graphs show relative abundance of human IGHV (left panel), IGKV (middle panel) and IGLV (right panel) genes in Sequence Read Archive accession SRP010970 (bottom), antibodies obtained from Delta breakthrough infection (top), and Vax2 (middle). FIGs.6d-f, same as FIG.6a. FIGs.6-c are graphs showing relative abundance of human IGHV (left panel), IGKV (middle panel), and IGLV (right panel) genes in Sequence Read Archive accession SRP010970 (bottom), antibodies obtained from Omicron BA.1 breakthrough infection (top), and Vax3 (middle). Statistical significance was determined by the two-sided binomial test. * = pİ0.05, ** = pİ0.01, *** = pİ0.001, **** = pİ0.0001. FIG. 6g shows number of nucleotide somatic hypermutations (SHM) in IGHV and IGLV in WT-RBD-specific sequences, separately after Delta or Omicron breakthrough infection, to Vax2 (Cho et al., 2021), and Vax3 (Muecksch et al., 2022). The bars and numbers in FIG.6g represent the median value. [0063] FIGs. 7a and 7b show mAb affinity and epitopes of the antibodies. FIG. 7a is a graph showing affinity measurements (KDs) for WT RBD measured by BLI for antibodies cloned from vaccinated individuals after Delta or Omicron breakthrough infection, compared to Vax2 (Cho et al., 2021), and Vax3(Muecksch et al., 2022). FIG. 7b shows the results of epitope mapping performed by competition BLI, comparing mAbs cloned from vaccinated individuals after Delta (n=48) or Omicron BA.1 (n=49) breakthrough infection, compared to Vax2 (Cho et al., 2021), and Vax3(Muecksch et al., 2022). Pie charts show the distribution of the antibody classes among all RBD-binding antibodies (upper panel), WT neutralizing antibodies only (middle panel) or non- neutralizing antibodies only (lower panel). Bars represent geometric mean values. Statistical significance was determined by using FIG.7a, by a two-tailed Kruskal Wallis test with subsequent Dunn’s multiple comparisons; FIG.7b, by a two-tailed Chi-square test. [0064] FIG.8 shows neutralizing breadth of the antibodies. Ring plots show fraction of mAbs in FIGs. 3c-e that are neutralizing (IC50 1-1000 ng/mL, white), or non-neutralizing (IC50>1000 ng/mL, black) for mutant or variant SARS-CoV-2 pseudovirus indicated across the top at the time point indicated to the left. The number inside the circle indicates the number of antibodies tested. The deletions/substitutions corresponding to viral variants were incorporated into a spike protein that also includes the R683G substitution, which disrupts the furin cleavage site and increases particle infectivity. Neutralizing activity against mutant pseudoviruses was compared to a wild- type (WT) SARS-CoV-2 spike sequence (NC_045512), carrying R683G where appropriate. All experiments were performed at least in duplicate and repeated twice. Statistical significance was determined by using a two-sided Fisher’s exact test. DETAILED DESCRIPTION OF THE INVENTION [0065] SARS-CoV-2 represents a serious public health concern. Methods to diagnose and treat persons who are infected with SARS-CoV-2 provide the opportunity to either prevent or control further spread of infection by SARS-CoV-2. These methods are especially important due to the ability of SARS-CoV-2 to infect persons through an airborne route. This invention is based, at least in part, on unexpected neutralizing activities of the disclosed anti-SARS-CoV-2 antibodies or antigen-binding fragments thereof. These antibodies and antigen-binding fragments constitute a novel therapeutic strategy in protection from SARS-CoV-2 infections. 1.1. Definitions [0066] To aid in understanding the detailed description of the compositions and methods according to the disclosure, a few express definitions are provided to facilitate an unambiguous disclosure of the various aspects of the disclosure. Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this disclosure belongs. [0067] The terms “a”, “an”, and “the”, as used herein, include plural references unless the context clearly dictates otherwise. [0068] The term “about”, as used herein, in reference to a number or range of numbers, is understood to mean the stated number and numbers +/- 10% thereof, or 10% below the lower listed limit and 10% above the higher listed limit for the values listed for a range. [0069] The term “between”, as used in a phrase as such “between A and B” or “between A-B” refers to a range including both A and B. [0070] The terms “or” and “and/or”, as used herein, include any, and all, combinations of one or more of the associated listed items. [0071] The terms “including”, “includes”, “included”, and other forms, as used herein, are not limiting. [0072] The terms “comprise” and its grammatical equivalents, as used herein, specify the presence of stated features, integers, steps, operations, elements, and/or components, but do not preclude the presence or addition of one or more other features, integers, steps, operations, elements, components, and/or groups thereof. [0073] A “neutralizing antibody” is one that can neutralize the ability of that pathogen to initiate and/or perpetuate an infection in a host and/or in target cells in vitro. “Broadly neutralizing anti- SARS-CoV-2 antibodies” refer to antibodies that neutralize more than one SARS-CoV-2 virus strains/variants in a neutralization assay. A broad neutralizing anti-SARS-CoV-2 antibody may neutralize at least 2, 3, 4, 5, 6, 7, 8, 9 or more different strains/variants of SARS-CoV-2. [0074] The term “antibody” as referred to herein includes whole antibodies and any antigen- binding fragment or single chains thereof. Whole antibodies are glycoproteins comprising at least two heavy (H) chains and two light (L) chains inter-connected by disulfide bonds. Each heavy chain is comprised of a heavy chain variable region (abbreviated herein as VH) and a heavy chain constant region. The heavy chain constant region is comprised of three domains, CH1, CH2, and CH3. Each light chain is comprised of a light chain variable region (abbreviated herein as VL) and a light chain constant region. The light chain constant region is comprised of one domain, CL. The VH and VL regions can be further subdivided into regions of hypervariability, termed complementarity determining regions (CDR), interspersed with regions that are more conserved, termed framework regions (FR). Each VH and VL is composed of three CDRs and four FRs, arranged from amino-terminus to carboxy-terminus in the following order: FR1, CDR1, FR2, CDR2, FR3, CDR3, FR4. The heavy chain variable region CDRs and FRs are HFRl, HCDRl, HFR2, HCDR2, HFR3, HCDR3, HFR4. The light chain variable region CDRs and FRs are LFR1, LCDR1, LFR2, LCDR2, LFR3, LCDR3, LFR4. The variable regions of the heavy and light chains contain a binding domain that interacts with an antigen. The constant regions of the antibodies can mediate the binding of the immunoglobulin to host tissues or factors, including various cells of the immune system (e.g., effector cells) and the first component (CIq) of the classical complement system. [0075] The term “antigen-binding fragment or portion” of an antibody (or simply “antibody fragment or portion”), as used herein, refers to one or more fragments of an antibody that retain the ability to specifically bind to an antigen (e.g., a spike or S protein of SARS-CoV-2 virus). It has been shown that the antigen-binding function of an antibody can be performed by fragments of a full-length antibody. Examples of binding fragments encompassed within the term “antigen- binding fragment or portion” of an antibody include (i) a Fab fragment, a monovalent fragment consisting of the VL, VH, CL, and CHI domains; (ii) a F(ab')2 fragment, a bivalent fragment comprising two Fab fragments linked by a disulfide bridge at the hinge region; (iii) a Fab' fragment, which is essentially a Fab with part of the hinge region (see, FUNDAMENTAL IMMUNOLOGY (Paul ed., 3rd ed. 1993)); (iv) a Fd fragment consisting of the VH and CHI domains; (v) a Fv fragment consisting of the VL and VH domains of a single arm of an antibody, (vi) a dAb fragment (Ward et al., (1989) Nature 341:544-546), which consists of a VH domain; (vii) an isolated CDR; and (viii) a nanobody, a heavy chain variable region containing a single variable domain and two constant domains. Furthermore, although the two domains of the Fv fragment, VL and VH, are coded for by separate genes, they can be joined, using recombinant methods, by a synthetic linker that enables them to be made as a single protein chain in which the VL and VH regions pair to form monovalent molecules (known as single chain Fv or scFv); see, e.g., Bird et al. (1988) Science 242:423-426; and Huston et al. (1988) Proc. Natl. Acad. Sci. USA 85:5879-5883). Such single chain antibodies are also intended to be encompassed within the term “antigen-binding fragment or portion” of an antibody. These antibody fragments are obtained using conventional techniques known to those with skill in the art, and the fragments are screened for utility in the same manner as are intact antibodies. [0076] The term “antigen” refers to a molecule or a portion of a molecule capable of being bound by an antibody or an antigen-binding fragment thereof and additionally capable of being used in an animal to produce antibodies capable of binding to an epitope of that antigen. An antigen may be a polypeptide, carbohydrate, nucleic acid, lipid, hapten, or other naturally occurring or synthetic compound. [0077] An “isolated antibody,” as used herein, is intended to refer to an antibody that is substantially free of other antibodies having different antigenic specificities (e.g., an isolated antibody that specifically binds to a spike or S protein of SARS-CoV-2 virus is substantially free of antibodies that specifically bind antigens other than the neuraminidase). An isolated antibody can be substantially free of other cellular material and/or chemicals. [0078] The terms “monoclonal antibody” or “monoclonal antibody composition” as used herein refer to a preparation of antibody molecules of single molecular composition. A monoclonal antibody composition displays a single binding specificity and affinity for a particular epitope. [0079] The term “human antibody” is intended to include antibodies having variable regions in which both the framework and CDR regions are derived from human germline immunoglobulin sequences. Furthermore, if the antibody contains a constant region, the constant region also is derived from human germline immunoglobulin sequences. The human antibodies of the invention can include amino acid residues not encoded by human germline immunoglobulin sequences (e.g., mutations introduced by random or site-specific mutagenesis in vitro or by somatic mutation in vivo). However, the term “human antibody,” as used herein, is not intended to include antibodies in which CDR sequences derived from the germline of another mammalian species, such as a mouse, have been grafted onto human framework sequences. [0080] The term “human monoclonal antibody” refers to antibodies displaying a single binding specificity, which have variable regions in which both the framework and CDR regions are derived from human germline immunoglobulin sequences. In one embodiment, the human monoclonal antibodies can be produced by a hybridoma that includes a B cell obtained from a transgenic nonhuman animal, e.g., a transgenic mouse, having a genome comprising a human heavy chain transgene and a light chain transgene fused to an immortalized cell. [0081] The term “recombinant human antibody,” as used herein, includes all human antibodies that are prepared, expressed, created, or isolated by recombinant means, such as (a) antibodies isolated from an animal (e.g., a mouse) that is transgenic or transchromosomal for human immunoglobulin genes or a hybridoma prepared therefrom (described further below), (b) antibodies isolated from a host cell transformed to express the human antibody, e.g., from a transfectoma, (c) antibodies isolated from a recombinant, combinatorial human antibody library, and (d) antibodies prepared, expressed, created or isolated by any other means that involve splicing of human immunoglobulin gene sequences to other DNA sequences. Such recombinant human antibodies have variable regions in which the framework and CDR regions are derived from human germline immunoglobulin sequences. In some embodiments, however, such recombinant human antibodies can be subjected to in vitro mutagenesis (or, when an animal transgenic for human Ig sequences is used, in vivo somatic mutagenesis) and thus the amino acid sequences of the VH and VL regions of the recombinant antibodies are sequences that, while derived from and related to human germline VH and VL sequences, may not naturally exist within the human antibody germline repertoire in vivo. [0082] The term “isotype” refers to the antibody class (e.g., IgM or IgG1) that is encoded by the heavy chain constant region genes. The phrases “an antibody recognizing an antigen” and “an antibody specific for an antigen” are used interchangeably herein with the term “an antibody which binds specifically to an antigen.” [0083] The term “human antibody derivatives” refers to any modified form of the human antibody, e.g., a conjugate of the antibody and another agent or antibody. The term “humanized antibody” is intended to refer to antibodies in which CDR sequences derived from the germline of another mammalian species, such as a mouse, have been grafted onto human framework sequences. Additional framework region modifications can be made within the human framework sequences. [0084] The term “chimeric antibody” is intended to refer to antibodies in which the variable region sequences are derived from one species, and the constant region sequences are derived from another species, such as an antibody in which the variable region sequences are derived from a mouse antibody, and the constant region sequences are derived from a human antibody. The term can also refer to an antibody in which its variable region sequence or CDR(s) is derived from one source (e.g., an IgA1 antibody), and the constant region sequence or Fc is derived from a different source (e.g., a different antibody, such as an IgG, IgA2, IgD, IgE or IgM antibody). [0085] The term “effective amount” or “therapeutically effective amount”, as used herein, refers to an amount of a therapeutic (e.g., a pharmaceutical composition provided herein) which is sufficient to treat, diagnose, prevent, delay the onset of, reduce and/or ameliorate the severity and/or duration of a given condition, disorder or disease and/or a symptom related thereto. The term also encompasses an amount necessary for the reduction, slowing, or amelioration of the advancement or progression of a given disease, reduction, slowing, or amelioration of the recurrence, development or onset of a given disease, and/or to improve or enhance the prophylactic or therapeutic effect (s) of another therapy or to serve as a bridge to another therapy. [0086] The invention encompasses isolated or substantially purified nucleic acids, peptides, polypeptides, or proteins. In the context of the present invention, an “isolated” nucleic acid, DNA or RNA molecule or an “isolated” polypeptide is a nucleic acid, DNA molecule, RNA molecule, or polypeptide that exists apart from its native environment and is therefore not a product of nature. An isolated nucleic acid, DNA molecule, RNA molecule, or polypeptide may exist in a purified form or may exist in a non-native environment such as, for example, a transgenic host cell. A “purified” nucleic acid molecule, peptide, polypeptide, or protein, or a fragment thereof, is substantially free of other cellular material, or culture medium when produced by recombinant techniques, or substantially free of chemical precursors or other chemicals when chemically synthesized. In one embodiment, an Āisolatedā nucleic acid is free of sequences that naturally flank the nucleic acid (i.e., sequences located at the 5' and 3’ ends of the nucleic acid) in the genomic DNA of the organism from which the nucleic acid is derived. For example, in various embodiments, the isolated nucleic acid molecule can contain less than about 5 kb, 4 kb, 3 kb, 2 kb, 1 kb, 0.5 kb, or 0.1 kb of nucleotide sequences that naturally flank the nucleic acid molecule in genomic DNA of the cell from which the nucleic acid is derived. A protein, peptide, or polypeptide that is substantially free of cellular material includes preparations of protein, peptide, or polypeptide having less than about 30%, 20%, 10%, or 5% (by dry weight) of contaminating protein. When the protein of the invention, or biologically active portion thereof, is recombinantly produced, preferably culture medium represents less than about 30%, 20%, 10%, or 5% (by dry weight) of chemical precursors or non-protein-of-interest chemicals. [0087] The terms “polypeptide,” “peptide,” and “protein” are used interchangeably herein to refer to polymers of amino acids of any length. The polymer may be linear or branched, it may comprise modified amino acids, and it may be interrupted by non-amino acids. The terms also encompass an amino acid polymer that has been modified; for example, disulfide bond formation, glycosylation, lipidation, acetylation, phosphorylation, pegylation, or any other manipulation, such as conjugation with a labeling component. As used herein, the term “amino acid” includes natural and/or unnatural or synthetic amino acids, including glycine and both the D or L optical isomers, and amino acid analogs and peptidomimetics. [0088] A peptide or polypeptide “fragment” as used herein refers to a less than full-length peptide, polypeptide, or protein. For example, a peptide or polypeptide fragment can have at least about 3, at least about 4, at least about 5, at least about 10, at least about 20, at least about 30, at least about 40 amino acids in length, or single unit lengths thereof. For example, fragment may be 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, or more amino acids in length. There is no upper limit to the size of a peptide fragment. However, in some embodiments, peptide fragments can be less than about 500 amino acids, less than about 400 amino acids, less than about 300 amino acids, or less than about 250 amino acids in length. Preferably the peptide fragment can elicit an immune response when used to inoculate an animal. A peptide fragment may be used to elicit an immune response by inoculating an animal with a peptide fragment in combination with an adjuvant, a peptide fragment that is coupled to an adjuvant, or a peptide fragment that is coupled to arsanilic acid, sulfanilic acid, an acetyl group, or a picryl group. A peptide fragment can include a non- amide bond and can be a peptidomimetic. [0089] As used herein, the term “conjugate” or “conjugation” or “linked” as used herein refers to the attachment of two or more entities to form one entity. A conjugate encompasses both peptide-small molecule conjugates as well as peptide-protein/peptide conjugates. [0090] The term “recombinant,” as used herein, refers to antibodies or antigen-binding fragments thereof of the invention created, expressed, isolated, or obtained by technologies or methods known in the art as recombinant DNA technology, which include, e.g., DNA splicing and transgenic expression. The term refers to antibodies expressed in a non-human mammal (including transgenic non-human mammals, e.g., transgenic mice), or a cell (e.g., CHO cells) expression system or isolated from a recombinant combinatorial human antibody library. [0091] A “nucleic acid” or “polynucleotide” refers to a DNA molecule (for example, but not limited to, a cDNA or genomic DNA) or an RNA molecule (for example, but not limited to, an mRNA), and includes DNA or RNA analogs. A DNA or RNA analog can be synthesized from nucleotide analogs. The DNA or RNA molecules may include portions that are not naturally occurring, such as modified bases, modified backbone, deoxyribonucleotides in an RNA, etc. The nucleic acid molecule can be single-stranded or double-stranded. [0092] The term “substantial identity” or “substantially identical,” when referring to a nucleic acid or fragment thereof, indicates that, when optimally aligned with appropriate nucleotide insertions or deletions with another nucleic acid (or its complementary strand), there is nucleotide sequence identity in at least about 90%, and more preferably at least about 95%, 96%, 97%, 98% or 99% of the nucleotide bases, as measured by any well-known algorithm of sequence identity, such as FASTA, BLAST or GAP, as discussed below. A nucleic acid molecule having substantial identity to a reference nucleic acid molecule may, in certain instances, encode a polypeptide having the same or substantially similar amino acid sequence as the polypeptide encoded by the reference nucleic acid molecule. [0093] As applied to polypeptides, the term “substantial similarity” or “substantially similar” means that two peptide sequences, when optimally aligned, such as by the programs GAP or BESTFIT using default gap weights, share at least 90% sequence identity, even more preferably at least 95%, 98% or 99% sequence identity. Preferably, residue positions, which are not identical, differ by conservative amino acid substitutions. A “conservative amino acid substitution” is one in which an amino acid residue is substituted by another amino acid residue having a side chain (R group) with similar chemical properties (e.g., charge or hydrophobicity). In general, a conservative amino acid substitution will not substantially change the functional properties of a protein. In cases where two or more amino acid sequences differ from each other by conservative substitutions, the percent or degree of similarity may be adjusted upwards to correct for the conservative nature of the substitution. Means for making this adjustment are well known to those of skill in the art. See, e.g., Pearson (1994) Methods Mol. Biol. 24: 307-331, which is herein incorporated by reference. Examples of groups of amino acids that have side chains with similar chemical properties include 1) aliphatic side chains: glycine, alanine, valine, leucine, and isoleucine; 2) aliphatic- hydroxyl side chains: serine and threonine; 3) amide-containing side chains: asparagine and glutamine; 4) aromatic side chains: phenylalanine, tyrosine, and tryptophan; 5) basic side chains: lysine, arginine, and histidine; 6) acidic side chains: aspartate and glutamate, and 7) sulfur-containing side chains: cysteine and methionine. Preferred conservative amino acids substitution groups are: valine-leucine-isoleucine, phenylalanine-tyrosine, lysine-arginine, alanine-valine, glutamate-aspartate, and asparagine-glutamine. Alternatively, a conservative replacement is any change having a positive value in the PAM250 log-likelihood matrix disclosed in Gonnet et al. (1992) Science 256: 1443 45, herein incorporated by reference. A “moderately conservative” replacement is any change having a nonnegative value in the PAM250 log- likelihood matrix. [0094] Sequence similarity for polypeptides is typically measured using sequence analysis software. Protein analysis software matches similar sequences using measures of similarity assigned to various substitutions, deletions, and other modifications, including conservative amino acid substitutions. For instance, GCG software contains programs such as GAP and BESTFIT, which can be used with default parameters to determine sequence homology or sequence identity between closely related polypeptides, such as homologous polypeptides from different species of organisms or between a wild-type protein and a mutein thereof. See, e.g., GCG Version 6.1. Polypeptide sequences also can be compared using FASTA with default or recommended parameters; a program in GCG Version 6.1. FASTA (e.g., FASTA2 and FASTA3) provides alignments and percent sequence identity of the regions of the best overlap between the query and search sequences (Pearson (2000) supra). Another preferred algorithm when comparing a sequence of the invention to a database containing a large number of sequences from different organisms is the computer program BLAST, especially BLASTP or TBLASTN, using default parameters. See, e.g., Altschul et al. (1990) J. Mol. Biol. 215: 403-410 and (1997) Nucleic Acids Res. 25:3389- 3402, each of which is herein incorporated by reference. [0095] As used herein, the term “affinity” refers to the strength of the sum total of noncovalent interactions between a single binding site of a molecule (e.g., an antibody) and its binding partner (e.g., an antigen). Unless indicated otherwise, as used herein, “binding affinity” refers to intrinsic binding affinity, which reflects a 1:1 interaction between members of a binding pair (e.g., antibody and antigen). The affinity of a molecule X for its partner Y can generally be represented by the dissociation constant (KD). Affinity can be measured by common methods known in the art, including those described herein. [0096] The term “specifically binds,” or “binds specifically to,” or the like, refers to an antibody that binds to a single epitope, e.g., under physiologic conditions., but which does not bind to more than one epitope. Accordingly, an antibody that specifically binds to a polypeptide will bind to an epitope that is present on the polypeptide, but which is not present on other polypeptides. Specific binding can be characterized by an equilibrium dissociation constant of at least about l x 10-8 M or less (e.g., a smaller KD denotes a tighter binding). Methods for determining whether two molecules specifically bind are well known in the art and include, for example, equilibrium dialysis, surface plasmon resonance, and the like. As described herein, antibodies have been identified by surface plasmon resonance, e.g., BIACORE™, which bind specifically to a spike or S protein of a SARS-CoV-2 virus. [0097] Preferably, the antibody binds to a spike or S protein with “high affinity,” namely with a KD of 1 X 10-7 M or less, more preferably 5 x 10-8 M or less, more preferably 3 x 10-8 M or less, more preferably 1 x 10-8 M or less, more preferably 5 x 10-9 M or less or even more preferably 1 x 10-9 M or less, as determined by surface plasmon resonance, e.g., BIACORE™. The term “does not substantially bind” to a protein or cells, as used herein, means does not bind or does not bind with a high affinity to the protein or cells, i.e., binds to the protein or cells with a KD of 1 x 10-6 M or more, more preferably 1 x 10-5 M or more, more preferably 1 x 10-4 M or more, more preferably 1 x 10-3 M or more, even more preferably 1 x 10-2 M or more. [0098] The term “Kassoc” or “Ka,” as used herein, is intended to refer to the association rate of a particular antibody-antigen interaction, whereas the term “Kdis” or “Kd,” as used herein, is intended to refer to the dissociation rate of a particular antibody-antigen interaction. The term “KD,” as used herein, is intended to refer to the dissociation constant, which is obtained from the ratio of Kd to Ka (i.e., Kd/Ka) and is expressed as a molar concentration (M). KD values for antibodies can be determined using methods well established in the art. A preferred method for determining the KD of an antibody is by using surface plasmon resonance, preferably using a biosensor system such as a BIACORE™ system. [0099] Antibodies that “compete with another antibody for binding to a target” refer to antibodies that inhibit (partially or completely) the binding of the other antibody to the target. Whether two antibodies compete with each other for binding to a target, i.e., whether and to what extent one antibody inhibits the binding of the other antibody to a target, may be determined using known competition experiments. In some embodiments, an antibody competes with, and inhibits binding of another antibody to a target by at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% or 100%. The level of inhibition or competition may be different depending on which antibody is the “blocking antibody” (i.e., the cold antibody that is incubated first with the target). Competition assays can be conducted as described, for example, in Ed Harlow and David Lane, Cold Spring Harb Protoc; 2006 or in Chapter 11 of “Using Antibodies” by Ed Harlow and David Lane, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, USA 1999. Competing antibodies bind to the same epitope, an overlapping epitope or to adjacent epitopes (e.g., as evidenced by steric hindrance). Other competitive binding assays include: solid phase direct or indirect radioimmunoassay (RIA), solid phase direct or indirect enzyme immunoassay (EIA), sandwich competition assay (see Stahli et al., Methods in Enzymology 9:242 (1983)); solid phase direct biotin-avidin EIA (see Kirkland et al., J. Immunol. 137:3614 (1986)); solid phase direct labeled assay, solid phase direct labeled sandwich assay (see Harlow and Lane, Antibodies: A Laboratory Manual, Cold Spring Harbor Press (1988)); solid phase direct label RIA using 1-125 label (see Morel et al., Mol. Immunol. 25(1):7 (1988)); solid phase direct biotin-avidin EIA (Cheung et al., Virology 176:546 (1990)); and direct labeled RIA. (Moldenhauer et al., Scand. J. Immunol. 32:77 (1990)). [00100] The term “epitope” as used herein refers to an antigenic determinant that interacts with a specific antigen-binding site in the variable region of an antibody molecule known as a paratope. A single antigen may have more than one epitope. Thus, different antibodies may bind to different areas on an antigen and may have different biological effects. The term “epitope” also refers to a site on an antigen to which B and/or T cells respond. It also refers to a region of an antigen that is bound by an antibody. Epitopes may be defined as structural or functional. Functional epitopes are generally a subset of the structural epitopes and have those residues that directly contribute to the affinity of the interaction. Epitopes may also be conformational, that is, composed of non- linear amino acids. In some embodiments, epitopes may include determinants that are chemically active surface groupings of molecules such as amino acids, sugar side chains, phosphoryl groups, or sulfonyl groups, and, In some embodiments, may have specific three-dimensional structural characteristics, and/or specific charge characteristics. An epitope typically includes at least 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15 amino acids in a unique spatial conformation. Methods for determining what epitopes are bound by a given antibody (i.e., epitope mapping) are well known in the art and include, for example, immunoblotting and immune-precipitation assays, wherein overlapping or contiguous peptides from a spike or S protein are tested for reactivity with a given antibody. Methods of determining spatial conformation of epitopes include techniques in the art and those described herein, for example, x-ray crystallography and 2-dimensional nuclear magnetic resonance (see, e.g., Epitope Mapping Protocols in Methods in Molecular Biology, Vol. 66, G. E. Morris, Ed. (1996)). [00101] The term “epitope mapping” refers to the process of identification of the molecular determinants for antibody-antigen recognition. [00102] The term “binds to an epitope” or “recognizes an epitope” with reference to an antibody or antibody fragment refers to continuous or discontinuous segments of amino acids within an antigen. Those of skill in the art understand that the terms do not necessarily mean that the antibody or antibody fragment is in direct contact with every amino acid within an epitope sequence. [00103] The term “binds to the same epitope” with reference to two or more antibodies means that the antibodies bind to the same, overlapping, or encompassing continuous or discontinuous segments of amino acids. Those of skill in the art understand that the phrase “binds to the same epitope” does not necessarily mean that the antibodies bind to or contact exactly the same amino acids. The precise amino acids that the antibodies contact can differ. For example, a first antibody can bind to a segment of amino acids that is completely encompassed by the segment of amino acids bound by a second antibody. In another example, a first antibody binds one or more segments of amino acids that significantly overlap the one or more segments bound by the second antibody. For the purposes herein, such antibodies are considered to “bind to the same epitope.” [00104] As used herein, the term “immune response” refers to a biological response within a vertebrate against foreign agents, which response protects the organism against these agents and diseases caused by them. An immune response is mediated by the action of a cell of the immune system (for example, a T lymphocyte, B lymphocyte, natural killer (NK) cell, macrophage, eosinophil, mast cell, dendritic cell, or neutrophil) and soluble macromolecules produced by any of these cells or the liver (including antibodies, cytokines, and complement) that results in selective targeting, binding to, damage to, destruction of, and/or elimination from the vertebrate's body of invading pathogens, cells or tissues infected with pathogens, cancerous or other abnormal cells, or, in cases of autoimmunity or pathological inflammation, normal human cells or tissues. An immune reaction includes, e.g., activation or inhibition of a T cell, e.g., an effector T cell or a Th cell, such as a CD4+ or CD8+ T cell, or the inhibition of a Treg cell. [00105] The term “detectable label” as used herein refers to a molecule capable of detection, including, but not limited to, radioactive isotopes, fluorescers, chemiluminescers, chromophores, enzymes, enzyme substrates, enzyme cofactors, enzyme inhibitors, chromophores, dyes, metal ions, metal sols, ligands (e.g., biotin, avidin, streptavidin or haptens), intercalating dyes and the like. The term “fluorescer” refers to a substance or a portion thereof that is capable of exhibiting fluorescence in the detectable range. [00106] In many embodiments, the terms “subject” and “patient” are used interchangeably irrespective of whether the subject has or is currently undergoing any form of treatment. As used herein, the terms “subject” and “subjects” may refer to any vertebrate, including, but not limited to, a mammal (e.g., cow, pig, camel, llama, horse, goat, rabbit, sheep, hamsters, guinea pig, cat, dog, rat, and mouse, a non-human primate (for example, a monkey, such as a cynomolgus monkey, chimpanzee, etc.) and a human). The subject may be a human or a non-human. In more exemplary aspects, the mammal is a human. As used herein, the expression “a subject in need thereof” or “a patient in need thereof” means a human or non-human mammal that exhibits one or more symptoms or indications of disorders (e.g., neuronal disorders, autoimmune diseases, and cardiovascular diseases), and/or who has been diagnosed with inflammatory disorders. In some embodiments, the subject is a mammal. In some embodiments, the subject is human. [00107] The term “host cell”, as used herein, refers to a particular subject cell into which an exogenous nucleic acid molecule may be introduced and the progeny or potential progeny of such a cell. Progeny of such a cell may not be identical to the parent cell comprising the nucleic acid molecule due to mutations or environmental influences that may occur in succeeding generations or integration of the nucleic acid molecule into the host cell genome. [00108] As used herein, the term “disease” is intended to be generally synonymous and is used interchangeably with the terms “disorder” and “condition” (as in medical condition), in that all reflect an abnormal condition (e.g., inflammatory disorder) of the human or animal body or of one of its parts that impairs normal functioning, is typically manifested by distinguishing signs and symptoms, and causes the human or animal to have a reduced duration or quality of life. [00109] As used herein, the term “treating” or “treatment” of any disease or disorder refers in one embodiment, to ameliorating the disease or disorder (i.e., arresting or reducing the development of the disease or at least one of the clinical symptoms thereof). In another embodiment, “treating” or “treatment” refers to ameliorating at least one physical parameter, which may not be discernible by the patient. In yet another embodiment, “treating” or “treatment” refers to modulating the disease or disorder, either physically, (e.g., stabilization of a discernible symptom), physiologically, (e.g., stabilization of a physical parameter), or both. In yet another embodiment, “treating” or “treatment” refers to preventing or delaying the onset or development or progression of the disease or disorder. [00110] The terms “prevent,” “preventing,” “prevention,” “prophylactic treatment” and the like refer to reducing the probability of developing a disorder or condition in a subject, who does not have, but is at risk of or susceptible to developing a disorder or condition. [00111] The terms “decrease,” “reduced,” “reduction,” “decrease,” or “inhibit” are all used herein generally to mean a decrease by a statistically significant amount. However, for avoidance of doubt, “reduced,” “reduction,” “decrease,” or “inhibit” means a decrease by at least 10% as compared to a reference level, for example, a decrease by at least about 20%, or at least about 30%, or at least about 40%, or at least about 50%, or at least about 60%, or at least about 70%, or at least about 80%, or at least about 90% or up to and including a 100% decrease (e.g., absent level as compared to a reference sample), or any decrease between 10-100% as compared to a reference level. [00112] As used herein, the term “agent” denotes a chemical compound, a mixture of chemical compounds, a biological macromolecule (such as a nucleic acid, an antibody, a protein or portion thereof, e.g., a peptide), or an extract made from biological materials such as bacteria, plants, fungi, or animal (particularly mammalian) cells or tissues. The activity of such agents may render it suitable as a “therapeutic agent,” which is a biologically, physiologically, or pharmacologically active substance (or substances) that acts locally or systemically in a subject. [00113] As used herein, the terms “therapeutic agent,” “therapeutic capable agent,” or “treatment agent” are used interchangeably and refer to a molecule or compound that confers some beneficial effect upon administration to a subject. The beneficial effect includes enablement of diagnostic determinations; amelioration of a disease, symptom, disorder, or pathological condition; reducing or preventing the onset of a disease, symptom, disorder, or condition; and generally counteracting a disease, symptom, disorder or pathological condition. [00114] The term “therapeutic effect” is art-recognized and refers to a local or systemic effect in animals, particularly mammals, and more particularly humans caused by a pharmacologically active substance. [00115] The term “effective amount,” “effective dose,” or “effective dosage” is defined as an amount sufficient to achieve or at least partially achieve a desired effect. A “therapeutically effective amount” or “therapeutically effective dosage” of a drug or therapeutic agent is any amount of the drug that, when used alone or in combination with another therapeutic agent, promotes disease regression evidenced by a decrease in severity of disease symptoms, an increase in frequency and duration of disease symptom-free periods, or a prevention of impairment or disability due to the disease affliction. A “prophylactically effective amount” or a “prophylactically effective dosage” of a drug is an amount of the drug that, when administered alone or in combination with another therapeutic agent to a subject at risk of developing a disease or of suffering a recurrence of disease, inhibits the development or recurrence of the disease. The ability of a therapeutic or prophylactic agent to promote disease regression or inhibit the development or recurrence of the disease can be evaluated using a variety of methods known to the skilled practitioner, such as in human subjects during clinical trials, in animal model systems predictive of efficacy in humans, or by assaying the activity of the agent in in vitro assays. [00116] Doses are often expressed in relation to bodyweight. Thus, a dose which is expressed as [g, mg, or other unit]/kg (or g, mg etc.) usually refers to [g, mg, or other unit] “per kg (or g, mg etc.) bodyweight,” even if the term “bodyweight” is not explicitly mentioned. [00117] As used herein, the term “composition” or “pharmaceutical composition” refers to a mixture of at least one component useful within the invention with other components, such as carriers, stabilizers, diluents, dispersing agents, suspending agents, thickening agents, and/or excipients. The pharmaceutical composition facilitates administration of one or more components of the invention to an organism. [00118] As used herein, the term “pharmaceutically acceptable” refers to a material, such as a carrier or diluent, which does not abrogate the biological activity or properties of the composition, and is relatively non-toxic, i.e., the material may be administered to an individual without causing undesirable biological effects or interacting in a deleterious manner with any of the components of the composition in which it is contained. [00119] As used herein, the term “pharmaceutically acceptable carrier” includes a pharmaceutically acceptable salt, pharmaceutically acceptable material, composition or carrier, such as a liquid or solid filler, diluent, excipient, solvent or encapsulating material, involved in carrying or transporting a compound(s) of the present invention within or to the subject such that it may perform its intended function. Typically, such compounds are carried or transported from one organ, or portion of the body, to another organ, or portion of the body. Each salt or carrier must be “acceptable” in the sense of being compatible with the other ingredients of the formulation and not injurious to the subject. Some examples of materials that may serve as pharmaceutically acceptable carriers include: sugars, such as lactose, glucose, and sucrose; starches, such as corn starch and potato starch; cellulose, and its derivatives, such as sodium carboxymethyl cellulose, ethyl cellulose, and cellulose acetate; powdered tragacanth; malt; gelatin; talc; excipients, such as cocoa butter and suppository waxes; oils, such as peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil, and soybean oil; glycols, such as propylene glycol; polyols, such as glycerin, sorbitol, mannitol, and polyethylene glycol; esters, such as ethyl oleate and ethyl laurate; agar; buffering agents, such as magnesium hydroxide and aluminum hydroxide; alginic acid; pyrogen- free water; isotonic saline; Ringer’s solution; ethyl alcohol; phosphate buffer solutions; diluent; granulating agent; lubricant; binder; disintegrating agent; wetting agent; emulsifier; coloring agent; release agent; coating agent; sweetening agent; flavoring agent; perfuming agent; preservative; antioxidant; plasticizer; gelling agent; thickener; hardener; setting agent; suspending agent; surfactant; humectant; carrier; stabilizer; and other non-toxic compatible substances employed in pharmaceutical formulations, or any combination thereof. As used herein, “pharmaceutically acceptable carrier” also includes any and all coatings, antibacterial and antifungal agents, and absorption delaying agents, and the like that are compatible with the activity of one or more components of this disclosure, and are physiologically acceptable to the subject. Supplementary active compounds may also be incorporated into the compositions. [00120] “Combination” therapy, as used herein, unless otherwise clear from the context, is meant to encompass administration of two or more therapeutic agents in a coordinated fashion and includes, but is not limited to, concurrent dosing. Specifically, combination therapy encompasses both co-administration (e.g., administration of a co-formulation or simultaneous administration of separate therapeutic compositions) and serial or sequential administration, provided that administration of one therapeutic agent is conditioned in some way on the administration of another therapeutic agent. For example, one therapeutic agent may be administered only after a different therapeutic agent has been administered and allowed to act for a prescribed period of time. See, e.g., Kohrt et al. (2011) Blood 117:2423. [00121] As used herein, the term “co-administration” or “co-administered” refers to the administration of at least two agent(s) or therapies to a subject. In some embodiments, the co- administration of two or more agents/therapies is concurrent. In other embodiments, a first agent/therapy is administered prior to a second agent/therapy. Those of skill in the art understand that the formulations and/or routes of administration of the various agents/therapies used may vary. [00122] As used herein, the term “contacting,” when used in reference to any set of components, includes any process whereby the components to be contacted are mixed into the same mixture (for example, are added into the same compartment or solution), and does not necessarily require actual physical contact between the recited components. The recited components can be contacted in any order or any combination (or sub-combination) and can include situations where one or some of the recited components are subsequently removed from the mixture, optionally prior to addition of other recited components. For example, “contacting A with B and C” includes any and all of the following situations: (i) A is mixed with C, then B is added to the mixture; (ii) A and B are mixed into a mixture; B is removed from the mixture, and then C is added to the mixture; and (iii) A is added to a mixture of B and C. [00123] “Sample,” “test sample,” and “patient sample” may be used interchangeably herein. The sample can be a sample of serum, urine plasma, amniotic fluid, cerebrospinal fluid, cells, or tissue. Such a sample can be used directly as obtained from a patient or can be pre-treated, such as by filtration, distillation, extraction, concentration, centrifugation, inactivation of interfering components, addition of reagents, and the like, to modify the character of the sample in some manner as discussed herein or otherwise as is known in the art. The terms “sample” and “biological sample” as used herein generally refer to a biological material being tested for and/or suspected of containing an analyte of interest, such as antibodies. The sample may be any tissue sample from the subject. The sample may comprise protein from the subject. [00124] As used herein, the term “in vitro” refers to events that occur in an artificial environment, e.g., in a test tube or reaction vessel, in cell culture, etc., rather than within a multi-cellular organism. [00125] As used herein, the term “in vivo” refers to events that occur within a multi-cellular organism, such as a non-human animal. [00126] As used herein, the singular forms “a,” “an,” and “the” include plural references unless the context clearly dictates otherwise. [00127] As used herein, the terms “including,” “comprising,” “containing,” or “having” and variations thereof are meant to encompass the items listed thereafter and equivalents thereof as well as additional subject matter unless otherwise noted. [00128] As used herein, the phrases “in one embodiment,” “in various embodiments,” “in some embodiments,” and the like are used repeatedly. Such phrases do not necessarily refer to the same embodiment, but they may unless the context dictates otherwise. [00129] As used herein, the terms “and/or” or “/” means any one of the items, any combination of the items, or all of the items with which this term is associated. [00130] As used herein, the word “substantially” does not exclude “completely,” e.g., a composition that is “substantially free” from Y may be completely free from Y. Where necessary, the word “substantially” may be omitted from the definition of the invention. [00131] As used herein, the term “each,” when used in reference to a collection of items, is intended to identify an individual item in the collection but does not necessarily refer to every item in the collection. Exceptions can occur if explicit disclosure or context clearly dictates otherwise. [00132] As used herein, the term “approximately” or “about,” as applied to one or more values of interest, refers to a value that is similar to a stated reference value. In some embodiments, the term “approximately” or “about” refers to a range of values that fall within 25%, 20%, 19%, 18%, 17%, 16%, 15%, 14%, 13%, 12%, 11%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, or less in either direction (greater than or less than) of the stated reference value unless otherwise stated or otherwise evident from the context (except where such number would exceed 100% of a possible value). Unless indicated otherwise herein, the term “about” is intended to include values, e.g., weight percents, proximate to the recited range that are equivalent in terms of the functionality of the individual ingredient, the composition, or the embodiment. [00133] As disclosed herein, a number of ranges of values are provided. It is understood that each intervening value, to the tenth of the unit of the lower limit, unless the context clearly dictates otherwise, between the upper and lower limits of that range is also specifically disclosed. Each smaller range between any stated value or intervening value in a stated range and any other stated or intervening value in that stated range is encompassed within the invention. The upper and lower limits of these smaller ranges may independently be included or excluded in the range, and each range where either, neither, or both limits are included in the smaller ranges is also encompassed within the invention, subject to any specifically excluded limit in the stated range. Where the stated range includes one or both of the limits, ranges excluding either or both of those included limits are also included in the invention. [00134] The use of any and all examples, or exemplary language (e.g., “such as”) provided herein, is intended merely to better illuminate the invention and does not pose a limitation on the scope of the invention unless otherwise claimed. No language in the specification should be construed as indicating any non-claimed element as essential to the practice of the invention. [00135] All methods described herein are performed in any suitable order unless otherwise indicated herein or otherwise clearly contradicted by context. In regard to any of the methods provided, the steps of the method may occur simultaneously or sequentially. When the steps of the method occur sequentially, the steps may occur in any order, unless noted otherwise. In cases in which a method comprises a combination of steps, each and every combination or sub-combination of the steps is encompassed within the scope of the disclosure, unless otherwise noted herein. [00136] Each publication, patent application, patent, and other reference cited herein is incorporated by reference in its entirety to the extent that it is not inconsistent with the present disclosure. Publications disclosed herein are provided solely for their disclosure prior to the filing date of the present invention. Nothing herein is to be construed as an admission that the present invention is not entitled to antedate such publication by virtue of prior invention. Further, the dates of publication provided may be different from the actual publication dates, which may need to be independently confirmed. [00137] It is understood that the examples and embodiments described herein are for illustrative purposes only and that various modifications or changes in light thereof will be suggested to persons skilled in the art and are to be included within the spirit and purview of this application and scope of the appended claims. 1.2. Antibodies and Antigen-binding fragments [00138] In one aspect, this disclosure provides novel isolated and recombinantly modified anti- SARS-CoV-2 antibodies or antigen-binding fragments thereof. In some embodiments, antibodies or antigen-binding fragments thereof described herein are broadly neutralizing antibodies that neutralize multiple SARS-CoV-2 virus strains. The antibodies are able to protect a subject prophylactically and therapeutically against a lethal challenge with a SARS-CoV-2 virus. [00139] In one aspect, this disclosure provides an isolated anti-SARS-CoV-2 antibody or antigen-binding fragment thereof that binds specifically to a SARS-CoV-2 antigen. In some embodiments, the SARS-CoV-2 antigen comprises a portion of a spike (S) polypeptide, such as a S polypeptide of a human or an animal SARS-CoV-2. In some embodiments, the antibodies described herein are monoclonal antibodies. [00140] In some embodiments, the monoclonal antibodies, or antigen-binding fragments thereof, specifically bind to the S polypeptide of a SARS-CoV-2, or a domain of the spike protein. In some embodiments, the monoclonal antibodies or antigen-binding fragments thereof bind to the receptor binding domain (RBD) of the S polypeptide (also referred to herein as the Spike protein). In some embodiments, the monoclonal antibodies or antigen-binding fragments thereof bind to the N- terminal domain (NTD) of the Spike protein. In some embodiments, the monoclonal antibodies or antigen-binding fragments thereof bind to the S2 domain of the Spike protein. [00141] In some embodiments, the monoclonal antibodies or antigen-binding fragments thereof bind to the RBD of the Spike protein (e.g., amino acids 319-541 of SEQ ID NO: 1353). In some embodiments, the monoclonal antibodies or antigen-binding fragments thereof bind to the N- terminal domain (NTD) (e.g., residues 14–305 of SEQ ID NO: 1353) of the S polypeptide. In some embodiments, the monoclonal antibodies or antigen-binding fragments thereof bind to the supersite b-hairpin (residues 152-158 of SEQ ID NO: 1353). In some embodiments, the monoclonal antibodies or antigen-binding fragments thereof bind to WKH^ȕ^-strand (residue 97-102 of SEQ ID NO: 1353), N4-loop (residues 178-188 of SEQ ID NO: 1353), and/or N-linked glycans at positions N122 and N149 of the S polypeptide. In some embodiments, the monoclonal antibodies or antigen-binding fragments thereof bind to residues 27-32, 57-60, 210-218, and/or 286-303 of the S polypeptide of SEQ ID NO: 1353. In some embodiments, the monoclonal antibodies or antigen-binding fragments thereof bind to residues 600-606 of the S polypeptide of SEQ ID NO: 1353. [00142] In some embodiments, the antibody or antigen-binding fragment thereof is capable of neutralizing a SARS-CoV-2 virus at an IC50 concentration of less than 50 (e.g., 1, 5, 10, 20, 30, 40, 50) μg/ml. [00143] The monoclonal antibodies or antigen-binding fragment thereof disclosed herein can be used for detecting and treating SARS-CoV-2. For instance, they can be useful in a cocktail approach designed to minimize the chances of escape that can readily occur with single antibody approaches. [00144] The spike protein is important because it is present on the outside of intact SARS-CoV- 2. Thus, it presents a target that can be used to inhibit or eliminate an intact virus before the virus has an opportunity to infect a cell. A representative amino acid sequence is provided below: [00145]
Figure imgf000047_0002
Figure imgf000047_0001
Figure imgf000048_0001
[00146] The total length of SARS-CoV-2 S is 1273 amino acids and consists of a signal peptide (amino acids 1–13) located at the N-terminus, the S1 subunit (14–685 residues), and the S2 subunit (686–1273 residues); the last two regions are responsible for receptor binding and membrane fusion, respectively. In the S1 subunit, there is an N-terminal domain (14–305 residues) and a receptor-binding domain (RBD, 319–541 residues); the fusion peptide (FP) (788–806 residues), heptapeptide repeat sequence 1 (HR1) (912–984 residues), HR2 (1163–1213 residues), TM domain (1213–1237 residues), and cytoplasm domain (1237–1273 residues) comprise the S2 subunit. S protein trimers visually form a characteristic bulbous, crown-like halo surrounding the viral particle. Based on the structure of coronavirus S protein monomers, the S1 and S2 subunits form the bulbous head and stalk region. [00147] Listed below in Table 2 are amino acid sequences of the heavy chain (HC) variable regions (HCVR) and light chain (LC) variable regions (LCVR) of exemplary antibodies. “HVCR” is also referred to herein as “VH” and LCVR is also referred to herein as “VL”. [00148] In some embodiments, the antibody or antigen-binding fragment thereof comprises: (i) a heavy chain variable region having an amino acid sequence with at least 75% (e.g., 75%, 78%, 80%, 82%, 84%, 85%, 86%, 88%, 90%, 92%, 94%, 95%, 96%, 97%, 98%, 99%) identity to one selected from those in Table 2; and/or (ii) a light chain variable region having an amino acid sequence with at least 75% (e.g., 75%, 78%, 80%, 82%, 84%, 85%, 86%, 88%, 90%, 92%, 94%, 95%, 96%, 97%, 98%, 99%) identity to one selected from those in Table 2. [00149] In some embodiments, the antibody or antigen-binding fragment thereof comprises: (i) a heavy chain variable region having an amino acid sequence selected from those in Table 2; and/or (ii) a light chain variable region having an amino acid sequence selected from those in Table 2. [00150] In some embodiments, the antibody or antigen-binding fragment thereof comprises a heavy chain variable region and a light chain variable region that comprises an amino acid sequence pair selected from those in Table 2. [00151] In some embodiments, the antibody or antigen-binding fragment thereof comprises three heavy chain complementarity determining regions (HCDRs) (HCDR1, HCDR2, and HCDR3) of a heavy chain variable region having the amino acid sequence of SEQ ID NO: 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31, 33, 35, 37, 39, 41, 43, 45, 47, 49, 51, 53, 55, 57, 59, 61, 63, 65, 67, 69, 71, 73, 75, 77, 79, 81, 83, 85, 87, 89, 91, 93, 95, 97, 99, 101, 103, 105, 107, 109, 111, 113, 115, 117, 119, 121, 123, 125, 127, 129, 131, 133, 135, 137, 139, 141, 143, 145, 147, 149, 151, 153, 155, 157, 159, 161, 163, 165, 167, 169, 171, 173, 175, 177, 179, 181, 183, 185, 187, 189, 191, 193, 195, 197, 199, 201, 203, 205, 207, 209, 211, 213, 215, 217, 219, 221, 223, 225, 227, 229, 231, 233, 235, 237, 239, 241, 243, 245, 247, 249, 251, 253, 255, 257, 259, 261, 263, 265, 267, 269, 271, 273, 275, 277, 279, 281, 283, 285, 287, 289, 291, 293, 295, 297, 299, 301, 303, 305, 307, 309, 311, 313, 315, 317, 319, 321, 323, 325, 327, 329, 331, 333, 335, 337, 339, 341, 343, 345, 347, 349, 351, 353, 355, 357, 359, 361, 363, 365, 367, 369, 371, 373, 375, 377, 379, 381, 383, 385, 387, 389, 391, 393, 395, 397, 399, 401, 403, 405, 407, 409, 411, 413, 415, 417, 419, 421, 423, 425, 427, 429, 431, 433, 435, 437, 439, 441, 443, 445, 447, 449, 451, 453, 455, 457, 459, 461, 463, 465, 467, 469, 471, 473, 475, 477, 479, 481, 483, 485, 487, 489, 491, 493, 495, 497, 499, 501, 503, 505, 507, 509, 511, 513, 515, 517, 519, 521, 523, 525, 527, 529, 531, 533, 535, 537, 539, 541, 543, 545, 547, 549, 551, 553, 555, 557, 559, 561, 563, 565, 567, 569, 571, 573, 575, 577, 579, 581, 583, 585, 587, 589, 591, 593, 595, 597, 599, 601, 603, 605, 607, 609, 611, 613, 615, 617, 619, 621, 623, 625, 627, 629, 631, 633, 635, 637, 639, 641, 643, 645, 647, 649, 651, 653, 655, 657, 659, 661, 663, 665, 667, 669, 671, 673, or 675; and three light chain CDRs (LCDR1, LCDR2, and LCDR3) of a light chain variable region having the amino acid sequence of SEQ ID NO: 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 44, 46, 48, 50, 52, 54, 56, 58, 60, 62, 64, 66, 68, 70, 72, 74, 76, 78, 80, 82, 84, 86, 88, 90, 92, 94, 96, 98, 100, 102, 104, 106, 108, 110, 112, 114, 116, 118, 120, 122, 124, 126, 128, 130, 132, 134, 136, 138, 140, 142, 144, 146, 148, 150, 152, 154, 156, 158, 160, 162, 164, 166, 168, 170, 172, 174, 176, 178, 180, 182, 184, 186, 188, 190, 192, 194, 196, 198, 200, 202, 204, 206, 208, 210, 212, 214, 216, 218, 220, 222, 224, 226, 228, 230, 232, 234, 236, 238, 240, 242, 244, 246, 248, 250, 252, 254, 256, 258, 260, 262, 264, 266, 268, 270, 272, 274, 276, 278, 280, 282, 284, 286, 288, 290, 292, 294, 296, 298, 300, 302, 304, 306, 308, 310, 312, 314, 316, 318, 320, 322, 324, 326, 328, 330, 332, 334, 336, 338, 340, 342, 344, 346, 348, 350, 352, 354, 356, 358, 360, 362, 364, 366, 368, 370, 372, 374, 376, 378, 380, 382, 384, 386, 388, 390, 392, 394, 396, 398, 400, 402, 404, 406, 408, 410, 412, 414, 416, 418, 420, 422, 424, 426, 428, 430, 432, 434, 436, 438, 440, 442, 444, 446, 448, 450, 452, 454, 456, 458, 460, 462, 464, 466, 468, 470, 472, 474, 476, 478, 480, 482, 484, 486, 488, 490, 492, 494, 496, 498, 500, 502, 504, 506, 508, 510, 512, 514, 516, 518, 520, 522, 524, 526, 528, 530, 532, 534, 536, 538, 540, 542, 544, 546, 548, 550, 552, 554, 556, 558, 560, 562, 564, 566, 568, 570, 572, 574, 576, 578, 580, 582, 584, 586, 588, 590, 592, 594, 596, 598, 600, 602, 604, 606, 608, 610, 612, 614, 616, 618, 620, 622, 624, 626, 628, 630, 632, 634, 636, 638, 640, 642, 644, 646, 648, 650, 652, 654, 656, 658, 660, 662, 664, 666, 668, 670, 672, 674, or 676. [00152] A person of ordinary skill in the art will understand that various CDR numbering schemes (such as the Kabat, Chothia, Enhanced Chothia, IMGT, AHoAbM, Contact numbering schemes) can be used to determine CDR positions. [00153] In some embodiments, the antibody or antigen-binding fragment thereof comprises: a heavy chain variable region comprising an amino acid sequence with at least 75% (e.g., 75%, 78%, 80%, 82%, 84%, 85%, 86%, 88%, 90%, 92%, 94%, 95%, 96%, 97%, 98%, 99%) identity to the amino acid sequence of SEQ ID NO: 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31, 33, 35, 37, 39, 41, 43, 45, 47, 49, 51, 53, 55, 57, 59, 61, 63, 65, 67, 69, 71, 73, 75, 77, 79, 81, 83, 85, 87, 89, 91, 93, 95, 97, 99, 101, 103, 105, 107, 109, 111, 113, 115, 117, 119, 121, 123, 125, 127, 129, 131, 133, 135, 137, 139, 141, 143, 145, 147, 149, 151, 153, 155, 157, 159, 161, 163, 165, 167, 169, 171, 173, 175, 177, 179, 181, 183, 185, 187, 189, 191, 193, 195, 197, 199, 201, 203, 205, 207, 209, 211, 213, 215, 217, 219, 221, 223, 225, 227, 229, 231, 233, 235, 237, 239, 241, 243, 245, 247, 249, 251, 253, 255, 257, 259, 261, 263, 265, 267, 269, 271, 273, 275, 277, 279, 281, 283, 285, 287, 289, 291, 293, 295, 297, 299, 301, 303, 305, 307, 309, 311, 313, 315, 317, 319, 321, 323, 325, 327, 329, 331, 333, 335, 337, 339, 341, 343, 345, 347, 349, 351, 353, 355, 357, 359, 361, 363, 365, 367, 369, 371, 373, 375, 377, 379, 381, 383, 385, 387, 389, 391, 393, 395, 397, 399, 401, 403, 405, 407, 409, 411, 413, 415, 417, 419, 421, 423, 425, 427, 429, 431, 433, 435, 437, 439, 441, 443, 445, 447, 449, 451, 453, 455, 457, 459, 461, 463, 465, 467, 469, 471, 473, 475, 477, 479, 481, 483, 485, 487, 489, 491, 493, 495, 497, 499, 501, 503, 505, 507, 509, 511, 513, 515, 517, 519, 521, 523, 525, 527, 529, 531, 533, 535, 537, 539, 541, 543, 545, 547, 549, 551, 553, 555, 557, 559, 561, 563, 565, 567, 569, 571, 573, 575, 577, 579, 581, 583, 585, 587, 589, 591, 593, 595, 597, 599, 601, 603, 605, 607, 609, 611, 613, 615, 617, 619, 621, 623, 625, 627, 629, 631, 633, 635, 637, 639, 641, 643, 645, 647, 649, 651, 653, 655, 657, 659, 661, 663, 665, 667, 669, 671, 673, or 675; or comprising the amino acid sequence of SEQ ID NO: 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31, 33, 35, 37, 39, 41, 43, 45, 47, 49, 51, 53, 55, 57, 59, 61, 63, 65, 67, 69, 71, 73, 75, 77, 79, 81, 83, 85, 87, 89, 91, 93, 95, 97, 99, 101, 103, 105, 107, 109, 111, 113, 115, 117, 119, 121, 123, 125, 127, 129, 131, 133, 135, 137, 139, 141, 143, 145, 147, 149, 151, 153, 155, 157, 159, 161, 163, 165, 167, 169, 171, 173, 175, 177, 179, 181, 183, 185, 187, 189, 191, 193, 195, 197, 199, 201, 203, 205, 207, 209, 211, 213, 215, 217, 219, 221, 223, 225, 227, 229, 231, 233, 235, 237, 239, 241, 243, 245, 247, 249, 251, 253, 255, 257, 259, 261, 263, 265, 267, 269, 271, 273, 275, 277, 279, 281, 283, 285, 287, 289, 291, 293, 295, 297, 299, 301, 303, 305, 307, 309, 311, 313, 315, 317, 319, 321, 323, 325, 327, 329, 331, 333, 335, 337, 339, 341, 343, 345, 347, 349, 351, 353, 355, 357, 359, 361, 363, 365, 367, 369, 371, 373, 375, 377, 379, 381, 383, 385, 387, 389, 391, 393, 395, 397, 399, 401, 403, 405, 407, 409, 411, 413, 415, 417, 419, 421, 423, 425, 427, 429, 431, 433, 435, 437, 439, 441, 443, 445, 447, 449, 451, 453, 455, 457, 459, 461, 463, 465, 467, 469, 471, 473, 475, 477, 479, 481, 483, 485, 487, 489, 491, 493, 495, 497, 499, 501, 503, 505, 507, 509, 511, 513, 515, 517, 519, 521, 523, 525, 527, 529, 531, 533, 535, 537, 539, 541, 543, 545, 547, 549, 551, 553, 555, 557, 559, 561, 563, 565, 567, 569, 571, 573, 575, 577, 579, 581, 583, 585, 587, 589, 591, 593, 595, 597, 599, 601, 603, 605, 607, 609, 611, 613, 615, 617, 619, 621, 623, 625, 627, 629, 631, 633, 635, 637, 639, 641, 643, 645, 647, 649, 651, 653, 655, 657, 659, 661, 663, 665, 667, 669, 671, 673, or 675; and/or a light chain variable region comprising an amino acid sequence with at least 75% (e.g., 75%, 78%, 80%, 82%, 84%, 85%, 86%, 88%, 90%, 92%, 94%, 95%, 96%, 97%, 98%, 99%) identity to the amino acid sequence of SEQ ID NO: 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 44, 46, 48, 50, 52, 54, 56, 58, 60, 62, 64, 66, 68, 70, 72, 74, 76, 78, 80, 82, 84, 86, 88, 90, 92, 94, 96, 98, 100, 102, 104, 106, 108, 110, 112, 114, 116, 118, 120, 122, 124, 126, 128, 130, 132, 134, 136, 138, 140, 142, 144, 146, 148, 150, 152, 154, 156, 158, 160, 162, 164, 166, 168, 170, 172, 174, 176, 178, 180, 182, 184, 186, 188, 190, 192, 194, 196, 198, 200, 202, 204, 206, 208, 210, 212, 214, 216, 218, 220, 222, 224, 226, 228, 230, 232, 234, 236, 238, 240, 242, 244, 246, 248, 250, 252, 254, 256, 258, 260, 262, 264, 266, 268, 270, 272, 274, 276, 278, 280, 282, 284, 286, 288, 290, 292, 294, 296, 298, 300, 302, 304, 306, 308, 310, 312, 314, 316, 318, 320, 322, 324, 326, 328, 330, 332, 334, 336, 338, 340, 342, 344, 346, 348, 350, 352, 354, 356, 358, 360, 362, 364, 366, 368, 370, 372, 374, 376, 378, 380, 382, 384, 386, 388, 390, 392, 394, 396, 398, 400, 402, 404, 406, 408, 410, 412, 414, 416, 418, 420, 422, 424, 426, 428, 430, 432, 434, 436, 438, 440, 442, 444, 446, 448, 450, 452, 454, 456, 458, 460, 462, 464, 466, 468, 470, 472, 474, 476, 478, 480, 482, 484, 486, 488, 490, 492, 494, 496, 498, 500, 502, 504, 506, 508, 510, 512, 514, 516, 518, 520, 522, 524, 526, 528, 530, 532, 534, 536, 538, 540, 542, 544, 546, 548, 550, 552, 554, 556, 558, 560, 562, 564, 566, 568, 570, 572, 574, 576, 578, 580, 582, 584, 586, 588, 590, 592, 594, 596, 598, 600, 602, 604, 606, 608, 610, 612, 614, 616, 618, 620, 622, 624, 626, 628, 630, 632, 634, 636, 638, 640, 642, 644, 646, 648, 650, 652, 654, 656, 658, 660, 662, 664, 666, 668, 670, 672, 674, or 676; or comprising the amino acid sequence of SEQ ID NO: 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 44, 46, 48, 50, 52, 54, 56, 58, 60, 62, 64, 66, 68, 70, 72, 74, 76, 78, 80, 82, 84, 86, 88, 90, 92, 94, 96, 98, 100, 102, 104, 106, 108, 110, 112, 114, 116, 118, 120, 122, 124, 126, 128, 130, 132, 134, 136, 138, 140, 142, 144, 146, 148, 150, 152, 154, 156, 158, 160, 162, 164, 166, 168, 170, 172, 174, 176, 178, 180, 182, 184, 186, 188, 190, 192, 194, 196, 198, 200, 202, 204, 206, 208, 210, 212, 214, 216, 218, 220, 222, 224, 226, 228, 230, 232, 234, 236, 238, 240, 242, 244, 246, 248, 250, 252, 254, 256, 258, 260, 262, 264, 266, 268, 270, 272, 274, 276, 278, 280, 282, 284, 286, 288, 290, 292, 294, 296, 298, 300, 302, 304, 306, 308, 310, 312, 314, 316, 318, 320, 322, 324, 326, 328, 330, 332, 334, 336, 338, 340, 342, 344, 346, 348, 350, 352, 354, 356, 358, 360, 362, 364, 366, 368, 370, 372, 374, 376, 378, 380, 382, 384, 386, 388, 390, 392, 394, 396, 398, 400, 402, 404, 406, 408, 410, 412, 414, 416, 418, 420, 422, 424, 426, 428, 430, 432, 434, 436, 438, 440, 442, 444, 446, 448, 450, 452, 454, 456, 458, 460, 462, 464, 466, 468, 470, 472, 474, 476, 478, 480, 482, 484, 486, 488, 490, 492, 494, 496, 498, 500, 502, 504, 506, 508, 510, 512, 514, 516, 518, 520, 522, 524, 526, 528, 530, 532, 534, 536, 538, 540, 542, 544, 546, 548, 550, 552, 554, 556, 558, 560, 562, 564, 566, 568, 570, 572, 574, 576, 578, 580, 582, 584, 586, 588, 590, 592, 594, 596, 598, 600, 602, 604, 606, 608, 610, 612, 614, 616, 618, 620, 622, 624, 626, 628, 630, 632, 634, 636, 638, 640, 642, 644, 646, 648, 650, 652, 654, 656, 658, 660, 662, 664, 666, 668, 670, 672, 674, or 676. [00154] In some embodiments, the antibody or antigen-binding fragment thereof comprises a heavy chain variable region (HCVR) and a light chain variable region (LCVR) that comprise a HCVR/LCVR amino acid sequence pair of SEQ ID NOs: 1-2, 3-4, 5-6, 7-8, 9-10, 11-12, 13-14, 15-16, 17-18, 19-20, 21-22, 23-24, 25-26, 27-28, 29-30, 31-32, 33-34, 35-36, 37-38, 39-40, 41-42, 43-44, 45-46, 47-48, 49-50, 51-52, 53-54, 55-56, 57-58, 59-60, 61-62, 63-64, 65-66, 67-68, 69-70, 71-72, 73-74, 75-76, 77-78, 79-80, 81-82, 83-84, 85-86, 87-88, 89-90, 91-92, 93-94, 95-96, 97-98, 99-100, 101-102, 103-104, 105-106, 107-108, 109-110, 111-112, 113-114, 115-116, 117-118, 119-120, 121-122, 123-124, 125-126, 127-128, 129-130, 131-132, 133-134, 135-136, 137-138, 139-140, 141-142, 143-144, 145-146, 147-148, 149-150, 151-152, 153-154, 155-156, 157-158, 159-160, 161-162, 163-164, 165-166, 167-168, 169-170, 171-172, 173-174, 175-176, 177-178, 179-180, 181-182, 183-184, 185-186, 187-188, 189-190, 191-192, 193-194, 195-196, 197-198, 199-200, 201-202, 203-204, 205-206, 207-208, 209-210, 211-212, 213-214, 215-216, 217-218, 219-220, 221-222, 223-224, 225-226, 227-228, 229-230, 231-232, 233-234, 235-236, 237-238, 239-240, 241-242, 243-244, 245-246, 247-248, 249-250, 251-252, 253-254, 255-256, 257-258, 259-260, 261-262, 263-264, 265-266, 267-268, 269-270, 271-272, 273-274, 275-276, 277-278, 279-280, 281-282, 283-284, 285-286, 287-288, 289-290, 291-292, 293-294, 295-296, 297-298, 299-300, 301-302, 303-304, 305-306, 307-308, 309-310, 311-312, 313-314, 315-316, 317-318, 319-320, 321-322, 323-324, 325-326, 327-328, 329-330, 331-332, 333-334, 335-336, 337-338, 339-340, 341-342, 343-344, 345-346, 347-348, 349-350, 351-352, 353-354, 355-356, 357-358, 359-360, 361-362, 363-364, 365-366, 367-368, 369-370, 371-372, 373-374, 375-376, 377-378, 379-380, 381-382, 383-384, 385-386, 387-388, 389-390, 391-392, 393-394, 395-396, 397-398, 399-400, 401-402, 403-404, 405-406, 407-408, 409-410, 411-412, 413-414, 415-416, 417-418, 419-420, 421-422, 423-424, 425-426, 427-428, 429-430, 431-432, 433-434, 435-436, 437-438, 439-440, 441-442, 443-444, 445-446, 447-448, 449-450, 451-452, 453-454, 455-456, 457-458, 459-460, 461-462, 463-464, 465-466, 467-468, 469-470, 471-472, 473-474, 475-476, 477-478, 479-480, 481-482, 483-484, 485-486, 487-488, 489-490, 491-492, 493-494, 495-496, 497-498, 499-500, 501-502, 503-504, 505-506, 507-508, 509-510, 511-512, 513-514, 515-516, 517-518, 519-520, 521-522, 523-524, 525-526, 527-528, 529-530, 531-532, 533-534, 535-536, 537-538, 539-540, 541-542, 543-544, 545-546, 547-548, 549-550, 551-552, 553-554, 555-556, 557-558, 559-560, 561-562, 563-564, 565-566, 567-568, 569-570, 571-572, 573-574, 575-576, 577-578, 579-580, 581-582, 583-584, 585-586, 587-588, 589-590, 591-592, 593-594, 595-596, 597-598, 599-600, 601-602, 603-604, 605-606, 607-608, 609-610, 611-612, 613-614, 615-616, 617-618, 619-620, 621-622, 623-624, 625-626, 627-628, 629-630, 631-632, 633-634, 635-636, 637-638, 639-640, 641-642, 643-644, 645-646, 647-648, 649-650, 651-652, 653-654, 655-656, 657-658, 659-660, 661-662, 663-664, 665-666, 667-668, 669-670, 671-672, 673-674, or 675-676. [00155] In some embodiments, the antibody is selected from B1030, B1032, B2014, B2058, and B2117 antibodies, as listed in Table 2. Sequences for exemplary antibodies are provided in Table 3 below: Table 3: Exemplary Antibody Sequences
Figure imgf000053_0001
Figure imgf000054_0001
[00156] In some embodiments, the antibody or antigen-binding fragment thereof comprises a variable heavy chain CDR1 (HCDR1) comprising an amino acid sequence of SEQ ID NO: 1354. In some embodiments, the antibody or antigen-binding fragment thereof comprises a HCDR1 comprising an amino acid sequence of SEQ ID NO:1360. In some embodiments, the antibody or antigen-binding fragment thereof comprises a HCDR1 comprising an amino acid sequence of SEQ ID NO:1366. In some embodiments, the antibody or antigen-binding fragment thereof comprises a HCDR1 comprising an amino acid sequence of SEQ ID NO:1372. In some embodiments, the antibody or antigen-binding fragment thereof comprises a HCDR1 comprising an amino acid sequence of SEQ ID NO:1378. In some embodiments, the HCDR1 comprises an amino acid sequence that is at least 75% or at least 87% identical to one of SEQ ID NOs: 1354, 1360, 1366, 1372, or 1378. In some embodiments, the HCDR1 comprises an amino acid sequence that is 100% identical to one of SEQ ID NOs: 1354, 1360, 1366, 1372, or 1378 except for the substitution or deletion of one or two amino acids. [00157] In some embodiments, the antibody or antigen-binding fragment thereof comprises a variable heavy chain CDR2 HCDR2 comprising an amino acid sequence of SEQ ID NO: 1355. In some embodiments, the antibody or antigen-binding fragment thereof comprises a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1361. In some embodiments, the antibody or antigen-binding fragment thereof comprises a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1367. In some embodiments, the antibody or antigen-binding fragment thereof comprises a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1373. In some embodiments, the antibody or antigen-binding fragment thereof comprises a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1379. In some embodiments, the antibody or antigen-binding fragment thereof comprises a HCDR2 comprising an amino acid sequence that is at least 71%, at least 75%, at least 85%, or at least 87% identical to one of SEQ ID NOs: 1355, 1361, 1367, 1373, or 1379. In some embodiments, the HCDR2 comprises an amino acid sequence that is at least 100% identical to one of SEQ ID NOs: 1355, 1361, 1367, 1373, or 1379 except for the substitution or deletion of one or two amino acids. [00158] In some embodiments, the antibody or antigen-binding fragment thereof comprises a variable heavy chain CDR3 (HCDR3) comprising an amino acid sequence of SEQ ID NO: 1356. In some embodiments, the antibody or antigen-binding fragment thereof comprises a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1362. In some embodiments, the antibody or antigen-binding fragment thereof comprises a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1368. In some embodiments, the antibody or antigen-binding fragment thereof comprises a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1374. In some embodiments, the antibody or antigen-binding fragment thereof comprises a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1380. In some embodiments, the antibody or antigen-binding fragment thereof comprises a HCDR3 comprising an amino acid sequence that is at least at least 75%, at least 81%, at least 85%, at least 86%, at least 87%, at least 89%, at least 93%, at least 94%, or at least 95% identical to one of SEQ ID NOs: 1356, 1362, 1368, 1374, or 1380. In some embodiments, the HCDR3 comprises an amino acid sequence that is at least 100% identical to one of SEQ ID NOs: 1356, 1362, 1368, 1374, or 1380 except for the substitution or deletion of one or two amino acids. [00159] In some embodiments, the antibody or antigen-binding fragment thereof comprises a variable light chain CDR1 (LCDR1) comprising an amino acid sequence of SEQ ID NO: 1357. In some embodiments, the antibody or antigen-binding fragment thereof comprises a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1363. In some embodiments, the antibody or antigen-binding fragment thereof comprises a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1369. In some embodiments, the antibody or antigen-binding fragment thereof comprises a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1375. In some embodiments, the antibody or antigen-binding fragment thereof comprises a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1381. In some embodiments, the antibody or antigen-binding fragment thereof comprises an LCDR1 comprises an amino acid sequence that is at least 77% or at least 88% identical to one of SEQ ID NOs: 1357, 1363, 1369, 1375, or 1381. In some embodiments, the LCDR1 comprises an amino acid sequence that is at least 100% identical to one of SEQ ID NOs: 1357, 1363, 1369, 1375, or 1381 except for the substitution or deletion of one or two amino acids. [00160] In some embodiments, the antibody or antigen-binding fragment thereof comprises a variable light chain CDR2 (LCDR2) comprising an amino acid sequence of SEQ ID NO: 1358. In some embodiments, the antibody or antigen-binding fragment thereof comprises a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1364. In some embodiments, the antibody or antigen-binding fragment thereof comprises a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1370. In some embodiments, the antibody or antigen-binding fragment thereof comprises a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1379. In some embodiments, the antibody or antigen-binding fragment thereof comprises a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1382. In some embodiments, the antibody or antigen-binding fragment thereof comprises an LCDR2 comprises an amino acid sequence that is at least 66% identical to one of SEQ ID NOs: 1358, 1364, 1370, 1376, or 1382. In some embodiments, the LCDR2 comprises an amino acid sequence that is at least 100% identical to one of SEQ ID NOs: 1358, 1364, 1370, 1376, or 1382 except for the substitution or deletion of one amino acid. [00161] In some embodiments, the antibody or antigen-binding fragment thereof comprises a variable light chain CDR3 (LCDR3) comprising an amino acid sequence of SEQ ID NO: 1359. In some embodiments, the antibody or antigen-binding fragment thereof comprises a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1365. In some embodiments, the antibody or antigen-binding fragment thereof comprises a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1371. In some embodiments, the antibody or antigen-binding fragment thereof comprises a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1377. In some embodiments, the antibody or antigen-binding fragment thereof comprises a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1383. In some embodiments, the antibody or antigen-binding fragment thereof comprises an LCDR3 comprises an amino acid sequence that is at least 50%, at least 75%, at least 77%, at least 81%, at least 84%, at least 87%, at least 88%, at least 90%, or at least 92% identical to one of SEQ ID NOs: 1359, 1365, 1371, 1377, or 1383. In some embodiments, the LCDR3 comprises an amino acid sequence that is at least 100% identical to one of SEQ ID NOs: 1359, 1365, 1371, 1377, or 1383 except for the substitution or deletion of one or two amino acids. [00162] In some embodiments, the antibody or antigen-binding fragment thereof comprises: 1) a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1354, 1360, 1366, 1372, or 1378; 2) a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1355, 1361, 1367, 1373, or 1379; 3) a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1356, 1362, 1368, 1374, or 1380; 4) a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1357, 1363, 1369, 1375, or 1381; 5) a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1358, 1364, 1370, 1376, or 1382; and 6) a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1359, 1365, 1371, 1377, or 1383. [00163] In some embodiments, the antibody or antigen-binding fragment thereof comprises a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1354, a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1355, a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1356, a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1357, a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1358, and a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1359. In some embodiments, the antibody or antigen-binding fragment thereof comprises a HCDR1 consisting of SEQ ID NO: 1354, a HCDR2 consisting of SEQ ID NO: 1355, a HCDR3 consisting of SEQ ID NO: 1356, a LCDR1 consisting of SEQ ID NO: 1357, a LCDR2 consisting of SEQ ID NO: 1358, and a LCDR3 consisting of SEQ ID NO: 1359. [00164] In some embodiments, the antibody or antigen-binding fragment thereof comprises a VH comprising an amino acid sequence that is at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% identical to SEQ ID NO: 25. In some embodiments, the antibody or antigen-binding fragment thereof comprises a VH comprising an amino acid sequence of SEQ ID NO: 25. In some embodiments, the antibody or antigen-binding fragment thereof comprises a VH consisting of SEQ ID NO: 25. In some embodiments, the antibody or antigen-binding fragment thereof comprises a VL comprising an amino acid sequence that is at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% identical to SEQ ID NO: 26. In some embodiments, the antibody or antigen-binding fragment thereof comprises a VL comprising an amino acid sequence of SEQ ID NO: 26. In some embodiments, the antibody or antigen-binding fragment thereof comprises a VH consisting of SEQ ID NO: 26. [00165] In some embodiments, the antibody or antigen-binding fragment thereof comprises a VH comprising an amino acid sequence that is at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% identical to SEQ ID NO: 25 and a VL comprising an amino acid sequence that is at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% identical to SEQ ID NO: 26. In some embodiments, the antibody or antigen-binding fragment thereof comprises a VH comprising an amino acid sequence of SEQ ID NO: 25 and a VL comprising an amino acid sequence of SEQ ID NO: 26. In some embodiments, the antibody or antigen-binding fragment thereof comprises a VL consisting of SEQ ID NO: 25 and a VL consisting of SEQ ID NO: 26. [00166] In some embodiments, the antibody or antigen-binding fragment thereof comprises a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1360, a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1361, a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1362, a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1363, a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1364, and a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1365. In some embodiments, the antibody or antigen-binding fragment thereof comprises a HCDR1 consisting of SEQ ID NO: 1360, a HCDR2 consisting of SEQ ID NO: 1361, a HCDR3 consisting of SEQ ID NO: 1362, a LCDR1 consisting of SEQ ID NO: 1363, a LCDR2 consisting of SEQ ID NO: 1364, and a LCDR3 consisting of SEQ ID NO: 1365. [00167] In some embodiments, the antibody or antigen-binding fragment thereof comprises a VH comprising an amino acid sequence that is at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% identical to SEQ ID NO: 395. In some embodiments, the antibody or antigen-binding fragment thereof comprises a VH comprising an amino acid sequence of SEQ ID NO: 395. In some embodiments, the antibody or antigen-binding fragment thereof comprises a VH consisting of SEQ ID NO: 395. In some embodiments, the antibody or antigen-binding fragment thereof comprises a VL comprising an amino acid sequence that is at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% identical to SEQ ID NO: 396. In some embodiments, the antibody or antigen-binding fragment thereof comprises a VL comprising an amino acid sequence of SEQ ID NO: 396. In some embodiments, the antibody or antigen- binding fragment thereof comprises a VL consisting of SEQ ID NO: 396. [00168] In some embodiments, the antibody or antigen-binding fragment thereof comprises a VH comprising an amino acid sequence that is at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% identical to SEQ ID NO: 395 and a VL comprising an amino acid sequence that is at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% identical to SEQ ID NO: 396. In some embodiments, the antibody or antigen-binding fragment thereof comprises a VH comprising an amino acid sequence of SEQ ID NO: 395 and a VL comprising an amino acid sequence of SEQ ID NO: 396. In some embodiments, the antibody or antigen-binding fragment thereof comprises a VH consisting of SEQ ID NO: 395 and a VL consisting of SEQ ID NO: 396. [00169] In some embodiments, the antibody or antigen-binding fragment thereof comprises a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1366, a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1367, a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1368, a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1369, a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1370, and a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1371. In some embodiments, the antibody or antigen-binding fragment thereof comprises a HCDR1 consisting of SEQ ID NO: 1366, a HCDR2 consisting of SEQ ID NO: 1367, a HCDR3 consisting of SEQ ID NO: 1368, a LCDR1 consisting of SEQ ID NO: 1369, a LCDR2 consisting of SEQ ID NO: 1370, and a LCDR3 consisting of SEQ ID NO: 1371. [00170] In some embodiments, the antibody or antigen-binding fragment thereof comprises a VH comprising an amino acid sequence that is at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% identical to SEQ ID NO: 397. In some embodiments, the antibody or antigen-binding fragment thereof comprises a VH comprising an amino acid sequence of SEQ ID NO: 397. In some embodiments, the antibody or antigen-binding fragment thereof comprises a VH consisting of SEQ ID NO: 397. In some embodiments, the antibody or antigen-binding fragment thereof comprises a VL comprising an amino acid sequence that is at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% identical to SEQ ID NO: 398. In some embodiments, the antibody or antigen-binding fragment thereof comprises a VL comprising an amino acid sequence of SEQ ID NO: 398. In some embodiments, the antibody or antigen- binding fragment thereof comprises a VL consisting of SEQ ID NO: 398. [00171] In some embodiments, the antibody or antigen-binding fragment thereof comprises a VH comprising an amino acid sequence that is at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% identical to SEQ ID NO: 397 and a VL comprising an amino acid sequence that is at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% identical to SEQ ID NO: 398. In some embodiments, the antibody or antigen-binding fragment thereof comprises a VH comprising an amino acid sequence of SEQ ID NO: 397 and a VL comprising an amino acid sequence of SEQ ID NO: 398. In some embodiments, the antibody or antigen-binding fragment thereof comprises a VH consisting of SEQ ID NO: 397 and a VL consisting of SEQ ID NO: 398. [00172] In some embodiments, the antibody or antigen-binding fragment thereof comprises a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1372, a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1373, a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1374, a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1375, a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1376, and a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1377. In some embodiments, the antibody or antigen-binding fragment thereof comprises a HCDR1 consisting of SEQ ID NO: 1372, a HCDR2 consisting of SEQ ID NO: 1373, a HCDR3 consisting of SEQ ID NO: 1374, a LCDR1 consisting of SEQ ID NO: 1375, a LCDR2 consisting of SEQ ID NO: 1376, and a LCDR3 consisting of SEQ ID NO: 1377. [00173] In some embodiments, the antibody or antigen-binding fragment thereof comprises a VH comprising an amino acid sequence that is at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% identical to SEQ ID NO: 191. In some embodiments, the antibody or antigen-binding fragment thereof comprises a VH comprising an amino acid sequence of SEQ ID NO: 191. In some embodiments, the antibody or antigen-binding fragment thereof comprises a VH consisting of SEQ ID NO: 191. In some embodiments, the antibody or antigen-binding fragment thereof comprises a VL comprising an amino acid sequence that is at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% identical to SEQ ID NO: 192. In some embodiments, the antibody or antigen-binding fragment thereof comprises a VL comprising an amino acid sequence of SEQ ID NO: 192. In some embodiments, the antibody or antigen- binding fragment thereof comprises a VL consisting of SEQ ID NO: 192. [00174] In some embodiments, the antibody or antigen-binding fragment thereof comprises a VH comprising an amino acid sequence that is at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% identical to SEQ ID NO: 191 and a VL comprising an amino acid sequence that is at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% identical to SEQ ID NO: 192. In some embodiments, the antibody or antigen-binding fragment thereof comprises a VH comprising an amino acid sequence of SEQ ID NO: 191 and a VL comprising an amino acid sequence of SEQ ID NO: 192. In some embodiments, the antibody or antigen-binding fragment thereof comprises a VH consisting of SEQ ID NO: 191 and a VL consisting of SEQ ID NO: 192. [00175] In some embodiments, the antibody or antigen-binding fragment thereof comprises a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1378, a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1379, a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1380, a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1381, a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1382, and a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1383. In some embodiments, the antibody or antigen-binding fragment thereof comprises a HCDR1 consisting of SEQ ID NO: 1378, a HCDR2 consisting of SEQ ID NO: 1379, a HCDR3 consisting of SEQ ID NO: 1380, a LCDR1 consisting of SEQ ID NO: 1381, a LCDR2 consisting of SEQ ID NO: 1382, and a LCDR3 consisting of SEQ ID NO: 1383. [00176] In some embodiments, the antibody or antigen-binding fragment thereof comprises a VH comprising an amino acid sequence that is at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% identical to SEQ ID NO: 219. In some embodiments, the antibody or antigen-binding fragment thereof comprises a VH comprising an amino acid sequence of SEQ ID NO: 219. In some embodiments, the antibody or antigen-binding fragment thereof comprises a VH consisting of SEQ ID NO: 219. In some embodiments, the antibody or antigen-binding fragment thereof comprises a VL comprising an amino acid sequence that is at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% identical to SEQ ID NO: 220. In some embodiments, the antibody or antigen-binding fragment thereof comprises a VL comprising an amino acid sequence of SEQ ID NO: 220. In some embodiments, the antibody or antigen- binding fragment thereof comprises a VL consisting of SEQ ID NO: 220. [00177] In some embodiments, the antibody or antigen-binding fragment thereof comprises a VH comprising an amino acid sequence that is at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% identical to SEQ ID NO: 219 and a VL comprising an amino acid sequence that is at least about 70%, about 75%, about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% identical to SEQ ID NO: 220. In some embodiments, the antibody or antigen-binding fragment thereof comprises a VH comprising an amino acid sequence of SEQ ID NO: 219 and a VL comprising an amino acid sequence of SEQ ID NO: 220. In some embodiments, the antibody or antigen-binding fragment thereof comprises a VH consisting of SEQ ID NO: 219 and a VL consisting of SEQ ID NO: 220. [00178] In some embodiments, the antibody or antigen-binding fragment thereof binds to an epitope within the N-terminal domain (NTD) of the S polypeptide. In some embodiments, the antibody or antigen-binding fragment thereof binds to an epitope within the receptor binding domain (RBD) of the S polypeptide. In some embodiments, the antibody or antigen-binding fragment thereof binds to an epitope within the subdomain (SD) of the S polypeptide. In some embodiments, the antibody or antigen-binding fragment thereof binds to an epitope within the S2 subunit of the S polypeptide. [00179] In some embodiments, the antibody or antigen-binding fragment thereof binds to an epitope outside of the N-terminal domain (NTD) of the S polypeptide. In some embodiments, the antibody or antigen-binding fragment thereof binds to an epitope outside of the receptor binding domain (RBD) of the S polypeptide. In some embodiments, the antibody or antigen-binding fragment thereof binds to an epitope outside of the subdomain (SD) of the S polypeptide. In some embodiments, the antibody or antigen-binding fragment thereof binds to an epitope outside of the S2 subunit of the S polypeptide. [00180] In some embodiments, the antibody or antigen-binding fragment thereof is a neutralizing antibody or antigen-binding fragment thereof. In some embodiments, the antibody or antigen-binding fragment thereof neutralizes SARS-CoV-2. In some embodiments, the antibody or antigen-binding fragment thereof inhibits the fusion of SARS-CoV-2 and host cell membrane. [00181] In some embodiments, the antibody or antigen-binding fragment thereof is a non- neutralizing antibody or antigen-binding fragment thereof. In some embodiments, the antibody or antigen-binding fragment thereof is cytotoxic to a SARS-CoV-2 infected host cell. [00182] In some embodiments, the monoclonal antibodies, or antigen-binding fragments thereof, described herein are an IgG1, IgG2, IgG3, or IgG4 isotype. In some embodiments, the monoclonal antibodies, or antigen-binding fragments thereof, described herein are IgG1 isotypes. [00183] In some embodiments, the antibody or antigen-binding fragment thereof comprises (a) a first target binding site that specifically binds to an epitope within the S polypeptide, and (b) a second target binding site that binds to a different epitope on the S polypeptide or on a different molecule. In some embodiments, the multivalent antibody is a bivalent or bispecific antibody. [00184] In some embodiments, the antibody or the antigen-binding fragment thereof further comprises a variant Fc constant region. In some embodiments, the antibody is a monoclonal antibody. In some embodiments, the antibody is a chimeric antibody, a humanized antibody, or a humanized monoclonal antibody. In some embodiments, the antibody or antigen-binding fragment thereof is a single-chain antibody, Fab or Fab2 fragment. [00185] In some embodiments, the antibody or the antigen-binding fragment thereof further comprises a variant Fc constant region. The antibody can be a monoclonal antibody. In some embodiments, the antibody can be a chimeric antibody, a humanized antibody, or a humanized monoclonal antibody. In some embodiments, the antibody or antigen-binding fragment thereof can be a single-chain antibody, Fab or Fab2 fragment. [00186] In some embodiments, the antibody or antigen-binding fragment thereof can be detectably labeled or conjugated to a toxin, a therapeutic agent, a polymer (e.g., polyethylene glycol (PEG)), a receptor, an enzyme, or a receptor ligand. For example, an antibody of the present invention may be coupled to a toxin (e.g., a tetanus toxin). Such antibodies may be used to treat animals, including humans, that are infected with the virus that is etiologically linked to SARS- CoV-2. The toxin-coupled antibody is thought to bind to a portion of a spike protein presented on an infected cell, and then kill the infected cell. [00187] In another example, an antibody of the present invention may be coupled to a detectable tag. Such antibodies may be used within diagnostic assays to determine if an animal, such as a human, is infected with SARS-CoV-2. Examples of detectable tags include fluorescent proteins (i.e., green fluorescent protein, red fluorescent protein, yellow fluorescent protein), fluorescent markers (i.e., fluorescein isothiocyanate, rhodamine, texas red), radiolabels (i.e., 3H, 32P, 125I), enzymes (i.e.^^ȕ-JDODFWRVLGDVH^^KRUVHUDGLVK^SHUR[LGDVH^^ȕ-glucuronidase, alkaline phosphatase), or an affinity tag (i.e., avidin, biotin, streptavidin). Methods to couple antibodies to a detectable tag are known in the art. Harlow et al., Antibodies: A Laboratory Manual, page 319 (Cold Spring Harbor Pub.1988). Fragment [00188] In some embodiments, an antibody provided herein is an antibody fragment. Antibody fragments include, but are not limited to, Fab, Fab', Fab'-SH, F(ab')2, Fv, and single-chain Fv (scFv) fragments, and other fragments described below, e.g., diabodies, triabodies tetrabodies, and single- domain antibodies. For a review of certain antibody fragments, see Hudson et al., Nat. Med.9:129- 134 (2003). For a review of scFv fragments, see, e.g., Pluckthun, in The Pharmacology of Monoclonal Antibodies, vol. 113, Rosenburg and Moore eds., (Springer-Verlag, New York), pp. 269-315 (1994); see also WO 93/16185; and U.S. Pat. Nos. 5,571,894 and 5,587,458. For discussion of Fab and F(ab')2 fragments comprising salvage receptor binding epitope residues and having increased in vivo half-life, see U.S. Pat. No.5,869,046. [00189] Diabodies are antibody fragments with two antigen-binding sites that may be bivalent or bispecific. See, for example, EP 404,097; WO 1993/01161; Hudson et al., Nat. Med. 9:129- 134 (2003); and Hollinger et al., Proc. Natl. Acad. Sci. USA 90: 6444-6448 (1993). Triabodies and tetrabodies are also described in Hudson et al., Nat. Med.9:129-134 (2003). [00190] Single-domain antibodies are antibody fragments comprising all or a portion of the heavy chain variable domain or all or a portion of the light chain variable domain of an antibody. In some embodiments, a single-domain antibody is a human single-domain antibody (DOMANTIS, Inc., Waltham, Mass.; see, e.g., U.S. Pat. No.6,248,516). [00191] Antibody fragments can be made by various techniques, including but not limited to proteolytic digestion of an intact antibody as well as production by recombinant host cells (e.g., E. coli or phage), as described herein. Chimeric and Humanized Antibodies [00192] In some embodiments, an antibody provided herein is a chimeric antibody. Certain chimeric antibodies are described, e.g., in U.S. Pat. No.4,816,567; and Morrison et al., Proc. Natl. Acad. Sci. USA, 81:6851-6855 (1984)). In one example, a chimeric antibody comprises a non- human variable region (e.g., a variable region derived from a mouse, rat, hamster, rabbit, or non- human primate, such as a monkey) and a human constant region. In a further example, a chimeric antibody is a “class switched” antibody in which the class or subclass has been changed from that of the parent antibody. Chimeric antibodies include antigen-binding fragments thereof. [00193] In some embodiments, a chimeric antibody is a humanized antibody. Typically, a non- human antibody is humanized to reduce immunogenicity to humans, while retaining the specificity and affinity of the parental non-human antibody. Generally, a humanized antibody comprises one or more variable domains in which HVRs, e.g., CDRs, (or portions thereof) are derived from a non-human antibody, and FRs (or portions thereof) are derived from human antibody sequences. A humanized antibody optionally will also comprise at least a portion of a human constant region. In some embodiments, some FR residues in a humanized antibody are substituted with corresponding residues from a non-human antibody (e.g., the antibody from which the HVR residues are derived), e.g., to restore or improve antibody specificity or affinity. [00194] Humanized antibodies and methods of making them are reviewed, e.g., in Almagro and Fransson, Front. Biosci.13:1619-1633 (2008), and are further described, e.g., in Riechmann et al., Nature 332:323-329 (1988); Queen et al., Proc. Nat'l Acad. Sci. USA 86:10029-10033 (1989); U.S. Pat. Nos. 5,821,337, 7,527,791, 6,982,321, and 7,087,409; Kashmiri et al., Methods 36:25-34 (2005) (describing specificity determining region (SDR) grafting); Padlan, Mol. Immunol.28:489- 498 (1991) (describing “resurfacing”); Dall'Acqua et al., Methods 36:43-60 (2005) (describing “FR shuffling”); and Osbourn et al., Methods 36:61-68 (2005) and Klimka et al., Br. J. Cancer, 83:252-260 (2000) (describing the “guided selection” approach to FR shuffling). [00195] Human framework regions that may be used for humanization include but are not limited to: framework regions selected using the “best-fit” method (see, e.g., Sims et al. J. Immunol. 151:2296 (1993)); framework regions derived from the consensus sequence of human antibodies of a particular subgroup of light or heavy chain variable regions (see, e.g., Carter et al. Proc. Natl. Acad. Sci. USA, 89:4285 (1992); and Presta et al. J. Immunol., 151:2623 (1993)); human mature (somatically mutated) framework regions or human germline framework regions (see, e.g., Almagro and Fransson, Front. Biosci.13:1619-1633 (2008)); and framework regions derived from screening FR libraries (see, e.g., Baca et al., J. Biol. Chem.272:10678-10684 (1997) and Rosok et al., J. Biol. Chem.271:22611-22618 (1996)). Human Antibodies [00196] In some embodiments, an antibody provided herein is a human antibody. Human antibodies can be produced using various techniques known in the art or using techniques described herein. Human antibodies are described generally in van Dijk and van de Winkel, Curr. Opin. Pharmacol.5: 368-74 (2001) and Lonberg, Curr. Opin. Immunol.20:450-459 (2008). [00197] Human antibodies may be prepared by administering an immunogen to a transgenic animal that has been modified to produce intact human antibodies or intact antibodies with human variable regions in response to antigenic challenge. Such animals typically contain all or a portion of the human immunoglobulin loci, which replace the endogenous immunoglobulin loci, or which are present extrachromosomally or integrated randomly into the animal's chromosomes. In such transgenic mice, the endogenous immunoglobulin loci have generally been inactivated. For review of methods for obtaining human antibodies from transgenic animals, see Lonberg, Nat. Biotech. 23:1117-1125 (2005). See also, e.g., U.S. Pat. Nos. 6,075,181 and 6,150,584 describing XENOMOUSE™ technology; U.S. Pat. No.5,770,429 describing HUMAB technology; U.S. Pat. No.7,041,870 describing K-M MOUSE technology, and U.S. Patent Application Publication No. US 2007/0061900, describing VELOCIMOUSE™ technology). Human variable regions from intact antibodies generated by such animals may be further modified, e.g., by combining with a different human constant region. [00198] Human antibodies can also be made by hybridoma-based methods. Human myeloma and mouse-human heteromyeloma cell lines for the production of human monoclonal antibodies have been described. (See, e.g., Kozbor J. Immunol., 133: 3001 (1984); Brodeur et al., Monoclonal Antibody Production Techniques and Applications, pp. 51-63 (Marcel Dekker, Inc., New York, 1987); and Boerner et al., J. Immunol., 147: 86 (1991).) Human antibodies generated via human B-cell hybridoma technology are also described in Li et al., Proc. Natl. Acad. Sci. USA, 103:3557- 3562 (2006). Additional methods include those described, for example, in U.S. Pat. No.7,189,826 (describing production of monoclonal human IgM antibodies from hybridoma cell lines) and Ni, Xiandai Mianyixue, 26(4):265-268 (2006) (describing human-human hybridomas). Human hybridoma technology (Trioma technology) is also described in Vollmers and Brandlein, Histology and Histopathology, 20(3):927-937 (2005) and Vollmers and Brandlein, Methods and Findings in Experimental and Clinical Pharmacology, 27(3):185-91 (2005). [00199] Human antibodies may also be generated by isolating Fv clone variable domain sequences selected from human-derived phage display libraries. Such variable domain sequences may then be combined with a desired human constant domain. Techniques for selecting human antibodies from antibody libraries are described below. [00200] Antibodies of the invention may be isolated by screening combinatorial libraries for antibodies with the desired activity or activities. For example, a variety of methods are known in the art for generating phage display libraries and screening such libraries for antibodies possessing the desired binding characteristics. Such methods are reviewed, e.g., in Hoogenboom et al., in Methods in Molecular Biology 178:1-37 (O'Brien et al., ed., Human Press, Totowa, N.J., 2001) and further described, e.g., in the McCafferty et al., Nature 348:552-554; Clackson et al., Nature 352: 624-628 (1991); Marks et al., J. Mol. Biol. 222: 581-597 (1992); Marks and Bradbury, in Methods in Molecular Biology 248:161-175 (Lo, ed., Human Press, Totowa, N.J., 2003); Sidhu et al., J. Mol. Biol. 338(2): 299-310 (2004); Lee et al., J. Mol. Biol. 340(5): 1073-1093 (2004); Fellouse, Proc. Natl. Acad. Sci. USA 101(34): 12467-12472 (2004); and Lee et al., J. Immunol. Methods 284(1-2): 119-132 (2004). [00201] In certain phage display methods, repertoires of VH and VL genes are separately cloned by polymerase chain reaction (PCR) and recombined randomly in phage libraries, which can then be screened for antigen-binding phage as described in Winter et al., Ann. Rev. Immunol., 12: 433- 455 (1994). Phage typically displays antibody fragments, either as scFv fragments or as Fab fragments. Libraries from immunized sources provide high-affinity antibodies to the immunogen without the requirement of constructing hybridomas. Alternatively, the naive repertoire can be cloned (e.g., from human) to provide a single source of antibodies to a wide range of non-self and also self-antigens without any immunization as described by Griffiths et al., EMBO J, 12: 725-734 (1993). Finally, naive libraries can also be made synthetically by cloning unrearranged V-gene segments from stem cells and using PCR primers containing random sequences to encode the highly variable CDR3 regions and to accomplish rearrangement in vitro, as described by Hoogenboom and Winter, J. Mol. Biol., 227: 381-388 (1992). Patent publications describing human antibody phage libraries include, for example, U.S. Pat. No. 5,750,373, and US Patent Publication Nos. 2005/0079574, 2005/0119455, 2005/0266000, 2007/0117126, 2007/0160598, 2007/0237764, 2007/0292936, and 2009/0002360. Antibodies or antibody fragments isolated from human antibody libraries are considered human antibodies or human antibody fragments herein. Variants [00202] In some embodiments, amino acid sequence variants of the antibodies provided herein are contemplated. For example, it may be desirable to improve the binding affinity and/or other biological properties of the antibody. Amino acid sequence variants of an antibody may be prepared by introducing appropriate modifications into the nucleotide sequence encoding the antibody, or by peptide synthesis. Such modifications include, for example, deletions from, and/or insertions into and/or substitutions of residues within the amino acid sequences of the antibody. Any combination of deletion, insertion, and substitution can be made to arrive at the final construct, provided that the final construct possesses the desired characteristics, e.g., antigen- binding. [00203] Substitution of one or more CDR residues or omission of one or more CDRs is also possible. Antibodies have been described in the scientific literature in which one or two CDRs can be dispensed with for binding. Padlan et al. (1995 FASEB J. 9:133-139) analyzed the contact regions between antibodies and their antigens, based on published crystal structures, and concluded that only about one fifth to one third of CDR residues actually contact the antigen. Padlan also found many antibodies in which one or two CDRs had no amino acids in contact with an antigen (see also, Vajdos et al.2002 J Mol Biol 320:415-428). [00204] CDR residues not contacting antigen can be identified based on previous studies (for example residues H60-H65 in CDRH2 are often not required), from regions of Kabat CDRs lying outside Chothia CDRs, by molecular modeling and/or empirically. If a CDR or residue(s) thereof is omitted, it is usually substituted with an amino acid occupying the corresponding position in another human antibody sequence or a consensus of such sequences. Positions for substitution within CDRs and amino acids to substitute can also be selected empirically. [00205] The fragments, whether attached to other sequences or not, can also include insertions, deletions, substitutions, or other selected modifications of particular regions or specific amino acids residues, provided the activity of the antibody or antibody fragment is not significantly altered or impaired compared to the non-modified antibody or antibody fragment. These modifications can provide for some additional property, such as to remove/add amino acids capable of disulfide bonding, to increase its bio-longevity, to alter its secretory characteristics, etc. In any case, the antibody or antibody fragment must possess a bioactive property, such as specific binding to its cognate antigen. Functional or active regions of the antibody or antibody fragment may be identified by mutagenesis of a specific region of the protein, followed by expression and testing of the expressed polypeptide. Such methods are readily apparent to a skilled practitioner in the art and can include site-specific mutagenesis of the nucleic acid encoding the antibody or antibody fragment. (Zoller, M.J. Curr. Opin. Biotechnol.3:348-354, 1992). [00206] As discussed herein there are numerous variants and derivatives of the antibodies, CDRs, VH, and VL proteins disclosed herein. Protein variants and derivatives are well understood to those of skill in the art and in can involve amino acid sequence modifications. For example, amino acid sequence modifications typically fall into one or more of three classes: substitutional, insertional or deletional variants. As used herein, “insertions” refer to a change in an amino acid or nucleotide sequence resulting in the addition of one or more amino acid or nucleotide residues, respectively, as compared to the parent, often the naturally occurring, molecule. Insertions include amino and/or carboxyl terminal fusions as well as intrasequence insertions of single or multiple amino acid residues. Insertions ordinarily will be smaller insertions than those of amino or carboxyl terminal fusions, for example, on the order of one to four residues. Immunogenic fusion protein derivatives, such as those described in the examples, are made by fusing a polypeptide sufficiently large to confer immunogenicity to the target sequence by cross-linking in vitro or by recombinant cell culture transformed with DNA encoding the fusion. Deletions are characterized by the removal of one or more amino acid residues from the protein sequence. Typically, no more than about from 2 to 6 residues are deleted at any one site within the protein molecule. These variants ordinarily are prepared by site specific mutagenesis of nucleotides in the DNA encoding the protein, thereby producing DNA encoding the variant, and thereafter expressing the DNA in recombinant cell culture. Techniques for making substitution mutations at predetermined sites in DNA having a known sequence are well known, for example M13 primer mutagenesis and PCR mutagenesis. Amino acid substitutions are typically of single residues but can occur at a number of different locations at once; insertions usually will be on the order of about from 1 to 10 amino acid residues; and deletions will range about from 1 to 30 residues. Deletions or insertions preferably are made in adjacent pairs, i.e., a deletion of 2 residues or insertion of 2 residues. Substitutions, deletions, insertions, or any combination thereof may be combined to arrive at a final construct. The mutations must not place the sequence out of reading frame and preferably will not create complementary regions that could produce secondary mRNA structure. Substitutional variants are those in which at least one residue has been removed and a different residue inserted in its place. Such substitutions generally are made in accordance with the following Table B and are referred to as conservative substitutions. Table A: Amino Acid Abbreviations
Figure imgf000070_0001
Figure imgf000071_0001
Table B: Amino Acid Substitutions - Exemplary Conservative Substitutions
Figure imgf000071_0002
[00207] Substantial changes in function or immunological identity are made by selecting substitutions that are less conservative than those in Table B, i.e., selecting residues that differ more significantly in their effect on maintaining (a) the structure of the polypeptide backbone in the area of the substitution, for example as a sheet or helical conformation, (b) the charge or hydrophobicity of the molecule at the target site or (c) the bulk of the side chain. Conservative amino acid substitutions include the ones in which the amino acid residue is replaced with an amino acid residue having similar structural or chemical properties. Families of amino acid residues having similar side chains have been defined in the art. These families include amino acids with basic side chains (e.g., lysine, arginine, histidine), acidic side chains (e.g., aspartic acid, glutamic acid), uncharged polar side chains (e.g., glycine, asparagine, glutamine, serine, threonine, tyrosine, cystine, tryptophan), nonpolar side chains (e.g., alanine, valine, leucine, isoleucine, proline, phenylalanine, methionine), beta-branched side chains (e.g., threonine, valine, isoleucine) and aromatic side chains (e.g., tyrosine, phenylalanine, tryptophan, histidine). [00208] The substitutions which in general are expected to produce the greatest changes in the protein properties will be those in which (a) a hydrophilic residue, e.g. seryl or threonyl, is substituted for (or by) a hydrophobic residue, e.g. leucyl, isoleucyl, phenylalanyl, valyl or alanyl; (b) a cysteine or proline is substituted for (or by) any other residue; (c) a residue having an electropositive side chain, e.g., lysyl, arginyl, or histidyl, is substituted for (or by) an electronegative residue, e.g., glutamyl or aspartyl; or (d) a residue having a bulky side chain, e.g., phenylalanine, is substituted for (or by) one not having a side chain, e.g., glycine, in this case, (e) by increasing the number of sites for sulfation and/or glycosylation. [00209] The replacement of one amino acid residue with another that is biologically and/or chemically similar is known to those skilled in the art as a conservative substitution. For example, a conservative substitution would be replacing one hydrophobic residue for another, or one polar residue for another. The substitutions include combinations such as, for example, Gly, Ala; Val, Ile, Leu; Asp, Glu; Asn, Gln; Ser, Thr; Lys, Arg; and Phe, Tyr. Such conservatively substituted variations of each explicitly disclosed sequence are included within the mosaic polypeptides provided herein. Substitution, Insertion, and Deletion Variants [00210] In some embodiments, antibody variants having one or more amino acid substitutions are provided. Sites of interest for substitutional mutagenesis include the HVRs and FRs. Conservative substitutions are defined herein. Amino acid substitutions may be introduced into an antibody of interest and the products screened for a desired activity, e.g., retained/improved antigen-binding, decreased immunogenicity, or improved antibody-dependent cell-mediated cytotoxicity (ADCC) and complement-dependent cytotoxicity (CDC). [00211] Accordingly, an antibody of the invention can comprise one or more conservative modifications of the CDRs, heavy chain variable region, or light variable regions described herein. A conservative modification or functional equivalent of a peptide, polypeptide, or protein disclosed in this invention refers to a polypeptide derivative of the peptide, polypeptide, or protein, e.g., a protein having one or more point mutations, insertions, deletions, truncations, a fusion protein, or a combination thereof. It substantially retains the activity to of the parent peptide, polypeptide, or protein (such as those disclosed in this invention). In general, a conservative modification or functional equivalent is at least 60% (e.g., any number between 60% and 100%, inclusive, e.g., 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, and 99%) identical to a parent. Accordingly, within the scope of this invention are heavy chain variable region or light variable regions having one or more point mutations, insertions, deletions, truncations, a fusion protein, or a combination thereof, as well as antibodies having the variant regions. [00212] As used herein, the percent homology between two amino acid sequences is equivalent to the percent identity between the two sequences. The percent identity between the two sequences is a function of the number of identical positions shared by the sequences (i.e., % homology=# of identical positions/total # of positions x 100), 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. The comparison of sequences and determination of percent identity between two sequences can be accomplished using a mathematical algorithm, as described in the non-limiting examples below. [00213] The percent identity between two amino acid sequences can be determined using the algorithm of E. Meyers and W. Miller (Comput. Appl. Biosci., 4:11-17 (1988)) which has been incorporated into the ALIGN program (version 2.0), using a PAM120 weight residue table, a gap length penalty of 12 and a gap penalty of 4. In addition, the percent identity between two amino acid sequences can be determined using the Needleman and Wunsch (J. Mol. Biol. 48:444-453 (1970)) algorithm, which has been incorporated into the GAP program in the GCG software package (available at gcg.com), using either a Blossum 62 matrix or a PAM250 matrix, and a gap weight of 16, 14, 12, 10, 8, 6, or 4 and a length weight of 1, 2, 3, 4, 5, or 6. [00214] Additionally or alternatively, the protein sequences of the present invention can further be used as a “query sequence” to perform a search against public databases to, for example, identify related sequences. Such searches can be performed using the XBLAST program (version 2.0) of Altschul, et al. (1990) J. Mol. Biol. 215:403-10. BLAST protein searches can be performed with the XBLAST program, score=50, wordlength=3 to obtain amino acid sequences homologous to the antibody molecules of the invention. To obtain gapped alignments for comparison purposes, Gapped BLAST can be utilized as described in Altschul et al., (1997) Nucleic Acids Res. 25(17):3389-3402. When utilizing BLAST and Gapped BLAST programs, the default parameters of the respective programs (e.g., XBLAST and NBLAST) can be used. (See ncbi.nlm.nih.gov). [00215] As used herein, the term “conservative modifications” refers to amino acid modifications that do not significantly affect or alter the binding characteristics of the antibody containing the amino acid sequence. Such conservative modifications include amino acid substitutions, additions, and deletions. Modifications can be introduced into an antibody of the invention by standard techniques known in the art, such as site-directed mutagenesis and PCR- mediated mutagenesis. Conservative amino acid substitutions are ones in which the amino acid residue is replaced with an amino acid residue having a similar side chain. Families of amino acid residues having similar side chains have been defined in the art. These families include: (i) amino acids with basic side chains (e.g., lysine, arginine, histidine), (ii) acidic side chains (e.g., aspartic acid, glutamic acid), (iii) uncharged polar side chains (e.g., glycine, asparagine, glutamine, serine, threonine, tyrosine, cysteine, tryptophan), (iv) nonpolar side chains (e.g., alanine, valine, leucine, isoleucine, proline, phenylalanine, methionine), (v) beta-branched side chains (e.g., threonine, valine, isoleucine), and (vi) aromatic side chains (e.g., tyrosine, phenylalanine, tryptophan, histidine). [00216] Non-conservative substitutions will entail exchanging a member of one of these classes for another class. [00217] An exemplary substitutional variant is an affinity matured antibody, which may be conveniently generated, e.g., using phage display-based affinity maturation techniques such as those described in, e.g., Hoogenboom et al., in Methods in Molecular Biology 178:1-37 (O'Brien et al., ed., Human Press, Totowa, N.J., (2001). Amino acid sequence insertions include amino- and/or carboxyl-terminal fusions ranging in length from one residue to polypeptides containing a hundred or more residues, as well as intrasequence insertions of single or multiple amino acid residues. Examples of terminal insertions include an antibody with an N-terminal methionyl residue. Other insertional variants of the antibody molecule include the fusion to the N- or C- terminus of the antibody to an enzyme (e.g., for ADEPT) or a polypeptide which increases the serum half-life of the antibody. Glycosylation Variants [00218] In some embodiments, an antibody provided herein is altered to increase or decrease the extent to which the antibody is glycosylated. Addition or deletion of glycosylation sites to an antibody may be conveniently accomplished by altering the amino acid sequence such that one or more glycosylation sites are created or removed. [00219] For example, an aglycoslated antibody can be made (i.e., the antibody lacks glycosylation). Glycosylation can be altered to, for example, increase the affinity of the antibody for antigen. Such carbohydrate modifications can be accomplished by, for example, altering one or more sites of glycosylation within the antibody sequence. For example, one or more amino acid substitutions can be made that result in elimination of one or more variable region framework glycosylation sites to thereby eliminate glycosylation at that site. Such aglycosylation may increase the affinity of the antibody for antigen. Such an approach is described in further detail in U.S. Patent Nos. 5,714,350 and 6,350,861 by Co et al. [00220] Glycosylation of the constant region on N297 may be prevented by mutating the N297 residue to another residue, e.g., N297A, and/or by mutating an adjacent amino acid, e.g., 298 to thereby reduce glycosylation on N297. [00221] Additionally or alternatively, an antibody can be made that has an altered type of glycosylation, such as a hypofucosylated antibody having reduced amounts of fucosyl residues or an antibody having increased bisecting GlcNac structures. Such altered glycosylation patterns have been demonstrated to increase the ADCC ability of antibodies. Such carbohydrate modifications can be accomplished by, for example, expressing the antibody in a host cell with altered glycosylation machinery. Cells with altered glycosylation machinery have been described in the art and can be used as host cells in which to express recombinant antibodies described herein to thereby produce an antibody with altered glycosylation. For example, EP 1,176,195 by Hanai et al. describes a cell line with a functionally disrupted FUT8 gene, which encodes a fucosyltransferase, such that antibodies expressed in such a cell line exhibit hypofucosylation. PCT Publication WO 03/035835 by Presta describes a variant Chinese Hamster Ovary cell line, Led 3 cells, with reduced ability to atach fucose to Asn(297)-lmked carbohydrates, also resulting in hypofucosylation of antibodies expressed in that host cell (see also Shields, R.L. et al. (2002) J . Biol. Chem. 277:26733-26740). PCT Publication WO 99/54342 by Umana et al. describes cell lines engineered to express glycoprotein-modifying glycosyltransferases (e.g., beta(l,4)-N- acetylglucosammyltransferase III (GnTIII)) such that antibodies expressed in the engineered cell lines exhibit increased bisecting GlcNac structures which result in increased ADCC activity of the antibodies (see also Umana et al. (1999) Nat. Biotech. 17: 176-180).
Fc Region Variants
[00222] The variable regions of the antibody described herein can be linked (e.g., covalently linked or fused) to an Fc, e.g., an IgGl, IgG2, IgG3 or IgG4 Fc, which may be of any allotype or isoallotype, e.g., for IgGl: Glm, Glml(a), Glm2(x), Glm3(f), Glml7(z); for IgG2: G2m, G2m23(n); for IgG3: G3m, G3m21(gl), G3m28(g5), G3ml l(b0), G3m5(bl), G3ml3(b3), G3ml4(b4), G3ml0(b5), G3ml5(s), G3ml6(t), G3m6(c3), G3m24(c5), G3m26(u), G3m27(v); and for K: Km, Kml, Km2, Km3 (see, e.g,, Jefferies et al. (2009) mAbs 1 : 1). In some embodiments, the antibodies variable regions described herein are linked to an Fc that binds to one or more activating Fc receptors (Fcyl, Fcylla, or Fcyllla), and thereby stimulate ADCC and may cause T cell depletion. In some embodiments, the antibody variable regions described herein are linked to an Fc that causes depletion.
[00223] In some embodiments, the antibody variable regions described herein may be linked to an Fc comprising one or more modifications, typically to alter one or more functional properties of the antibody, such as serum half-life, complement fixation, Fc receptor binding, and/or antigendependent cellular cytotoxicity. Furthermore, an antibody described herein may be chemically modified (e.g., one or more chemical moieties can be atached to the antibody) or be modified to alter its glycosylation, to alter one or more functional properties of the antibody. The numbering of residues in the Fc region is that of the EU index of Kabat.
[00224] The Fc region encompasses domains derived from the constant region of an immunoglobulin, preferably a human immunoglobulin, including a fragment, analog, variant, mutant, or derivative of the constant region. Suitable immunoglobulins include IgGl, IgG2, IgG3, IgG4, and other classes such as IgA, IgD, IgE, and IgM. The constant region of an immunoglobulin is defined as a naturally- occurring or synthetically-produced polypeptide homologous to the immunoglobulin C -terminal region, and can include a CHI domain, a hinge, a CH2 domain, a CH3 domain, or a CH4 domain, separately or in combination. In some embodiments, an antibody of this invention has an Fc region other than that of a wild-type IgAl . The antibody can have an Fc region from that of IgG (e.g., IgGl, IgG2, IgG3, and IgG4) or other classes such as IgA2, IgD, IgE, and IgM. The Fc can be a mutant form of IgAl.
[00225] The constant region of an immunoglobulin is responsible for many important antibody functions, including Fc receptor (FcR) binding and complement fixation. There are five major classes of heavy chain constant region, classified as IgA, IgG, IgD, IgE, IgM, each with characteristic effector functions designated by isotype. For example, IgG is separated into four subclasses known as IgGl, IgG2, IgG3, and IgG4.
[00226] Ig molecules interact with multiple classes of cellular receptors. For example, IgG molecules interact with three classes of Fey receptors (FcyR) specific for the IgG class of antibody, namely FcyRI, FcyRII, and FcyRUL. The important sequences for the binding of IgG to the FcyR receptors have been reported to be located in the CH2 and CH3 domains. The serum half-life of an antibody is influenced by the ability of that antibody to bind to an FcR.
[00227] In some embodiments, the Fc region is a variant Fc region, e.g., an Fc sequence that has been modified (e.g., by amino acid substitution, deletion and/or insertion) relative to a parent Fc sequence (e.g., an unmodified Fc polypeptide that is subsequently modified to generate a variant), to provide desirable structural features and/or biological activity. For example, one may make modifications in the Fc region in order to generate an Fc variant that (a) has increased or decreased ADCC, (b) increased or decreased CDC, (c) has increased or decreased affinity for Clq and/or (d) has increased or decreased affinity for an Fc receptor relative to the parent Fc. Such Fc region variants will generally comprise at least, one ammo acid modification in the Fc region. Combining amino acid modifications is thought to be particularly desirable. For example, the variant Fc region may include two, three, four, five, etc., substitutions therein, e.g., of the specific Fc region positions identified herein.
[00228] A variant Fc region may also comprise a sequence alteration wherein amino acids involved in disulfide bond formation are removed or replaced with other amino acids. Such removal may avoid reaction with other cysteine-containing proteins present in the host cell used to produce the antibodies described herein. Even when cy steine residues are removed, single chain Fc domains can still form a dimeric Fc domain that is held together non-covalently. In other embodiments, the Fc region may be modified to make it more compatible with a selected host cell. For example, one may remove the PA sequence near the N-terminus of a typical native Fc region, which may be recognized by a digestive enzyme in E. coli, such as proline iminopeptidase. In other embodiments, one or more glycosylation sites within the Fc domain may be removed. Residues that are typically glycosylated (e.g., asparagine) may confer cytolytic response. Such residues may be deleted or substituted with unglycosylated residues (e.g., alanine). In other embodiments, sites involved in interaction with complement, such as the Clq binding site, may be removed from the Fc region. For example, one may delete or substitute the EKK sequence of human IgGl. In some embodiments, sites that affect binding to Fc receptors may be removed, preferably sites other than salvage receptor binding sites. In other embodiments, an Fc region may be modified to remove an ADCC site. ADCC sites are known in the art; see, for example, Molec. Immunol. 29 (5): 633-9 (1992) with regard to ADCC sites in IgGl. Specific examples of variant Fc domains are disclosed, for example, in WO 97/34631 and WO 96/32478. [00229] In one embodiment, the hinge region of Fc is modified such that the number of cysteine residues in the hinge region is altered, e.g., increased or decreased. This approach is described further in U.S. Patent No. 5,677,425 by Bodmer et al. The number of cysteine residues in the hinge region of Fc is altered to, for example, facilitate assembly of the light and heavy chains or to increase or decrease the stability of the antibody. In one embodiment, the Fc hinge region of an antibody is mutated to decrease the biological half-life of the antibody. More specifically, one or more amino acid mutations are introduced into the CH2-CH3 domain interface region of the Fc- hinge fragment such that the antibody has impaired Staphylococcal protein A (SpA) binding relative to native Fc-hinge domain SpA binding. This approach is described in further detail in U.S. Patent No.6,165,745 by Ward et al. [00230] In yet other embodiments, the Fc region is altered by replacing at least one amino acid residue with a different amino acid residue to alter the effector function(s) of the antibody. For example, one or more amino acids selected from amino acid residues 234, 235, 236, 237, 297, 318, 320, and 322 can be replaced with a different amino acid residue such that the antibody has an altered affinity for an effector ligand but retains the antigen-binding ability of the parent antibody. The effector ligand to which affinity is altered can be, for example, an Fc receptor or the CI component of complement. This approach is described in further detail in U.S. Patent Nos. 5,624,821 and 5,648,260, both by Winter et al. [00231] In another example, one or more amino acids selected from amino acid residues 329, 331, and 322 can be replaced with a different ammo acid residue such that the antibody has altered Clq binding and/or reduced or abolished CDC. This approach is described in further detail in U.S. Patent Nos. 6,194,551 by Idusogie et al.
[00232] In another example, one or more amino acid residues within amino acid positions 231 and 239 are altered to thereby alter the ability of the antibody to fix complement. This approach is described further in PCT Publication WO 94/29351 by Bodmer et al.
[00233] In yet another example, the Fc region may be modified to increase ADCC and/or to increase the affinity for an Fey receptor by modifying one or more amino acids at the following positions: 234, 235, 236, 238, 239, 240, 241 , 243, 244, 245, 247, 248, 249, 252, 254, 255, 256, 258, 262, 263, 264, 265, 267, 268, 269, 270, 272, 276, 278, 280, 283, 285, 286, 289, 290, 292, 293,
294, 295, 296, 298, 299, 301, 303, 305, 307, 309, 312, 313, 315, 320, 322, 324, 325, 326, 327, 329,
330, 331, 332, 333, 334, 335, 337, 338, 340, 360, 373, 376, 378, 382, 388, 389, 398, 414, 416, 419,
430, 433, 434, 435, 436, 437, 438 or 439. Exemplary substitutions include 236A, 239D, 239E,
268D, 267E, 268E, 268F, 324T, 332D, and 332E. Exemplary variants include 239D/332E, 236A/332E, 236A/239D/332E, 268F/324T, 267E/268F, 267E/324T, and 267E/268F7324T. Other modifications for enhancing FcyR and complement interactions include but are not limited to substitutions 298A, 333A, 334A, 326A, 2471, 339D, 339Q, 280H, 290S, 2981'1. 298V, 243L, 292P, 300L, 396L, 3051, and 396L. These and other modifications are reviewed in Strohl, 2009, Current Opinion in Biotechnology 20:685-691.
[00234] Fc modifications that increase binding to an Fey receptor include amino acid modifications at any one or more of amino acid positions 238, 239, 248, 249, 252, 254, 255, 256, 258, 265, 267, 268, 269, 270, 272, 279, 280, 283, 285, 298, 289, 290, 292, 293, 294, 295, 296, 298, 301, 303, 305, 307, 312, 315, 324, 327, 329, 330, 335, 337, 3338, 340, 360, 373, 376, 379, 382, 388, 389, 398, 414, 416, 419, 430, 434, 435, 437, 438 or 439 of the Fc region, wherein the numbering of the residues in the Fc region is that of the EU index as in Kabat (WO00/42072).
[00235] Other Fc modifications that can be made to Fes are those for reducing or ablating binding to FcyR and/or complement proteins, thereby reducing or ablating Fc-mediated effector functions such as ADCC, antibody-dependent cellular phagocytosis (ADCP), and CDC. Exemplary modifications include but are not limited to substitutions, insertions, and deletions at positions 234, 235, 236, 237, 267, 269, 325, and 328, wherein numbering is according to the EU index. Exemplary substitutions include but are not limited to 234G, 235G, 236R, 237K, 267R, 269R, 325L, and 328R, wherein numbering is according to the EU index. An Fc variant may comprise 236R/328R. Other modifications for reducing FcyR and complement interactions include substitutions 297A, 234 A, 235A, 237A, 318A, 228P, 236E, 268Q, 309L, 330S, 331 S, 220S, 226S, 229S, 238S, 233P, and 234V, as well as removal of the glycosylation at position 297 by mutational or enzymatic means or by production in organisms such as bacteria that do not glycosylate proteins. These and other modifications are reviewed in Strohl, 2009, Current Opinion in Biotechnology 20:685-691.
[00236] Optionally, the Fc region may comprise a non-naturally occurring amino acid residue at additional and/or alternative positions known to one skilled in the art (see, e.g., ELS. Pat. Nos. 5,624,821; 6,277,375; 6,737,056; 6,194,551; 7,317,091; 8,101,720; WO00/42072; WOOl/58957; W002/06919; WO04/016750; W004/029207; WO04/035752; WO04/074455; WO04/099249; W004/063351; W005/070963; W005/040217, WO05/092925 and W006/020114).
[00237] Fc variants that enhance affinity for an inhibitory receptor FcyRIIb may also be used. Such variants may provide an Fc fusion protein with immune-modulatory activities related to FcyRIIb cells, including, for example, B cells and monocytes. In one embodiment, the Fc variants provide selectively enhanced affinity to FcyRIIb relative to one or more activating receptors. Modifications for altering binding; to FcyRIIb include one or more modifications at a position selected from the group consisting of 234, 235, 236, 237, 239, 266, 267, 268, 325, 326, 327, 328, and 332, according to the EU index. Exemplary substitutions for enhancing FcyRIIb affinity include but are not limited to 234D, 234E, 234F, 234W, 235D, 235F, 235R, 235Y, 236D, 236N, 237D, 237N, 239D, 239E, 266M, 267D, 267E, 268D, 268E, 327D, 327E, 328F, 328W, 328Y, and 332E. Exemplary substitutions include 235Y, 236D, 239D, 266M, 267E, 268D, 268E, 328F, 328W, and 328Y. Other Fc variants for enhancing binding to FcyRIIb include 235Y/267E, 236D/267E, 239D/268D, 239D/267E, 267E/268D, 267E/268E, and 267E/328F.
[00238] The affinities and binding properties of an Fc region for its ligand may be determined by a variety of in vitro assay methods (biochemical or immunological based assays) known in the art, including but not limited to equilibrium methods (e.g., ELISA, or radioimmunoassay), or kinetics (e.g., BIACORE™ analysis), and other methods such as indirect binding assays, competitive inhibition assays, fluorescence resonance energy transfer (FRET), gel electrophoresis and chromatography (e.g., gel filtration). These and other methods may utilize a label on one or more of the components being examined and/or employ a variety of detection methods, including but not limited to chromogenic, fluorescent, luminescent, or isotopic labels. A detailed description of binding affinities and kinetics can be found in Paul, W. E., ed., Fundamental Immunology, 4th Ed., Lippincott-Raven, Philadelphia (1999), which focuses on antibody-immunogen interactions. [00239] In some embodiments, the antibody is modified to increase its biological half-life. Various approaches are possible. For example, this may be done by increasing the binding affinity of the Fc region for FcRn. For example, one or more of the following residues can be mutated: 252, 254, 256, 433, 435, 436, as described in U.S. Pat. No. 6,277,375. Specific exemplary substitutions include one or more of the following: T252L, T254S, and/or T256F. Alternatively, to increase the biological half-life, the antibody can be altered within the CH1 or CL region to contain a salvage receptor binding epitope taken from two loops of a CH2 domain of an Fc region of an IgG, as described in U.S. Patent Nos. 5,869,046 and 6,121,022 by Presta et al. Other exemplary variants that increase binding to FcRn and/or improve pharmacokinetic properties include substitutions at positions 259, 308, 428, and 434, including for example 259I, 308F, 428L, 428M, 434S, 434H, 434F, 434Y, and 434M. Other variants that increase Fc binding to FcRn include: 250E, 250Q, 428L, 428F, 250Q/428L (Hinton et al„ 2004, J. Biol. Chem. 279(8): 6213- 6216, Hinton et al.2006 Journal of Immunology 176:346-356), 256A, 272A, 286A, 305A, 307A, 307Q, 311A, 312A, 376A, 378Q, 380A, 382A, 434A (Shields et al, Journal of Biological Chemistry, 2001, 276(9):6591-6604), 252F, 252T, 252Y, 252W, 254T, 256S, 256R, 256Q, 256E, 256D, 256T, 309P, 311S, 433R, 433S, 433I, 433P, 433Q, 434H, 434F, 434Y, 252Y/254T/256E, 433K/434F/436H, 308T/309P/311S (Dall Acqua et al. Journal of Immunology, 2002, 169:5171- 5180, Dall'Acqua et al., 2006, Journal of Biological Chemistry 281:23514-23524). Other modifications for modulating FcRn binding are described in Yeung et al., 2010, J Immunol, 182:7663-7671. In some embodiments, hybrid IgG isotypes with particular biological characteristics may be used. For example, an IgGl/IgG3 hybrid variant may be constructed by substituting IgG 1 positions in the CH2 and/or CH3 region with the amino acids from IgG3 at positions where the two isotypes differ. Thus, a hybrid variant IgG antibody may be constructed that comprises one or more substitutions, e.g., 274Q, 276K, 300F, 339T, 356E, 358M, 384S, 392N, 397M, 422I, 435R, and 436F. In other embodiments described herein, an IgGl/IgG2 hybrid variant may be constructed by substituting IgG2 positions in the CH2 and/or CH3 region with amino acids from IgGl at positions where the two isotypes differ. Thus, a hybrid variant IgG antibody may be constructed chat comprises one or more substitutions, e.g., one or more of the following ammo acid substitutions: 233E, 234L, 235L, 236G (referring to an insertion of a glycine at position 236), and 321 h.
[00240] Moreover, the binding sites on human IgGl for FcyRl, FcyRII, FcyRIII, and FcRn have been mapped, and variants with improved binding have been described (see Shields, R.L. et al. (2001) J. Bioi. Chem. 276:6591-6604). Specific mutations at positions 256, 290, 298, 333, 334, and 339 were shown to improve binding to FcyRIII. Additionally, the following combination mutants were shown to improve FcyRIII binding: T256A/S298A, S298A/E333A, S298A,fK224A, and S298A/E333A/K334A, which has been shown to exhibit enhanced FcyRIIIa binding and ADCC activity (Shields et al., 2001). Other IgGl variants with strongly enhanced binding to FcyRIIIa have been identified, including variants with S239D/I332E and S239D/I332E/A330L mutations which showed the greatest increase in affinity for FcyRIIIa, a decrease in FcyRIIb binding, and strong cytotoxic activity in cynomolgus monkeys (Lazar et al., 2006). Introduction of the triple mutations into antibodies such as alemtuzumab (CD52- specific), trastuzumab (HER2/neu- specific), rituximab (CD20- specific), and cetuximab (EGFR- specific) translated into greatly enhanced ADCC activity' in vitro, and the S239D/I332E variant showed an enhanced capacity to deplete B cells in monkeys (Lazar et al., 2006). In addition, IgGl mutants containing L235V, F243L, R292P, Y300L, and P396L mutations which exhibited enhanced binding to FcyRIIIa and concomitantly enhanced ADCC activity' in transgenic mice expressing human FcyRIIIa in models of B cell malignancies and breast cancer have been identified (Stavenhagen et al., 2007; Nordstrom et al., 2011). Other Fc mutants that, may be used include S298A/E333A/L334A, S239D/1332E, S239D/I332E/A330L, L235V/F243L/R292P/Y300L/ P396L, and M428L/N434S.
[00241] In some embodiments, an Fc is chosen that has reduced binding to FcyRs. An exemplary Fc, e.g., IgGl Fc, with reduced FcyR binding, comprises the following three amino acid substitutions: L234A, L235E, and G237A.
[00242] In some embodiments, an Fc is chosen that has reduced complement fixation. An exemplary Fc, e.g., IgGl Fc, with reduced complement fixation, has the following two ammo acid substitutions: A330S and P331S.
[00243] In some embodiments, an Fc is chosen that has essentially no effector function, i.e., it has reduced binding to FcyRs and reduced complement fixation. An exemplary Fc, e.g., IgGl Fc, that is effectorless, comprises the following five mutations: L234A, L235E, G237A, A330S, and P331S. [00244] When using an IgG4 constant domain, it is usually preferable to include the substitution S228P, which mimics the hinge sequence in IgGl and thereby stabilizes IgG4 molecules. Multivalent Antibodies [00245] In one embodiment, the antibodies of the invention may be monovalent or multivalent (e.g., bivalent, trivalent, etc.). As used herein, the term “valency” refers to the number of potential target binding sites associated with an antibody. Each target binding site specifically binds one target molecule or specific position or locus on a target molecule. When an antibody is monovalent, each binding site of the molecule will specifically bind to a single antigen position or epitope. When an antibody comprises more than one target binding site (multivalent), each target binding site may specifically bind to the same or different molecules (e.g., may bind to different ligands or different antigens, or different epitopes or positions on the same antigen). See, for example, U.S.P.N. 2009/0130105. [00246] In one embodiment, the antibodies are bispecific antibodies in which the two chains have different specificities, as described in Millstein et al., 1983, Nature, 305:537-539. Other embodiments include antibodies with additional specificities, such as trispecific antibodies. Other more sophisticated compatible multispecific constructs and methods of their fabrication are set forth in U.S.P.N. 2009/0155255, as well as WO 94/04690; Suresh et al., 1986, Methods in Enzymology, 121:210; and WO96/27011. [00247] As stated above, multivalent antibodies may immunospecifically bind to different epitopes of the desired target molecule or may immunospecifically bind to both the target molecule as well as a heterologous epitope, such as a heterologous polypeptide or solid support material. In some embodiments, the multivalent antibodies may include bispecific antibodies or trispecific antibodies. Bispecific antibodies also include cross-linked or “heteroconjugate” antibodies. For example, one of the antibodies in the heteroconjugate can be coupled to avidin, the other to biotin. Such antibodies have, for example, been proposed to target immune system cells to unwanted cells (U.S. Pat. No.4,676,980), and for treatment of HIV infection (WO 91/00360, WO 92/200373, and EP 03089). Heteroconjugate antibodies may be made using any convenient cross-linking methods. Suitable cross-linking agents are well known in the art and are disclosed in U.S. Pat. No.4,676,980, along with a number of cross-linking techniques. [00248] In some embodiments, antibody variable domains with the desired binding specificities (antibody-antigen combining sites) are fused to immunoglobulin constant domain sequences, such as an immunoglobulin heavy chain constant domain comprising at least part of the hinge, CH2, and/or CH3 regions, using methods well known to those of ordinary skill in the art. Antibody Derivatives [00249] An antibody provided herein may be further modified to contain additional nonproteinaceous moieties that are known in the art and readily available. The moieties suitable for derivatization of the antibody include but are not limited to water-soluble polymers. [00250] Non-limiting examples of water-soluble polymers include, but are not limited to, PEG, copolymers of ethylene glycol/propylene glycol, carboxymethylcellulose, dextran, polyvinyl alcohol, polyvinyl pyrrolidone, poly-1,3-dioxolane, poly-1,3,6-trioxane, ethylene/maleic anhydride copolymer, polyaminoacids (either homopolymers or random copolymers), and dextran or poly(n-vinyl pyrrolidone)polyethylene glycol, propropylene glycol homopolymers, polypropylene oxide/ethylene oxide co-polymers, polyoxyethylated polyols (e.g., glycerol), polyvinyl alcohol, and mixtures thereof. Polyethylene glycol propionaldehyde may have advantages in manufacturing due to its stability in water. The polymer may be of any molecular weight and may be branched or unbranched. The number of polymers attached to the antibody may vary, and if more than one polymer is attached, they can be the same or different molecules. In general, the number and/or type of polymers used for derivatization can be determined based on considerations including, but not limited to, the particular properties or functions of the antibody to be improved, whether the antibody derivative will be used in a therapy under defined conditions, etc. [00251] In another embodiment, conjugates of an antibody and nonproteinaceous moiety that may be selectively heated by exposure to radiation are provided. In one embodiment, the nonproteinaceous moiety is a carbon nanotube (Kam et al., Proc. Natl. Acad. Sci. USA 102: 11600- 11605 (2005)). The radiation may be of any wavelength and includes, but is not limited to, wavelengths that do not harm ordinary cells, but which heat the nonproteinaceous moiety to a temperature at which cells proximal to the antibody-nonproteinaceous moiety are killed. [00252] Another modification of the antibodies described herein is pegylation. An antibody can be pegylated to, for example, increase the biological (e.g., serum) half-life of the antibody. To pegylate an antibody, the antibody, or fragment thereof, typically is reacted with PEG, such as a reactive ester or aldehyde derivative of PEG, under conditions in which one or more PEG groups become attached to the antibody or antibody fragment. Preferably, the pegylation is carried out via an acylation reaction or an alkylation reaction with a reactive PEG molecule (or an analogous reactive water-soluble polymer). As used herein, the term “polyethylene glycol” is intended to encompass any of the forms of PEG that have been used to derivatize other proteins, such as mono (CI -CIO) alkoxy- or aryloxy-polyethylene glycol or polyethylene glycol-maleimide. In some embodiments, the antibody to be pegylated is an aglycosylated antibody. Methods for pegylating proteins are known in the art and can be applied to the antibodies described herein. See, for example, EP0154316 by Nishimura et al. and EP0401384 by Ishikawa et al. [00253] The present invention also encompasses a human monoclonal antibody described herein conjugated to a therapeutic agent, a polymer, a detectable label, or enzyme. In one embodiment, the therapeutic agent is a cytotoxic agent. In one embodiment, the polymer is PEG. 1.3. Nucleic Acids, Expression Cassettes, and Vectors [00254] The present invention provides isolated nucleic acid segments that encode the polypeptides, peptide fragments, and coupled proteins of the invention. The nucleic acid segments of the invention also include segments that encode for the same amino acids due to the degeneracy of the genetic code. For example, the amino acid threonine is encoded by ACU, ACC, ACA, and ACG and is therefore degenerate. It is intended that the invention includes all variations of the polynucleotide segments that encode for the same amino acids. Such mutations are known in the art (Watson et al., Molecular Biology of the Gene, Benjamin Cummings 1987). Mutations also include alteration of a nucleic acid segment to encode for conservative amino acid changes, for example, the substitution of leucine for isoleucine and so forth. Such mutations are also known in the art. Thus, the genes and nucleotide sequences of the invention include both the naturally occurring sequences as well as mutant forms. [00255] The nucleic acid segments of the invention may be contained within a vector. A vector may include, but is not limited to, any plasmid, phagemid, F-factor, virus, cosmid, or phage in a double- or single-stranded linear or circular form which may or may not be self transmissible or mobilizable. The vector can also transform a prokaryotic or eukaryotic host either by integration into the cellular genome or exist extra-chromosomally (e.g., autonomous replicating plasmid with an origin of replication). [00256] The nucleic acid segment in the vector can be under the control of, and operably linked to, an appropriate promoter or other regulatory elements for transcription in vitro or in a host cell, such as a eukaryotic cell, or a microbe, e.g., bacteria. The vector may be a shuttle vector that functions in multiple hosts. The vector may also be a cloning vector that typically contains one or a small number of restriction endonuclease recognition sites at which foreign DNA sequences can be inserted in a determinable fashion. Such insertion can occur without loss of essential biological function of the cloning vector. A cloning vector may also contain a marker gene that is suitable for use in the identification and selection of cells transformed with the cloning vector. Examples of marker genes are tetracycline resistance or ampicillin resistance. Many cloning vectors are commercially available (Stratagene, New England Biolabs, Clonetech). [00257] The nucleic acid segments of the invention may also be inserted into an expression vector. Typically, an expression vector contains prokaryotic DNA elements coding for a bacterial replication origin and an antibiotic resistance gene to provide for the amplification and selection of the expression vector in a bacterial host; regulatory elements that control initiation of transcription such as a promoter; and DNA elements that control the processing of transcripts such as introns, or a transcription termination/polyadenylation sequence. [00258] Methods to introduce nucleic acid segment into a vector are available in the art (Sambrook et al., Molecular Cloning: A Laboratory Manual, 3rd edition, Cold Spring Harbor Press, Cold Spring Harbor, N.Y. (2001)). Briefly, a vector into which a nucleic acid segment is to be inserted is treated with one or more restriction enzymes (restriction endonuclease) to produce a linearized vector having a blunt end, a Āstickyā end with a 5’ or a 3’ overhang, or any combination of the above. The vector may also be treated with a restriction enzyme and subsequently treated with another modifying enzyme, such as a polymerase, an exonuclease, a phosphatase or a kinase, to create a linearized vector that has characteristics useful for ligation of a nucleic acid segment into the vector. The nucleic acid segment that is to be inserted into the vector is treated with one or more restriction enzymes to create a linearized segment having a blunt end, a Āstickyā end with a 5’ or a 3’ overhang, or any combination of the above. The nucleic acid segment may also be treated with a restriction enzyme and subsequently treated with another DNA modifying enzyme. Such DNA modifying enzymes include, but are not limited to, polymerase, exonuclease, phosphatase or a kinase, to create a nucleic acid segment that has characteristics useful for ligation of a nucleic acid segment into the vector. [00259] The treated vector and nucleic acid segment are then ligated together to form a construct containing a nucleic acid segment according to methods available in the art (Sambrook et al., Molecular Cloning: A Laboratory Manual, 3rd edition, Cold Spring Harbor Press, Cold Spring Harbor, N.Y. (2001)). Briefly, the treated nucleic acid fragment, and the treated vector are combined in the presence of a suitable buffer and ligase. The mixture is then incubated under appropriate conditions to allow the ligase to ligate the nucleic acid fragment into the vector. [00260] The invention also provides an expression cassette that contains a nucleic acid sequence capable of directing expression of a particular nucleic acid segment of the invention, either in vitro or in a host cell. Also, a nucleic acid segment of the invention may be inserted into the expression cassette such that an anti-sense message is produced. The expression cassette is an isolatable unit such that the expression cassette may be in linear form and functional for in vitro transcription and translation assays. The materials and procedures to conduct these assays are commercially available from Promega Corp. (Madison, Wis.). For example, an in vitro transcript may be produced by placing a nucleic acid sequence under the control of a T7 promoter and then using T7 RNA polymerase to produce an in vitro transcript. This transcript may then be translated in vitro through use of a rabbit reticulocyte lysate. Alternatively, the expression cassette can be incorporated into a vector allowing for replication and amplification of the expression cassette within a host cell or also in vitro transcription and translation of a nucleic acid segment. [00261] Such an expression cassette may contain one or a plurality of restriction sites allowing for placement of the nucleic acid segment under the regulation of a regulatory sequence. The expression cassette can also contain a termination signal operably linked to the nucleic acid segment as well as regulatory sequences required for proper translation of the nucleic acid segment. The expression cassette containing the nucleic acid segment may be chimeric, meaning that at least one of its components is heterologous with respect to at least one of its other components. The expression cassette may also be one that is naturally occurring but has been obtained in a recombinant form useful for heterologous expression. Expression of the nucleic acid segment in the expression cassette may be under the control of a constitutive promoter or an inducible promoter, which initiates transcription only when the host cell is exposed to some particular external stimulus. [00262] The expression cassette may include in the 5’-3’ direction of transcription, a transcriptional and translational initiation region, a nucleic acid segment and a transcriptional and translational termination region functional in vivo and/or in vitro. The termination region may be native with the transcriptional initiation region, may be native with the nucleic acid segment, or may be derived from another source. [00263] The regulatory sequence can be a polynucleotide sequence located upstream (5' non- coding sequences), within, or downstream (3’ non-coding sequences) of a coding sequence, and which influences the transcription, RNA processing or stability, or translation of the associated coding sequence. Regulatory sequences can include, but are not limited to, enhancers, promoters, repressor binding sites, translation leader sequences, introns, and polyadenylation signal sequences. They may include natural and synthetic sequences as well as sequences which may be a combination of synthetic and natural sequences. While regulatory sequences are not limited to promoters, some useful regulatory sequences include constitutive promoters, inducible promoters, regulated promoters, tissue-specific promoters, viral promoters, and synthetic promoters. [00264] A promoter is a nucleotide sequence that controls the expression of the coding sequence by providing the recognition for RNA polymerase and other factors required for proper transcription. A promoter includes a minimal promoter, consisting only of all basal elements needed for transcription initiation, such as a TATA-box and/or initiator that is a short DNA sequence comprised of a TATA-box and other sequences that serve to specify the site of transcription initiation, to which regulatory elements are added for control of expression. A promoter may be derived entirely from a native gene, or be composed of different elements derived from different promoters found in nature, or even be comprised of synthetic DNA segments. A promoter may contain DNA sequences that are involved in the binding of protein factors that control the effectiveness of transcription initiation in response to physiological or developmental conditions. [00265] The invention also provides a construct containing a vector and an expression cassette. The vector may be selected from, but not limited to, any vector previously described. Into this vector may be inserted an expression cassette through methods known in the art and previously described (Sambrook et al., Molecular Cloning: A Laboratory Manual, 3rd edition, Cold Spring Harbor Press, Cold Spring Harbor, N.Y. (2001)). In one embodiment, the regulatory sequences of the expression cassette may be derived from a source other than the vector into which the expression cassette is inserted. In another embodiment, a construct containing a vector and an expression cassette is formed upon insertion of a nucleic acid segment of the invention into a vector that itself contains regulatory sequences. Thus, an expression cassette is formed upon insertion of the nucleic acid segment into the vector. Vectors containing regulatory sequences are available commercially, and methods for their use are known in the art (Clonetech, Promega, Stratagene). [00266] Provided herein are vectors comprising the polynucleotides or nucleic acid molecules disclosed herein. [00267] The host cell may be co-transfected with two vectors provided herein, the first vector encoding a heavy chain derived polypeptide and the second vector encoding a light chain derived polypeptide. The two vectors may contain identical selectable markers which enable equal expression of heavy and light chain polypeptides. Alternatively, a single vector may be used which encodes, and is capable of expressing, both heavy and light chain polypeptides. In such situations, the light chain should be placed before the heavy chain to avoid an excess of toxic free heavy chain (Proudfoot, Nature, 1986, 322:52; and Kohler, Proc. Natl. Acad. Sci. USA , 1980, 77:2197-9). [00268] In some embodiments, vectors include, without limitation, plasmids, phagemids, cosmids, transposons, artificial chromosomes such as yeast artificial chromosome (YAC), bacterial artificial chromosome (BAC), or P1-derived artificial chromosome (PAC), bacteriophages such as lambda phage or M13 phage, and animal viruses. In some embodiments, the coding sequences of the antibody or antigen-binding fragment thereof disclosed herein can be ligated into such vectors for expression in mammalian cells. [00269] In some embodiments, non-viral vectors are used to deliver one or more polynucleotides contemplated herein. In some embodiments, the recombinant vector comprising a polynucleotide encoding the antibody or antigen-binding fragment thereof described herein is a plasmid. Numerous suitable plasmid expression vectors are known to those of skill in the art, and many are commercially available. The following vectors are provided by way of example; for eukaryotic host cells: pXT1, pSG5 (Stratagene), pSVK3, pBPV, pMSG, and pSVLSV40 (Pharmacia). However, any other plasmid vector may be used so long as it is compatible with the host cell. [00270] In some embodiments, viral vectors are used to deliver one or more polynucleotides contemplated herein. Suitable viral vectors include, but are not limited to, viral vectors based on adenovirus (see, e.g., Li et al., Invest Opthalmol Vis Sci 35:25432549, 1994; Borras et al., Gene Ther 6:515524, 1999; Li and Davidson, PNAS 92:77007704, 1995; Sakamoto et al., H Gene Ther 5:10881097, 1999; WO 94/12649, WO 93/03769; WO 93/19191 ; WO 94/28938; WO 95/11984 and WO 95/00655); adeno-associated virus (see, e.g., U.S. Patent No. 7,078,387; Ali et al., Hum Gene Ther 9:81 86, 1998, Flannery et al„ PNAS 94:6916 6921 , 1997; Bennett et al., Invest Opthalmol Vis Sci 38:28572863, 1997; Jomary et al., Gene Ther 4:683690, 1997, Rolling et al., Hum Gene Ther 10:641648, 1999; Ali et al., Hum Mol Genet 5:591594, 1996; Srivastava in WO 93/09239, Samulski et al., J. Vir. (1989) 63:3822-3828; Mendelson et al„ Virol. (1988) 166:154- 165; and Flotte et al., PNAS (1993) 90:10613-10617); alphaviruses; arenaviruses; baculovirus; herpes simplex virus; human immunodeficiency virus (see, e.g., Miyoshi et al., PNAS 94:10319 23, 1997; Takahashi et al., J Virol 73:7812 7816, 1999); poliovirus; poxvirus; retrovirus (e.g., Murine Leukemia Virus, spleen necrosis virus, and vectors derived from retroviruses such as Rous Sarcoma Virus, Harvey Sarcoma Virus, avian leukosis virus, a lentivirus, human immunodeficiency virus, myeloproliferative sarcoma virus, and mammary tumor virus); SV40; vaccinia virus; and the like. Examples of vectors are pClneo vectors (Promega) for expression in mammalian cells; pLenti4/V5-DEST™, pLenti6/V5-DEST™, and pLenti6.2/V5-GW/lacZ (Invitrogen) for lentivirus-mediated gene transfer and expression in mammalian cells. [00271] In some embodiments, the vector is a non-integrating vector, including but not limited to, an episomal vector or a vector that is maintained extrachromosomally. As used herein, the term “episomal” refers to a vector that is able to replicate without integration into host’s chromosomal DNA and without gradual loss from a dividing host cell also meaning that said vector replicates extrachromosomally or episomally. The vector is engineered to harbor the sequence coding for the origin of DNA replication or “ori” from a lymphotrophic herpes virus or a gamma herpesvirus, an adenovirus, SV40, a bovine papilloma virus, or a yeast, specifically a replication origin of a lymphotrophic herpes virus or a gamma herpesvirus corresponding to oriP of EBV. In some embodiments, the lymphotrophic herpes virus may be Epstein Barr virus (EBV), Kaposi’s sarcoma herpes virus (KSHV), Herpes virus saimiri (HS), or Marek’s disease virus (MDV). Epstein Barr virus (EBV) and Kaposi’s sarcoma herpes virus (KSHV) are also examples of a gamma herpesvirus. A viral vector delivered by such viruses or viral particles may be referred to by the type of virus to deliver the viral vector (e.g., a lentiviral vector is a viral vector that is to be delivered by a lentivirus). A viral vector can contain viral elements (e.g., nucleotide sequences) necessary for packaging of the viral vector into the virus or viral particle, replicating the virus, or other desired viral activities. A virus containing a viral vector may be replication competent, replication deficient or replication defective. [00272] In some embodiments, the vector is an integrating vector. In some embodiments, a polynucleotide is introduced into a target or host cell using a transposon vector system. In some embodiments, the transposon vector system comprises a vector comprising transposable elements and a polynucleotide contemplated herein; and a transposase. In some embodiments, the transposon vector system is a single transposase vector system, see, e.g., WO 2008/027384. Exemplary transposases include, but are not limited to: piggyBac, Sleeping Beauty, Mos1, Tc1/mariner, Tol2, mini-Tol2, Tc3, MuA, Himar I, Frog Prince, and derivatives thereof. The piggyBac transposon and transposase are described, for example, in U.S. Patent 6,962,810, which is incorporated herein by reference in its entirety. The Sleeping Beauty transposon and transposase are described, for example, in Izsvak et al., J. Mol. Biol.302: 93-102 (2000), which is incorporated herein by reference in its entirety. The Tol2 transposon which was first isolated from the medaka fish Oryzias latipes and belongs to the hAT family of transposons is described in Kawakami et al. (2000). Mini-Tol2 is a variant of Tol2 and is described in Balciunas et al. (2006). The Tol2 and Mini-Tol2 transposons facilitate integration of a transgene into the genome of an organism when co-acting with the Tol2 transposase. The Frog Prince transposon and transposase are described, for example, in Miskey et al., Nucleic Acids Res.31:6873-6881 (2003). [00273] In some embodiments, a polynucleotide sequence encoding the antibody or antigen- binding fragment thereof disclosed herein is operably linked to one or more control elements that allow expression of the polynucleotide in both prokaryotic and eukaryotic cells. “Control elements” refer those non-translated regions of the vector which interact with host cellular proteins to carry out transcription and translation. Non-limiting examples of control elements include origin of replication, selection cassettes, constitutive and inducible promoters, enhancers, translation initiation signals (Shine Dalgarno sequence or Kozak sequence) introns, transcription terminators, 5’ and 3’ untranslated regions. See e.g., Bitter et al. (1987) Methods in Enzymology, 153:516-544) Such elements may vary in their strength and specificity. The transcriptional control element may be functional in either a eukaryotic cell (e.g., a mammalian cell) or a prokaryotic cell (e.g., bacterial or archaeal cell). [00274] In some embodiments, polynucleotides encoding the antibody or antigen-binding fragment thereof described herein are operably linked to a promoter and/or an enhancer. The term “promoter” as used herein refers to a recognition site of a polynucleotide (DNA or RNA) to which an RNA polymerase binds. An RNA polymerase initiates and transcribes polynucleotides operably linked to the promoter. In some embodiments, promoters operative in mammalian cells comprise an AT-rich region located approximately 25 to 30 bases upstream from the site where transcription is initiated and/or another sequence found 70 to 80 bases upstream from the start of transcription, a CNCAAT region where N may be any nucleotide. The term “enhancer” refers to a segment of DNA which contains sequences capable of providing enhanced transcription and in some instances can function independent of their orientation relative to another control sequence. An enhancer can function cooperatively or additively with promoters and/or other enhancer elements.
[00275] Non-limiting examples of suitable eukaryotic promoters (promoters functional in a eukaryotic cell) include those from cytomegalovirus (CMV) immediate early, herpes simplex virus (HSV) thymidine kinase, a viral simian virus 40 (SV40) (e.g., early and late SV40), a spleen focus forming virus (SFFV) promoter, long terminal repeats (LTRs) from retrovirus (e.g., a Moloney murine leukemia virus (MoMLV) LTR. promoter or a Rous sarcoma virus (RSV) LTR), a herpes simplex virus (HSV) (thymidine kinase) promoter, H5, P7.5, and Pl 1 promoters from vaccinia virus, an elongation factor 1 -alpha (EF1α) promoter, early growth response 1 (EGR1) promoter, a ferritin H (FerH) promoter, a ferritin L (FerL) promoter, a Glyceraldehyde 3 -phosphate dehydrogenase (GAPDH) promoter, a eukaryotic translation initiation factor 4A1 (EIF4A1) promoter, a heat shock 70kDa protein 5 (HSPA5) promoter, a heat shock protein 90kDa beta, member 1 (HSP90B1) promoter, a heat shock protein 70kDa (HSP70) promoter, a p-kinesin (p- KIN) promoter, the human ROSA 26 locus (Irions et al., Nature Biotechnology 25, 1477-1482 (2007)), a Ubiquitin C (UBC) promoter, a phosphoglycerate kinase-1 (PGK) promoter, a cytomegalovirus enhancer/ chicken p-actin (CAG) promoter, a P-actin promoter and a myeloproliferative sarcoma virus enhancer, negative control region deleted, dl587rev primerbinding site substituted (MND) promoter, and mouse metallothionein-1. Selection of the appropriate vector and promoter is well within the level of ordinary skill in the art.
[00276] In some embodiments, a polynucleotide sequence encoding the antibody or antigenbinding fragment thereof described herein is operably linked to a constitutive promoter. In such embodiments, the polynucleotides encoding the antibody or antigen-binding fragment thereof described herein are constitutively and/or ubiquitously expressed in a cell.
[00277] In some embodiments, a polynucleotide sequence encoding the antibody or antigenbinding fragment thereof described herein is operably linked to an inducible promoter. In such embodiments, polynucleotides encoding the antibody or antigen- binding fragment thereof described herein are conditionally expressed. As used herein, “conditional expression” may refer to any type of conditional expression including, but not limited to, inducible expression; repressible expression; expression in cells or tissues having a particular physiological, biological, or disease state (e.g., cell type or tissue specific expression) etc. Illustrative examples of inducible promoters/systems include, but are not limited to, steroid-inducible promoters such as promoters for genes encoding glucocorticoid or estrogen receptors (inducible by treatment with the corresponding hormone), metallothionine promoter (inducible by treatment with various heavy metals), MX-I promoter (inducible by interferon), the “GeneSwitch” mifepristone-regulatable system (Sirin et al., 2003, Gene, 323:67), the cumate inducible gene switch (WO 2002/088346), tetracycline-dependent regulatory systems, etc.
[00278] In some embodiments, the vectors described herein further comprise a transcription termination signal. Elements directing the efficient termination and polyadenylation of the heterologous nucleic acid transcripts increases heterologous gene expression. Transcription termination signals are generally found downstream of the polyadenylation signal. In some embodiments, vectors comprise a polyadenylation sequence 3’ of a polynucleotide encoding a polypeptide to be expressed. The term “poly A site” or “poly A sequence” as used herein denotes a DNA sequence which directs both the termination and polyadenylation of the nascent RNA transcript by RNA polymerase II. Polyadenylation sequences can promote mRNA stability' by addition of a poly A tail to the 3’ end of the coding sequence and thus, contribute to increased translational efficiency. Cleavage and polyadenylation is directed by a poly(A) sequence in the RNA. The core poly(A) sequence for mammalian pre-mRNAs has two recognition elements flanking a cleavage-polyadenylation site. Typically, an almost invariant AAUAAA hexamer lies 20-50 nucleotides upstream of a more variable element rich in U or GU residues. Cleavage of the nascent transcript occurs between these two elements and is coupled to the addition of up to 250 adenosines to the 5’ cleavage product. In some embodiments, the core poly(A) sequence is an ideal polyA sequence (e.g., AATAAA, ATTAAA, AGTAAA). In some embodiments, the poly(A) sequence is an SV40 polyA sequence, a bovine growth hormone polyA sequence (BGHpA), a rabbit P-globin polyA sequence (rPgpA), variants thereof, or another suitable heterologous or endogenous polyA sequence known in the art. [00279] The expression vector may also include nucleotide sequences encoding protein tags (e.g., 6xHis tag, hemagglutinin tag, green fluorescent protein, etc.) that are fused to the site-directed modifying polypeptide, thus resulting in a chimeric polypeptide. [00280] Methods of introducing polynucleotides and recombinant vectors into a host cell are known in the art. Suitable methods include e.g., viral or bacteriophage infection, transfection, conjugation, protoplast fusion, lipofection, electroporation, calcium phosphate precipitation, polyethyleneimine (PEI)-mediated transfection, DEAE-dextran mediated transfection, liposome- mediated transfection, particle gun technology, calcium phosphate precipitation, direct micro injection, nanoparticle-mediated nucleic acid delivery (see, e.g., Panyam et al., Adv Drug Deliv Rev. 2012 Sep 13. pii: S0169-409X(12)00283-9), microfluidics delivery methods (See e.g., International PCT Publication No. WO 2013/059343), and the like. [00281] In some embodiments, delivery via electroporation comprises mixing the cells with the polynucleotides encoding the antibody or antigen-binding fragment thereof in a cartridge, chamber, or cuvette and applying one or more electrical impulses of defined duration and amplitude. In some embodiments, cells are mixed with polynucleotides encoding the antibody or antigen-binding fragment thereof in a vessel connected to a device (e.g., a pump) which feeds the mixture into a cartridge, chamber, or cuvette wherein one or more electrical impulses of defined duration and amplitude are applied, after which the cells are delivered to a second vessel. Illustrative examples of polynucleotide delivery systems suitable for use in particular embodiments contemplated include, but are not limited to, those provided by Amaxa Biosystems, Maxcyte, Inc., BTX Molecular Delivery Systems, Neon® Transfection Systems, and Copernicus Therapeutics Inc. Lipofection reagents are sold commercially (e.g., Transfectam™ and Lipofectin™). Cationic and neutral lipids that are suitable for efficient lipofection of polynucleotides have been described in the literature. See e.g., Liu et al. (2003) Gene Therapy. 10:180–187; and Balazs et al. (2011) Journal of Drug Delivery.2011:1-12. [00282] In some embodiments, polynucleotides encoding the antibody or antigen-binding fragment thereof described herein are introduced to a cell in a non-viral delivery vehicle, such as a transposon, a nanoparticle (e.g., a lipid nanoparticle), a liposome, an exosome, an attenuated bacterium, or a virus-like particle. In some embodiments, the vehicle is an attenuated bacterium (e.g., naturally or artificially engineered to be invasive but attenuated to prevent pathogenesis including Listeria monocytogenes, certain Salmonella strains, Bifidobacterium longum, and modified Escherichia coli), bacteria having nutritional and tissue-specific tropism to target specific cells, and bacteria having modified surface proteins to alter target cell specificity. In some embodiments, the vehicle is a genetically modified bacteriophage (e.g., engineered phages having large packaging capacity, less immunogenicity, containing mammalian plasmid maintenance sequences and having incorporated targeting ligands). . In some embodiments, the vehicle is a biological liposome. For example, the biological liposome is a phospholipid-based particle derived from human cells (e.g., erythrocyte ghosts, which are red blood cells broken down into spherical structures derived from the subject and wherein tissue targeting can be achieved by attachment of various tissue or cell-specific ligands), secretory exosomes, or subject-derived membrane-bound nanovesicles (30-100 nm) of endocytic origin (e.g., can be produced from various cell types and can therefore be taken up by cells without the need for targeting ligands). [00283] In some embodiments, vectors comprising polynucleotides encoding the antibody or antigen-binding fragment thereof described herein are introduced to cells by viral delivery methods, e.g., by viral transduction. A number of viral based systems have been developed for gene transfer into mammalian cells. For example, retroviruses provide a convenient platform for gene delivery systems. The heterologous nucleic acid can be inserted into a vector and packaged in retroviral particles using techniques known in the art. The recombinant virus can then be isolated and delivered to the engineered mammalian cell in vitro or ex vivo. A number of retroviral systems are known in the art. In some embodiments, adenovirus vectors are used. A number of adenovirus vectors are known in the art. In some embodiments, lentivirus vectors are used. In some embodiments, self-inactivating lentiviral vectors are used. For example, self-inactivating lentiviral vectors carrying the immunomodulator (such as immune checkpoint inhibitor) coding sequence and/or self-inactivating lentiviral vectors carrying chimeric antigen receptors can be packaged with protocols known in the art. The resulting lentiviral vectors can be used to transduce a mammalian cell (such as primary human T cells) using methods known in the art. Vectors derived from retroviruses such as lentivirus are suitable tools to achieve long-term gene transfer, because they allow long-term, stable integration of a transgene and its propagation in progeny cells. Lentiviral vectors also have low immunogenicity, and can transduce nonproliferating cells. [00284] In some embodiments, the vehicle is a mammalian virus-like particle. For example, modified viral particles can be generated (e.g., by purification of the “empty” particles followed by ex vivo assembly of the virus with the desired cargo). [00285] In another aspect, this disclosure also provides (i) a nucleic acid molecule encoding a polypeptide chain of the antibody or antigen-binding fragment thereof described herein; (ii) a vector comprising the nucleic acid molecule as described; and (iii) a cultured host cell comprising the vector as described. Also provided is a method for producing a polypeptide, comprising: (a) obtaining the cultured host cell as described; (b) culturing the cultured host cell in a medium under conditions permitting expression of a polypeptide encoded by the vector and assembling of an antibody or fragment thereof; and (c) purifying the antibody or fragment from the cultured cell or the medium of the cell. [00286] Provided herein are polynucleotides encoding the antibody or antigen-binding fragment thereof that binds to the S polypeptide of a Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) disclosed herein. [00287] Polynucleotides disclosed herein can be at least about 5, at least about 10, at least about 15, at least about 20, at least about 25, at least about 30, at least about 40, at least about 50, at least about 100, at least about 200, at least about 300, at least about 400, at least about 500, at least about 1000, at least about 5000, at least about 10000, or at least about 15000 or more nucleotides in length, as well as all intermediate lengths. It will be readily understood that “intermediate lengths,” in this context, means any length between the quoted values, such as 6, 7, 8, 9, etc., 101, 102, 103, etc.; 151, 152, 153, etc.; 201, 202, 203, etc. [00288] The present disclosure further relates to variants of the polynucleotides disclosed herein. The polynucleotide variants can contain alterations in the coding regions, non-coding regions, or both. [00289] In some embodiments, a polynucleotide variant comprising a nucleotide sequence that is at least about 75 %, about 80 %, about 85 %, about 90 %, about 91 %, about 92 %, about 93 %, about 94 %, about 95 %, about 96 %, about 97 %, about 98%, or about 99 % identical to the nucleotide sequence of a polynucleotide disclosed herein. [00290] In some embodiments, a polynucleotide variant contains substitutions, additions, or deletions that alter the properties or activities of the encoded polypeptide. In some embodiments, a polynucleotide variant contains silent substitutions, additions, or deletions that does not alter the properties or activities of the encoded polypeptide. In some embodiments, a polynucleotide variant is produced to modulate or alter expression (or expression levels) of the encoded polypeptide. In some embodiments, a polynucleotide variant is produced to increase expression of the encoded polypeptide. In some embodiments, a polynucleotide variant is produced to decrease expression of the encoded polypeptide. In some embodiments, a polynucleotide variant has increased expression of the encoded polypeptide as compared to a parental polynucleotide sequence. In some embodiments, a polynucleotide variant has decreased expression of the encoded polypeptide as compared to a parental polynucleotide sequence. [00291] In some embodiments, polynucleotides are codon-optimized. As used herein, the term “codon-optimized” refers to substituting codons in a polynucleotide encoding a polypeptide in order to increase the expression, stability and/or activity of the polypeptide. Factors that influence codon optimization include, but are not limited to one or more of: (i) variation of codon biases between two or more organisms or genes or synthetically constructed bias tables, (ii) variation in the degree of codon bias within an organism, gene, or set of genes, (iii) systematic variation of codons including context, (iv) variation of codons according to their decoding tRNAs, (v) variation of codons according to GC %, either overall or in one position of the triplet, (vi) variation in degree of similarity to a reference sequence for example a naturally occurring sequence, (vii) variation in the codon frequency cutoff, (viii) structural properties of mRNAs transcribed from the DNA sequence, (ix) prior knowledge about the function of the DNA sequences upon which design of the codon substitution set is to be based, (x) systematic variation of codon sets for each amino acid, (xi) isolated removal of spurious translation initiation sites and/or (xii) elimination of fortuitous polyadenylation sites otherwise leading to truncated RNA transcripts. [00292] It will be appreciated by those of ordinary skill in the art that, as a result of the degeneracy of the genetic code, there are many nucleotide sequences that encode a polypeptide, or fragment of variant thereof, as described herein. Some of these polynucleotides bear minimal homology to the nucleotide sequence of any native gene. Nonetheless, polynucleotides that vary due to differences in codon usage are specifically contemplated in particular embodiments, for example polynucleotides that are optimized for human and/or primate codon selection. Further, alleles of the genes comprising the polynucleotide sequences provided herein may also be used. Alleles are endogenous genes that are altered as a result of one or more mutations, such as deletions, additions and/or substitutions of nucleotides. [00293] The polynucleotides contemplated herein, regardless of the length of the coding sequence itself, may be combined with other DNA sequences, such as promoters and/or enhancers, untranslated regions (UTRs), signal sequences, Kozak sequences, polyadenylation signals, additional restriction enzyme sites, multiple cloning sites, internal ribosomal entry sites (IRES), recombinase recognition sites (e.g., LoxP, FRT, and Att sites), termination codons, transcriptional termination signals, and polynucleotides encoding self-cleaving polypeptides, epitope tags, as disclosed elsewhere herein or as known in the art, such that their overall length may vary considerably. It is therefore contemplated that a polynucleotide fragment of almost any length may be employed in particular embodiments, with the total length preferably being limited by the ease of preparation and use in the intended recombinant DNA protocol. [00294] Polynucleotides can be prepared, isolated, purified, manipulated, and/or expressed using any of a variety of well-established techniques known and available in the art. 1.4. Methods of Production [00295] Antibodies may be produced using recombinant methods and compositions, e.g., as described in U.S. Pat. No. 4,816,567. In one embodiment, an isolated nucleic acid encoding an antibody described herein is provided. Such nucleic acid may encode an amino acid sequence comprising the VL and/or an amino acid sequence comprising the VH of the antibody (e.g., the light and/or heavy chains of the antibody). In a further embodiment, one or more vectors (e.g., expression vectors) comprising such nucleic acid are provided. In a further embodiment, a host cell comprising such nucleic acid is provided. In one such embodiment, a host cell comprises (e.g., has been transformed with): (1) a vector comprising a nucleic acid that encodes an amino acid sequence comprising the VL of the antibody and an amino acid sequence comprising the VH of the antibody, or (2) a first vector comprising a nucleic acid that encodes an amino acid sequence comprising the VL of the antibody and a second vector comprising a nucleic acid that encodes an amino acid sequence comprising the VH of the antibody. In one embodiment, the host cell is eukaryotic, e.g., a Chinese Hamster Ovary (CHO) cell or lymphoid cell (e.g., Y0, NS0, Sp20 cell). In one embodiment, a method of making an antibody is provided, wherein the method comprises culturing a host cell comprising a nucleic acid encoding the antibody, as provided above, under conditions suitable for expression of the antibody, and optionally recovering the antibody from the host cell (or host cell culture medium). [00296] For recombinant production of an antibody, a nucleic acid encoding an antibody, e.g., as described herein, is isolated and inserted into one or more vectors for further cloning and/or expression in a host cell. Such nucleic acid may be readily isolated and sequenced using conventional procedures (e.g., by using oligonucleotide probes that are capable of binding specifically to genes encoding the heavy and light chains of the antibody). [00297] Suitable host cells for cloning or expression of antibody-encoding vectors include prokaryotic or eukaryotic cells described herein. For example, antibodies may be produced in bacteria, in particular when glycosylation and Fc effector function are not needed. For expression of antibody fragments and polypeptides in bacteria, see, e.g., U.S. Pat. Nos.5,648,237, 5,789,199, and 5,840,523. (See also Charlton, Methods in Molecular Biology, Vol. 248 (B.K.C. Lo, ed., Humana Press, Totowa, N.J., 2003), pp. 245-254, describing expression of antibody fragments in E. coli.) After expression, the antibody may be isolated from the bacterial cell paste in a soluble fraction and can be further purified. [00298] In addition to prokaryotes, eukaryotic microbes such as filamentous fungi or yeast are suitable cloning or expression hosts for antibody-encoding vectors, including fungi and yeast strains whose glycosylation pathways have been “humanized,” resulting in the production of an antibody with a partially or fully human glycosylation pattern. See Gerngross, Nat. Biotech. 22:1409-1414 (2004), and Li et al., Nat. Biotech. 24:210-215 (2006). [00299] Suitable host cells for the expression of glycosylated antibodies are also derived from multicellular organisms (invertebrates and vertebrates). Examples of invertebrate cells include plant and insect cells. Numerous baculoviral strains have been identified, which may be used in conjunction with insect cells, particularly for transfection of Spodoptera frugiperda cells. [00300] Plant cell cultures can also be utilized as hosts. See, e.g., U.S. Pat. Nos. 5,959,177, 6,040,498, 6,420,548, 7,125,978, and 6,417,429 (describing PLANTIBODIES technology for producing antibodies in transgenic plants). [00301] Vertebrate cells may also be used as hosts. For example, mammalian cell lines that are adapted to grow in suspension may be useful. Other examples of useful mammalian host cell lines are monkey kidney CV1 line transformed by SV40 (COS-7); human embryonic kidney line (293 or 293 cells as described, e.g., in Graham et al., J. Gen Virol. 36:59 (1977)); baby hamster kidney cells (BHK); mouse sertoli cells (TM4 cells as described, e.g., in Mather, Biol. Reprod. 23:243- 251 (1980)); monkey kidney cells (CV1); African green monkey kidney cells (VERO-76); human cervical carcinoma cells (HELA); canine kidney cells (MDCK; buffalo rat liver cells (BRL 3A); human lung cells (W138); human liver cells (Hep G2); mouse mammary tumor (MMT 060562); TRI cells, as described, e.g., in Mather et al., Annals N.Y. Acad. Sci. 383:44-68 (1982); MRC 5 cells; and FS4 cells. Other useful mammalian host cell lines include CHO cells, including DHFR- CHO cells (Urlaub et al., Proc. Natl. Acad. Sci. USA 77:4216 (1980)); and myeloma cell lines such as Y0, NS0, and Sp2/0. For a review of certain mammalian host cell lines suitable for antibody production, see, e.g., Yazaki and Wu, Methods in Molecular Biology, Vol. 248 (B.K.C. Lo, ed., Humana Press, Totowa, N.J.), pp. 255-268 (2003). [00302] Monoclonal antibodies or functional fragments thereof (e.g., antigen-binding fragment thereof) may be made using, for example, the hybridoma method or the phage display method. [00303] In the hybridoma method (see, e.g., described in Kohler, et al., Nature, 1975, 256:495- 7), a mouse or other appropriate host animal, such as a hamster, is immunized as described above to elicit lymphocytes that produce or are capable of producing antibodies that will specifically bind to the protein used for immunization. Alternatively, lymphocytes may be immunized in vitro. After immunization, lymphocytes are isolated and then fused with a myeloma cell line using a suitable fusing agent, such as polyethylene glycol, to form a hybridoma cell (Goding, Monoclonal Antibodies: Principles and Practice 59-103 (1986)). [00304] The hybridoma cells thus prepared are seeded and grown in a suitable culture medium, which, in some embodiments, contains one or more substances that inhibit the growth or survival of the unfused, parental myeloma cells (also referred to as fusion partner). For example, if the parental myeloma cells lack the enzyme hypoxanthine guanine phosphoribosyl transferase (HGPRT or HPRT), the selective culture medium for the hybridomas typically will include hypoxanthine, aminopterin, and thymidine (HAT medium), which prevent the growth of HGPRT- deficient cells. [00305] Exemplary parental myeloma cells are those that fuse efficiently, support stable high- level production of antibody by the selected antibody-producing cells, and are sensitive to a selective medium that selects against the unfused parental cells. Exemplary myeloma cell lines are murine myeloma lines, such as SP-2 and derivatives, for example, X63-Ag8-653 cells available from the American Type Culture Collection (Manassas, VA), and those derived from MOPC-21 and MPC-11 mouse tumors available from the Salk Institute Cell Distribution Center (San Diego, CA). Human myeloma and mouse-human heteromyeloma cell lines also have been described for the production of human monoclonal antibodies (Kozbor, Immunol. 1984, 133:3001-05; and Brodeur, et al., Monoclonal Antibody Production Techniques and Applications, 1987, 51-63). [00306] Culture medium in which hybridoma cells are growing is assayed for production of monoclonal antibodies directed against the antigen. The binding specificity of monoclonal antibodies produced by hybridoma cells is determined by immunoprecipitation or by an in vitro binding assay, such as RIA or ELISA. The binding affinity of the monoclonal antibody can, for example, be determined by the Scatchard analysis described in Munson et al., Anal. Biochem., 1980, 107:220-39. [00307] Once hybridoma cells that produce antibodies of the desired specificity, affinity, and/or activity are identified, the clones may be subcloned by limiting dilution procedures and grown by standard methods (Goding, supra). Suitable culture media for this purpose include, for example, DMEM or RPMI-1640 medium. In addition, the hybridoma cells may be grown in vivo as ascites tumors in an animal, for example, by i.p. injection of the cells into mice. [00308] The monoclonal antibodies secreted by the subclones are suitably separated from the culture medium, ascites fluid, or serum by conventional antibody purification procedures such as, for example, affinity chromatography (e.g., using protein A or protein G-Sepharose) or ion- exchange chromatography, hydroxyapatite chromatography, gel electrophoresis, dialysis, etc. [00309] In the phage display method, synthetic antibody clones are selected by screening phage libraries containing phages that display various fragments of antibody variable region (Fv) fused to phage coat protein. Such phage libraries are screened against the desired antigen. Clones expressing Fv fragments capable of binding to the desired antigen are adsorbed to the antigen and thus separated from the non-binding clones in the library. The binding clones are then eluted from the antigen and can be further enriched by additional cycles of antigen absorption/elution. [00310] Variable domains can be displayed functionally on phage, either as single-chain Fv (scFv) fragments, in which VH and VL are covalently linked through a short, flexible peptide, or as Fab fragments, in which they are each fused to a constant domain and interact non-covalently, as described, for example, in Winter et al., 1994, Ann. Rev. Immunol.12:433-55. [00311] Repertoires of VH and VL genes can be separately cloned by PCR and recombined randomly in phage libraries, which can then be searched for antigen-binding clones as described in Winter et al, supra. Libraries from immunized sources provide high-affinity antibodies to the antigen without the requirement of constructing hybridomas. Alternatively, naive libraries can be cloned to provide a single source of human antibodies to a wide range of non-self and self-antigens without any immunization as described by Griffiths et al., EMBO J, 1993, 12:725-34. Finally, naive libraries can also be made synthetically by cloning the unrearranged V-gene segments from stem cells, and using PCR primers containing random sequence to encode the highly variable CDR3 regions and to accomplish rearrangement in vitro as described, for example, by Hoogenboom and Winter, J. Mol. Biol., 1992, 227:381-88. [00312] Screening of the libraries can be accomplished by various techniques known in the art. For example, the antigen can be used to coat the wells of adsorption plates, expressed on host cells affixed to adsorption plates or used in cell sorting, conjugated to biotin for capture with streptavidin-coated beads, or used in any other method for panning display libraries. The selection of antibodies with slow dissociation kinetics (e.g., good binding affinities) can be promoted by use of long washes and monovalent phage display as described in Bass, et al, Proteins, 1990, 8:309- 14 and WO 92/09690, and by use of a low coating density of antigen as described in Marks et al, BiotechnoL, 1992, 10:779-83. [00313] Exemplary phage display methods that can be used herein include those disclosed in Antibody Phage Display: Methods and Protocols (O’Brien and Aitken, eds., 2002); Brinkman, et al, J. Immunol. Methods, 1995, 182:41-50; Ames, et al., Immunol. Methods, 1995, 184:177-86; Kettleborough, et al., Eur. J. Immunol., 1994, 24:952-8; Persic, et al., Gene, 1997, 187:9-18; Burton et al., Advances in Immunology, 1994, 57:191-280; PCT Application No. PCT/GB91/01 134; International Publication Nos. WO 90/02809, WO 91/10737, WO 92/01047, WO 92/18619, WO 93/11236, WO 95/15982, WO 95/20401, and W097/13844; and U.S. Patent Nos.5,698,426, 5,223,409, 5,403,484, 5,580,717, 5,427,908, 5,750,753, 5,821,047, 5,571,698, 5,427,908, 5,516,637, 5,780,225, 5,658,727, 5,733,743 and 5,969,108. [00314] DNA encoding the monoclonal antibodies is readily isolated from the hybridoma cells and the screened libraries. Such DNA can be sequenced using conventional procedures (e.g., by using oligonucleotide probes that are capable of binding specifically to genes encoding the heavy and light chains of murine antibodies). Once isolated, the DNA may be placed into expression vectors, which are then transfected into host cells, such as E. coli cells, simian COS cells, Chinese Hamster Ovary (CHO) cells, or myeloma cells that do not otherwise produce antibody protein, to obtain the synthesis of monoclonal antibodies in the recombinant host cells. Review articles on recombinant expression in bacteria of DNA encoding the antibody include Skerra, et al., Curr. Opinion in Immunol., 1993, 5:256-62 and Pluckthun, Immunol. Revs., 1992, 130:151-88. [00315] Once an antibody molecule provided herein has been produced by recombinant expression, it may be purified by any method known in the art for purification of an immunoglobulin molecule, for example, by chromatography (e.g., ion exchange, affinity, particularly by affinity for the specific antigen after Protein A, and sizing column chromatography), centrifugation, differential solubility, or by any other standard technique for the purification of proteins. Further, the antibodies provided herein can be fused to heterologous polypeptide sequences described herein or otherwise known in the art to facilitate purification. Humanized, Human, and Recombinant Antibodies [00316] The antibodies disclosed herein can comprise “chimeric” sequences in which a portion of the heavy and/or light chain is identical with or homologous to corresponding sequences in antibodies derived from a particular species or belonging to a particular antibody class or subclass, while the remainder of the chain(s) is identical with or homologous to corresponding sequences in antibodies derived from another species or belonging to another antibody class or subclass, so long as they exhibit the desired biological activity (see U S. Pat. No. 4,816,567; and Morrison, et al. , Proc. Natl. Acad. Sci. USA , 1984, 81:6851-55). [00317] The antibodies disclosed herein can be humanized antibodies. A humanized antibody can comprise human framework region and human constant region sequences. In some embodiments, one or more FR region residues of the human immunoglobulin are replaced by corresponding nonhuman residues. In some embodiments, humanized antibodies comprise residues that are not found in the recipient antibody or in the donor antibody. These modifications are made to further refine antibody performance. [00318] Humanized antibodies can be produced using a variety of techniques known in the art, including but not limited to, CDR-grafting (European Patent No. EP 239,400; International publication No. WO 91/09967; and U S. Patent Nos. 5,225,539, 5,530,101, and 5,585,089), veneering or resurfacing (European Patent Nos. EP 592,106 and EP 519,596; Padlan, Molecular Immunology, 1991, 28(4/5):489-498; Studnicka, et al., Protein Engineering, 1994, 7(6):805-814; and Roguska, et al., Proc. Natl. Acad. Sci. USA, 1994, 91:969-73), chain shuffling (U.S. Patent No.5,565,332), and techniques disclosed in, e.g., U.S. Pat. No.6,407,213, U.S. Pat. No.5,766,886, WO 93/17105; Tan, et al., J. Immunol., 2002, 169:1119-25; Caldas, et al., Protein Eng., 2000, 13(5):353-60; Morea et al., Methods, 2000, 20(3):267-79, Baca, et al., J. Biol. Chem., 1997, 272(16): 10678-84; Roguska, et al., Protein Eng., 1996, 9(10):895904; Couto, et al., Cancer Res., 1995, 55 (23 Supp):5973s-5977s; Couto, et al., Cancer Res., 1995, 55(8): 1717-22; Sandhu, J.S., Gene, 1994, 150(2):409-10 and Pedersen, et al., J. Mol. Biol., 1994, 235(3):959-73. See also U.S. Patent Pub. No. US 2005/0042664 Al (Feb. 24, 2005), each of which is incorporated by reference herein in its entirety. In some embodiments, the humanized antibodies are constructed by CDR grafting, in which the amino acid sequences of the six CDRs of the parent non-human antibody (e.g., rodent) are grafted onto a human antibody framework. For example, Padlan, et al. determined that only about one third of the residues in the CDRs actually contact the antigen, and termed these the “specificity determining residues,” or SDRs (Padlan, et al, FASEB J., 1995, 9: 133-9). In the technique of SDR grafting, only the SDR residues are grafted onto the human antibody framework (see, e.g., Kashmiri, et al, Methods, 2005, 36:25-34). [00319] The choice of human variable domains, both light and heavy, to be used in making the humanized antibodies can be important to reduce antigenicity. For example, according to the so- called “best-fit” method, the sequence of the variable domain of a non-human (e.g., rodent) antibody is screened against the entire library of known human variable-domain sequences. The human sequence that is closest to that of the rodent may be selected as the human framework for the humanized antibody (Sims et al., J. Immunol., 1993, 151:2296-308; and Chothia et al., J. Mol. Biol., 1987, 196:901-17). [00320] Another method uses a particular framework derived from the consensus sequence of all human antibodies of a particular subgroup of light or heavy chains. The same framework may be used for several different humanized antibodies (Carter et al., Proc. Natl. Acad. Sci. USA, 1992, 89:4285-89; and Presta et al., J Immunol., 1993, 151:2623-32). In some embodiments, the framework is derived from the consensus sequences of the most abundant human subclasses, VL6 subgroup I (VL6I) and VH subgroup III (VHIII). [00321] In an alternative method based on comparison of CDRs, called superhumanization, FR homology is irrelevant. The method consists of comparison of the non-human sequence with the functional human germline gene repertoire. Those genes encoding the same or closely related canonical structures to the murine sequences are then selected. Next, within the genes sharing the canonical structures with the non-human antibody, those with highest homology within the CDRs are chosen as FR donors. Finally, the non-human CDRs are grafted onto these FRs (see, e.g., Tan et al., J. Immunol., 2002, 169:1119-25). [00322] Another method for antibody humanization is based on a metric of antibody humanness termed Human String Content (HSC). This method compares the mouse sequence with the repertoire of human germline genes, and the differences are scored as HSC. The target sequence is then humanized by maximizing its HSC rather than using a global identity measure to generate multiple diverse humanized variants (Lazar et al.,MoI. Immunol., 2007, 44:1986-98). [00323] In addition to the methods described above, empirical methods may be used to generate and select humanized antibodies. These methods include those that are based upon the generation of large libraries of humanized variants and selection of the best clones using enrichment technologies or high throughput screening techniques. Antibody variants may be isolated from phage, ribosome, and yeast display libraries as well as by bacterial colony screening (see, e.g., Hoogenboom, Nat. Biotechnol., 2005, 23:1105-16; Dufner, et al., Trends Biotechnol., 2006, 24:523-9; Feldhaus, et al., Nat. Biotechnol, 2003, 21:163-70; and Schlapschy et al., Protein Eng. Des. Sel, 2004, 17:847-60). [00324] In the FR library approach, a collection of residue variants are introduced at specific positions in the FR followed by screening of the library to select the FR that best supports the grafted CDR. The residues to be substituted may include some or all of the “Vernier” residues identified as potentially contributing to CDR structure (see, e.g., Foote and Winter, J. Mol. Biol., 1992, 224:487-99), or from the more limited set of target residues identified by Baca, et al, J. Biol. Chem., 1997, 272:10678-84. [00325] In FR shuffling, whole FRs are combined with the non-human CDRs instead of creating combinatorial libraries of selected residue variants (see, e.g., DalTAcqua et al., Methods, 2005, 36:43-60). The libraries may be screened for binding in a two-step process, first humanizing VL, followed by VH. Alternatively, a one-step FR shuffling process may be used. Such a process has been shown to be more efficient than the two-step screening, as the resulting antibodies exhibited improved biochemical and physicochemical properties including enhanced expression, increased affinity, and thermal stability (see, e.g., Damschroder, et al.,Mol. Immunol., 2007, 44:3049-60). [00326] The “Humaneering™” method is based on experimental identification of essential minimum specificity determinants (MSDs) and is based on sequential replacement of non-human fragments into libraries of human FRs and assessment of binding. It begins with regions of the CDR3 of non-human VH and VL chains and progressively replaces other regions of the non- human antibody into the human FRs, including the CDR1 and CDR2 of both VH and VL. This methodology typically results in epitope retention and identification of antibodies from multiple subclasses with distinct human V-segment CDRs. Humaneering™ allows for isolation of antibodies that are 91-96% homologous to human germline gene antibodies (see, e.g., Alfenito, Cambridge Healthtech Institute’s Third Annual PEGS, The Protein Engineering Summit, 2007). [00327] The “human engineering” method involves altering a non-human antibody or antibody fragment, such as a mouse or chimeric antibody or antibody fragment, by making specific changes to the amino acid sequence of the antibody so as to produce a modified antibody with reduced immunogenicity in a human that nonetheless retains the desirable binding properties of the original non-human antibodies. Generally, the technique involves classifying amino acid residues of a non- human (e.g., mouse) antibody as “low risk,” “moderate risk,” or “high risk” residues. The classification is performed using a global risk/reward calculation that evaluates the predicted benefits of making particular substitution (e.g., for immunogenicity in humans) against the risk that the substitution will affect the resulting antibody’s folding. The particular human amino acid residue to be substituted at a given position (e.g., low or moderate risk) of a non-human (e.g., mouse) antibody sequence can be selected by aligning an amino acid sequence from the non- human antibody’s variable regions with the corresponding region of a specific or consensus human antibody sequence. The amino acid residues at low or moderate risk positions in the non-human sequence can be substituted for the corresponding residues in the human antibody sequence according to the alignment. Techniques for making human engineered proteins are described in greater detail in Studnicka et al., Protein Engineering, 1994, 7:805-14; U.S. Pat. Nos. 5,766,886; 5,770,196; 5,821,123; and 5,869,619; and PCT Publication WO 93/11794. [00328] The antibodies disclosed herein can be composite human antibodies. A composite human antibody can be generated using, for example, Composite Human Antibody™ technology (Antitope Ltd., Cambridge, United Kingdom). To generate composite human antibodies, variable region sequences are designed from fragments of multiple human antibody variable region sequences in a manner that avoids T cell epitopes, thereby minimizing the immunogenicity of the resulting antibody. Such antibodies can comprise human constant region sequences, e.g., human light chain and/or heavy chain constant regions. [00329] The antibodies disclosed herein can be deimmunized antibodies. A deimmunized antibody is an antibody in which T-cell epitopes have been removed. Methods for making deimmunized antibodies have been described (see, e.g., Jones, et al., Methods Mol Biol., 2009, 525:405-23; and De Groot, et al., Cell. Immunol., 2006, 244:148-153). Deimmunized antibodies comprise T-cell epitope-depleted variable regions and human constant regions. Briefly, VH and VL of an antibody are cloned and T-cell epitopes are subsequently identified by testing overlapping peptides derived from the VH and VL of the antibody in a T cell proliferation assay. T cell epitopes are identified via in silico methods to identify peptide binding to human MHC class II. Mutations are introduced in the VH and VL to abrogate binding to human MHC class II. Mutated VH and VL are then utilized to generate the deimmunized antibody. [00330] It is further generally desirable that antibodies be humanized with retention of their affinity for the antigen and other favorable biological properties. To achieve this goal, according to one method, humanized antibodies are prepared by a process of analysis of the parental sequences and various conceptual humanized products using three-dimensional models of the parental and humanized sequences. Three-dimensional immunoglobulin models are commonly available and are familiar to those skilled in the art. Computer programs are available which illustrate and display probable three-dimensional conformational structures of selected candidate immunoglobulin sequences. These include, for example, WAM (Whitelegg and Rees, Protein Eng., 2000, 13:819-24), Modeller (Sali and Blundell, J. Mol. Biol., 1993, 234:779-815), and Swiss PDB Viewer (Guex and Peitsch, Electrophoresis, 1997, 18:2714-23). Inspection of these displays permits analysis of the likely role of the residues in the functioning of the candidate immunoglobulin sequence, e.g., the analysis of residues that influence the ability of the candidate immunoglobulin to bind its antigen. In this way, FR residues can be selected and combined from the recipient and import sequences so that the desired antibody characteristic, such as increased affinity for the target antigen(s), is achieved. In general, the CDR residues are directly and most substantially involved in influencing antigen-binding. [00331] The antibodies disclosed herein can be fully human antibodies, which possesses an amino acid sequence corresponding to that of an antibody produced by a human. This definition of a human antibody specifically excludes a humanized antibody comprising non-human antigen- binding residues. Human antibodies can be produced using various techniques known in the art, including phage-display libraries (Hoogenboom and Winter, J. Mol. Biol, 1991, 227:381; Marks, et al., 1991, J. Mol. Biol., 1991, 222:581) and yeast display libraries (Chao, et al., Nature Protocols , 2006, 1: 755-68). Also available for the preparation of human monoclonal antibodies are methods described in Cole, et al., Monoclonal Antibodies and Cancer Therapy 77 (1985): Boerner, et al., J. Immunol., 1991, 147(l):86-95; and van Dijk and van de Winkel, Curr. Opin. Pharmacol, 2001, 5: 368-74. Human antibodies can also be prepared by administering the antigen to a transgenic animal that has been modified to produce such antibodies in response to antigenic challenge, but whose endogenous loci have been disabled, e.g., mice (see, e.g., Jakobovits, Curr. Opin. Biotechnol., 1995, 6(5):561-66; Bruggemann and Taussing, Curr. Opin. Biotechnol., 1997, 8(4):455-58; and U.S. Pat. Nos. 6,075,181 and 6,150,584 regarding XENOMOUSE™ technology). See also, for example, Li, et al., Proc. Natl. Acad. Sci. USA , 2006, 103:3557-62, regarding human antibodies generated via a human B-cell hybridoma technology. [00332] The antibodies disclosed herein can be recombinant human antibodies, which are human antibodies prepared, expressed, created or isolated by recombinant means, such as antibodies expressed using a recombinant expression vector transfected into a host cell, antibodies isolated from a recombinant, combinatorial human antibody library, antibodies isolated from an animal (e.g., a mouse or cow) that is transgenic and/or transchromosomal for human immunoglobulin genes (see e.g., Taylor, L. D., et al., Nucl. Acids Res., 199220:6287-6295) or antibodies prepared, expressed, created or isolated by any other means that involves splicing of human immunoglobulin gene sequences to other DNA sequences. In some embodiments, such recombinant human antibodies can have variable and constant regions derived from human germline immunoglobulin sequences (See Kabat, E. A., et al. (1991) Sequences of Proteins of Immunological Interest Fifth Edition, U.S. Department of Health and Human Services, NIH Publication No.91-3242). In some embodiments, such recombinant human antibodies are subjected to in vitro mutagenesis (or, when an animal transgenic for human Ig sequences is used, in vivo somatic mutagenesis) and thus the amino acid sequences of the VH and VL regions of the recombinant antibodies are sequences that, while derived from and related to human germline VH and VL sequences, may not naturally exist within the human antibody germline repertoire in vivo. Modifications, Variations, and Derivatives [00333] Amino acid sequence modification(s) of the antibody or antigen-binding fragment thereof provided herein are contemplated. For example, it may be desirable to improve the binding affinity between the antibody or antigen-binding fragment thereof and the S polypeptide of a SARS-CoV-2; it may also be desirable to improve other biological properties of the antibody or antigen-binding fragment thereof, including but not limited to specificity, thermostability, expression level, or solubility. Thus, in addition to the antibody or antigen-binding fragment thereof described herein, it is contemplated that variants can be prepared. [00334] In some embodiments, the antibody or antigen-binding fragment thereof provided herein are chemically modified, for example, by the covalent attachment of any type of molecule to the antibody or antigen-binding fragment thereof. Exemplary non-limiting modifications include glycosylation, acetylation, pegylation, phosphorylation, amidation, derivatization by known protecting/blocking groups, proteolytic cleavage, linkage to a cellular ligand or other protein, etc. Additionally, the antibody or antigen-binding fragment thereof may contain one or more non-classical amino acids. [00335] In some embodiments, variations may be a substitution, deletion, or insertion of one or more codons encoding the antibody or antigen-binding fragment thereof that results in a change in the amino acid sequence as compared with the original sequence. The variation allowed may be determined by systematically making insertions, deletions, or substitutions of amino acids in the sequence and testing the resulting variants for activity exhibited by the full-length or mature native sequence. [00336] Amino acid substitutions can be the result of replacing one amino acid with another amino acid having similar structural and/or chemical properties, such as the replacement of a leucine with a serine, e.g., conservative amino acid replacements. Standard techniques known to those of skill in the art can be used to introduce mutations in the nucleotide sequence encoding a molecule provided herein, including, for example, site-directed mutagenesis and PCR-mediated mutagenesis which results in amino acid substitutions. [00337] A “conservative amino acid substitution” is one in which the amino acid residue is replaced with an amino acid residue having a side chain with a similar charge. Families of amino acid residues having side chains with similar charges have been defined in the art. These families include amino acids with basic side chains (e.g., lysine, arginine, histidine), acidic side chains (e.g., aspartic acid, glutamic acid), uncharged polar side chains (e.g., glycine, asparagine, glutamine, serine, threonine, tyrosine, cysteine), nonpolar side chains (e.g., alanine, valine, leucine, isoleucine, proline, phenylalanine, methionine, tryptophan), beta-branched side chains (e.g., threonine, valine, isoleucine) and aromatic side chains (e.g., tyrosine, phenylalanine, tryptophan, histidine). Alternatively, mutations can be introduced randomly along all or part of the coding sequence, such as by saturation mutagenesis, and the resultant mutants can be screened for biological activity to identify mutants that retain activity. Following mutagenesis, the encoded protein can be expressed and the activity of the protein can be determined. [00338] Substantial modifications in the biological properties of the antibody are accomplished by selecting substitutions that differ significantly in their effect on maintaining (a) the structure of the polypeptide backbone in the area of the substitution, for example, as a sheet or helical conformation, (b) the charge or hydrophobicity of the molecule at the target site, or (c) the bulk of the side chain. [00339] Substitutions may be in the range of about 1 to 100 amino acids. In some embodiments, the substitution includes fewer than about 100 amino acid substitutions, fewer than about 95 amino acid substitutions, fewer than about 90 amino acid substitutions, fewer than about 85 amino acid substitutions, fewer than about 80 amino acid substitutions, fewer than about 75 amino acid substitutions, fewer than about 70 amino acid substitutions, fewer than about 65 amino acid substitutions, fewer than about 50 amino acid substitutions, fewer than about 45 amino acid substitutions, fewer than about 40 amino acid substitutions, fewer than about 35 amino acid substitutions, fewer than about 30 amino acid substitutions, fewer than about 25 amino acid substitutions, fewer than about 20 amino acid substitutions, fewer than about 15 amino acid substitutions, fewer than about 10 amino acid substitutions, fewer than about 5 amino acid substitutions, fewer than about 4 amino acid substitutions, fewer than about 3 amino acid substitutions, or fewer than about 2 amino acid substitutions relative to the original molecule. [00340] Amino acid sequence insertions include amino- and/or carboxyl-terminal fusions ranging in length from 1 residue to polypeptides containing 100 or more residues, as well as intrasequence insertions of single or multiple amino acid residues. Examples of terminal insertions include an antibody with an N-terminal methionyl residue. Other insertional variants of the antibody molecule include the fusion to the N or C terminus of the antibody to an enzyme (e.g., for antibody-directed enzyme prodrug therapy) or a polypeptide which increases the serum half- life of the antibody. [00341] In some embodiments, the insertion is about 1 amino acid to about 100 amino acids. In some embodiments, the insertion is at least about 1 amino acid. In some embodiments, the insertion is at most about 100 amino acids. In some embodiments, the insertion is about 1 amino acid to about 5 amino acids, about 1 amino acid to about 10 amino acids, about 1 amino acid to about 20 amino acids, about 1 amino acid to about 30 amino acids, about 1 amino acid to about 40 amino acids, about 1 amino acid to about 50 amino acids, about 1 amino acid to about 60 amino acids, about 1 amino acid to about 70 amino acids, about 1 amino acid to about 80 amino acids, about 1 amino acid to about 90 amino acids, about 1 amino acid to about 100 amino acids, about 5 amino acids to about 10 amino acids, about 5 amino acids to about 20 amino acids, about 5 amino acids to about 30 amino acids, about 5 amino acids to about 40 amino acids, about 5 amino acids to about 50 amino acids, about 5 amino acids to about 60 amino acids, about 5 amino acids to about 70 amino acids, about 5 amino acids to about 80 amino acids, about 5 amino acids to about 90 amino acids, about 5 amino acids to about 100 amino acids, about 10 amino acids to about 20 amino acids, about 10 amino acids to about 30 amino acids, about 10 amino acids to about 40 amino acids, about 10 amino acids to about 50 amino acids, about 10 amino acids to about 60 amino acids, about 10 amino acids to about 70 amino acids, about 10 amino acids to about 80 amino acids, about 10 amino acids to about 90 amino acids, about 10 amino acids to about 100 amino acids, about 20 amino acids to about 30 amino acids, about 20 amino acids to about 40 amino acids, about 20 amino acids to about 50 amino acids, about 20 amino acids to about 60 amino acids, about 20 amino acids to about 70 amino acids, about 20 amino acids to about 80 amino acids, about 20 amino acids to about 90 amino acids, about 20 amino acids to about 100 amino acids, about 30 amino acids to about 40 amino acids, about 30 amino acids to about 50 amino acids, about 30 amino acids to about 60 amino acids, about 30 amino acids to about 70 amino acids, about 30 amino acids to about 80 amino acids, about 30 amino acids to about 90 amino acids, about 30 amino acids to about 100 amino acids, about 40 amino acids to about 50 amino acids, about 40 amino acids to about 60 amino acids, about 40 amino acids to about 70 amino acids, about 40 amino acids to about 80 amino acids, about 40 amino acids to about 90 amino acids, about 40 amino acids to about 100 amino acids, about 50 amino acids to about 60 amino acids, about 50 amino acids to about 70 amino acids, about 50 amino acids to about 80 amino acids, about 50 amino acids to about 90 amino acids, about 50 amino acids to about 100 amino acids, about 60 amino acids to about 70 amino acids, about 60 amino acids to about 80 amino acids, about 60 amino acids to about 90 amino acids, about 60 amino acids to about 100 amino acids, about 70 amino acids to about 80 amino acids, about 70 amino acids to about 90 amino acids, about 70 amino acids to about 100 amino acids, about 80 amino acids to about 90 amino acids, about 80 amino acids to about 100 amino acids, or about 90 amino acids to about 100 amino acids. In some embodiments, the insertion is about 1 amino acid, about 5 amino acids, about 10 amino acids, about 20 amino acids, about 30 amino acids, about 40 amino acids, about 50 amino acids, about 60 amino acids, about 70 amino acids, about 80 amino acids, about 90 amino acids, or about 100 amino acids. [00342] Amino acid sequence deletions include amino- and/or carboxyl-terminal deletions ranging in length from one residue to polypeptides containing a hundred or more residues, as well as intrasequence deletions of single or multiple amino acid residues. [00343] In some embodiments, the deletion is about 1 amino acid to about 100 amino acids. In some embodiments, the deletion is at least about 1 amino acid. In some embodiments, the deletion is at most about 100 amino acids. In some embodiments, the deletion is about 1 amino acid to about 5 amino acids, about 1 amino acid to about 10 amino acids, about 1 amino acid to about 20 amino acids, about 1 amino acid to about 30 amino acids, about 1 amino acid to about 40 amino acids, about 1 amino acid to about 50 amino acids, about 1 amino acid to about 60 amino acids, about 1 amino acid to about 70 amino acids, about 1 amino acid to about 80 amino acids, about 1 amino acid to about 90 amino acids, about 1 amino acid to about 100 amino acids, about 5 amino acids to about 10 amino acids, about 5 amino acids to about 20 amino acids, about 5 amino acids to about 30 amino acids, about 5 amino acids to about 40 amino acids, about 5 amino acids to about 50 amino acids, about 5 amino acids to about 60 amino acids, about 5 amino acids to about 70 amino acids, about 5 amino acids to about 80 amino acids, about 5 amino acids to about 90 amino acids, about 5 amino acids to about 100 amino acids, about 10 amino acids to about 20 amino acids, about 10 amino acids to about 30 amino acids, about 10 amino acids to about 40 amino acids, about 10 amino acids to about 50 amino acids, about 10 amino acids to about 60 amino acids, about 10 amino acids to about 70 amino acids, about 10 amino acids to about 80 amino acids, about 10 amino acids to about 90 amino acids, about 10 amino acids to about 100 amino acids, about 20 amino acids to about 30 amino acids, about 20 amino acids to about 40 amino acids, about 20 amino acids to about 50 amino acids, about 20 amino acids to about 60 amino acids, about 20 amino acids to about 70 amino acids, about 20 amino acids to about 80 amino acids, about 20 amino acids to about 90 amino acids, about 20 amino acids to about 100 amino acids, about 30 amino acids to about 40 amino acids, about 30 amino acids to about 50 amino acids, about 30 amino acids to about 60 amino acids, about 30 amino acids to about 70 amino acids, about 30 amino acids to about 80 amino acids, about 30 amino acids to about 90 amino acids, about 30 amino acids to about 100 amino acids, about 40 amino acids to about 50 amino acids, about 40 amino acids to about 60 amino acids, about 40 amino acids to about 70 amino acids, about 40 amino acids to about 80 amino acids, about 40 amino acids to about 90 amino acids, about 40 amino acids to about 100 amino acids, about 50 amino acids to about 60 amino acids, about 50 amino acids to about 70 amino acids, about 50 amino acids to about 80 amino acids, about 50 amino acids to about 90 amino acids, about 50 amino acids to about 100 amino acids, about 60 amino acids to about 70 amino acids, about 60 amino acids to about 80 amino acids, about 60 amino acids to about 90 amino acids, about 60 amino acids to about 100 amino acids, about 70 amino acids to about 80 amino acids, about 70 amino acids to about 90 amino acids, about 70 amino acids to about 100 amino acids, about 80 amino acids to about 90 amino acids, about 80 amino acids to about 100 amino acids, or about 90 amino acids to about 100 amino acids. In some embodiments, the deletion is about 1 amino acid, about 5 amino acids, about 10 amino acids, about 20 amino acids, about 30 amino acids, about 40 amino acids, about 50 amino acids, about 60 amino acids, about 70 amino acids, about 80 amino acids, about 90 amino acids, or about 100 amino acids. [00344] In some embodiments, a polypeptide variant comprising an amino acid sequence that is at least about 75 %, about 80 %, about 85 %, about 90 %, about 91 %, about 92 %, about 93 %, about 94 %, about 95 %, about 96 %, about 97 %, about 98%, or about 99 % identical to the amino acid sequence of a polypeptide disclosed herein. [00345] Molecule derived from antibodies disclosed herein are contemplated. A “molecule derived from an antibody” refers to a functional antigen-binding fragment of an antibody. It is a portion of an antibody heavy and/or light chain polypeptide that retains some or all of the binding activity of the antibody from which the fragment was derived. Non-limiting examples of functional fragments include single-chain Fvs (scFv), Fab fragments, F(ab’) fragments, F(ab)2 fragments, F(ab’)2 fragments, disulfide-linked Fvs (dsFv), Fd fragments, Fv fragments, diabody, triabody, tetrabody, and minibody. Such functional antigen-binding fragment can be found in, for example, Harlow and Lane, Antibodies: A Laboratory Manual (1989); Mol. Biology and Biotechnology: A Comprehensive Desk Reference (Myers, ed., 1995); Huston, et al, 1993, Cell Biophysics 22:189- 224; Pliickthun and Skerra, 1989, Meth. Enzymol. 178:497-515; and Day, Advanced Immunochemistrv (2d ed.1990). 1.5. Compositions and Kits [00346] The antibodies of this invention represent an excellent way for the development of antiviral therapies either alone or in antibody cocktails with additional anti-SARS-CoV-2 virus antibodies for the treatment of human SARS-CoV-2 infections in humans. [00347] In some embodiments, the present invention provides a pharmaceutical composition comprising the antibodies of the present invention described herein formulated together with a pharmaceutically acceptable carrier. The composition may optionally contain one or more additional pharmaceutically active ingredients, such as another antibody or a therapeutic agent. [00348] In some embodiments, the pharmaceutical comprises two or more of the antibody or antigen-binding fragment thereof described herein, such as any combinations of the antibody or antigen-binding fragment thereof comprising a heavy chain and a light chain that comprise the respective amino acid sequences described herein. In some embodiments, the present disclosure provides compositions comprising two or more monoclonal antibodies, or antigen- binding fragments thereof, described herein. In some embodiments, the present disclosure provides compositions comprising 2, 3, or 4 monoclonal antibodies, or antigen-binding fragments thereof, described herein. For example, in some embodiments, the present disclosure provides a composition comprising one of the following combinations of monoclonal antibodies or antigen- binding fragments thereof:
Figure imgf000114_0001
Figure imgf000115_0001
[00349] In some embodiments, the pharmaceutical compositions disclosed herein are for use in treating a SARS-CoV-2 infection in a subject. In some embodiments, the pharmaceutical compositions disclosed herein are for use in the prevention of a SARS-CoV-2 infection in a subject. [00350] The pharmaceutical compositions of the invention also can be administered in a combination therapy with, for example, another immune-stimulatory agent, an antiviral agent, a vaccine, etc. In some embodiments, a composition comprises an antibody of this invention at a concentration of at least 1 mg/ml, 5 mg/ml, 10 mg/ml, 50 mg/ml, 100 mg/ml, 150 mg/ml, 200 mg/ml, 1-300 mg/ml, or 100-300 mg/ml. [00351] In some embodiments, the second therapeutic agent comprises an anti-inflammatory drug or an antiviral compound. In some embodiments, the antiviral compound comprises: a nucleoside analog, a peptoid, an oligopeptide, a polypeptide, a protease inhibitor, a 3C-like protease inhibitor, a papain-like protease inhibitor, or an inhibitor of an RNA dependent RNA polymerase. In some embodiments, the antiviral compound may include: acyclovir, ganciclovir, vidarabine, foscarnet, cidofovir, amantadine, ribavirin, trifluorothymidine, zidovudine, didanosine, zalcitabine, or an interferon. In some embodiments, the interferon is an interferon-Į^ RU^ DQ^ interferon-ȕ^^ [00352] Also within the scope of this disclosure is use of the pharmaceutical composition in the preparation of a medicament for the diagnosis, prophylaxis, treatment, or combination thereof of a condition resulting from a SARS-CoV-2. [00353] The pharmaceutical composition can comprise any number of excipients. Excipients that can be used include carriers, surface-active agents, thickening or emulsifying agents, solid binders, dispersion or suspension aids, solubilizers, colorants, flavoring agents, coatings, disintegrating agents, lubricants, sweeteners, preservatives, isotonic agents, and combinations thereof. The selection and use of suitable excipients is taught in Gennaro, ed., Remington: The Science and Practice of Pharmacy, 20th Ed. (Lippincott Williams & Wilkins 2003), the disclosure of which is incorporated herein by reference. [00354] Preferably, a pharmaceutical composition is suitable for intravenous, intramuscular, subcutaneous, parenteral, spinal, or epidermal administration (e.g., by injection or infusion). Depending on the route of administration, the active compound can be coated in a material to protect it from the action of acids and other natural conditions that may inactivate it. The phrase “parenteral administration” as used herein means modes of administration other than enteral and topical administration, usually by injection, and includes, without limitation, intravenous, intramuscular, intraarterial, intrathecal, intracapsular, intraorbital, intracardiac, intradermal, intraperitoneal, transtracheal, subcutaneous, subcuticular, intraarticular, subcapsular, subarachnoid, intraspinal, epidural and intrasternal injection and infusion. Alternatively, an antibody of the present invention described herein can be administered via a non-parenteral route, such as a topical, epidermal, or mucosal route of administration, e.g., intranasally, orally, vaginally, rectally, sublingually, or topically. [00355] The pharmaceutical compositions of the invention may be prepared in many forms that include tablets, hard or soft gelatin capsules, aqueous solutions, suspensions, liposomes, and other slow-release formulations, such as shaped polymeric gels. An oral dosage form may be formulated such that the antibody is released into the intestine after passing through the stomach. Such formulations are described in U.S. Pat. No.6,306,434 and in the references contained therein. [00356] Oral liquid pharmaceutical compositions may be in the form of, for example, aqueous or oily suspensions, solutions, emulsions, syrups, or elixirs, or may be presented as a dry product for constitution with water or other suitable vehicle before use. Such liquid pharmaceutical compositions may contain conventional additives such as suspending agents, emulsifying agents, non-aqueous vehicles (which may include edible oils), or preservatives. [00357] An antibody can be formulated for parenteral administration (e.g., by injection, for example, bolus injection or continuous infusion) and may be presented in unit dosage form in ampules, prefilled syringes, small volume infusion containers, or multi-dose containers with an added preservative. The pharmaceutical compositions may take such forms as suspensions, solutions, or emulsions in oily or aqueous vehicles, and may contain formulatory agents such as suspending, stabilizing and/or dispersing agents. Pharmaceutical compositions suitable for rectal administration can be prepared as unit dose suppositories. Suitable carriers include saline solution and other materials commonly used in the art. [00358] For administration by inhalation, an antibody can be conveniently delivered from an insufflator, nebulizer, a pressurized pack, or other convenient means of delivering an aerosol spray. Pressurized packs may comprise a suitable propellant such as dichlorodifluoromethane, trichlorofluoromethane, dichlorotetrafluoroethane, carbon dioxide, or other suitable gas. In the case of a pressurized aerosol, the dosage unit may be determined by providing a valve to deliver a metered amount. [00359] Alternatively, for administration by inhalation or insufflation, an antibody may take the form of a dry powder composition, for example, a powder mix of a modulator and a suitable powder base such as lactose or starch. The powder composition may be presented in a unit dosage form in, for example, capsules or cartridges or, e.g., gelatin or blister packs from which the powder may be administered with the aid of an inhalator or insufflator. For intra-nasal administration, an antibody may be administered via a liquid spray, such as via a plastic bottle atomizer. [00360] Pharmaceutical compositions of the invention may also contain other ingredients such as flavorings, colorings, anti-microbial agents, or preservatives. It will be appreciated that the amount of an antibody required for use in treatment will vary not only with the particular carrier selected but also with the route of administration, the nature of the condition being treated, and the age and condition of the patient. Ultimately the attendant health care provider may determine a proper dosage. In addition, a pharmaceutical composition may be formulated as a single unit dosage form. [00361] The pharmaceutical composition of the present invention can be in the form of sterile aqueous solutions or dispersions. It can also be formulated in a microemulsion, liposome, or other ordered structure suitable to high drug concentration. [00362] An antibody of the present invention described herein can be administered as a sustained release formulation, in which case less frequent administration is required. Dosage and frequency vary depending on the half-life of the antibody in the patient. In general, human antibodies show the longest half-life, followed by humanized antibodies, chimeric antibodies, and nonhuman antibodies. The dosage and frequency of administration can vary depending on whether the treatment is prophylactic or therapeutic. In prophylactic applications, a relatively low dosage is administered at relatively infrequent intervals over a long period of time. Some patients continue to receive treatment for the rest of their lives. In therapeutic applications, a relatively high dosage at relatively short intervals is sometimes required until progression of the disease is reduced or terminated, and preferably until the patient shows partial or complete amelioration of symptoms of disease. Thereafter, the patient can be administered a prophylactic regime. [00363] The amount of active ingredient that can be combined with a carrier material to produce a single dosage form will vary depending upon the subject being treated and the particular mode of administration and will generally be that amount of the composition, which produces a therapeutic effect. Generally, out of one hundred percent, this amount will range from about 0.01% to about 99% of active ingredient, preferably from about 0.1% to about 70%, most preferably from about 1% to about 30% of active ingredient in combination with a pharmaceutically acceptable carrier. [00364] Dosage regimens can be adjusted to provide the optimum desired response (e.g., a therapeutic response). For example, a single bolus can be administered, several divided doses can be administered over time, or the dose can be proportionally reduced or increased as indicated by the exigencies of the therapeutic situation. It is especially advantageous to formulate parenteral compositions in dosage unit form for ease of administration and uniformity of dosage. Dosage unit form as used herein refers to physically discrete units suited as unitary dosages for the subjects to be treated; each unit contains a predetermined quantity of active compound calculated to produce the desired therapeutic effect in association with the required pharmaceutical carrier. Alternatively, the antibody can be administered as a sustained release formulation, in which case less frequent administration is required. For administration of the antibody, the dosage ranges from about 0.0001 to 800 mg/kg, and more usually 0.01 to 5 mg/kg, of the host body weight. For example, dosages can be 0.3 mg/kg body weight, 1 mg/kg body weight, 3 mg/kg body weight, 5 mg/kg body weight or 10 mg/kg body weight or within the range of 1-10 mg/kg. An exemplary treatment regime entails administration once per week, once every two weeks, once every three weeks, once every four weeks, once a month, once every 3 months or once every three to 6 months. Preferred dosage regimens for an antibody of the invention include 1 mg/kg body weight or 3 mg/kg body weight via intravenous administration, with the antibody being given using one of the following dosing schedules: (i) every four weeks for six dosages, then every three months; (ii) every three weeks; (iii) 3 mg/kg body weight once followed by 1 mg/kg body weight every three weeks. In some methods, dosage is adjusted to achieve a plasma antibody concentration of about 1-1000 ug /ml, and in some methods, about 25-300 μg /ml. A “therapeutically effective dosage” of an antibody of the invention preferably results in a decrease in seventy of disease symptoms, an increase in frequency and duration of disease symptom-free periods, or a prevention of impairment or disability due to the disease affliction. For example, for the treatment of SARS- CoV-2 infection in a subject, a “therapeutically effective dosage” preferably inhibits SARS-CoV- 2 virus replication or uptake by host cells by at least about 20%, more preferably by at least about 40%, even more preferably by at least about 60%, and still more preferably by at least about 80% relative to untreated subjects. A therapeutically effective amount of a therapeutic compound can neutralize SARS-CoV-2 virus, or otherwise ameliorate symptoms in a subject, which is typically a human or can be another mammal.
[00365] The pharmaceutical composition can be a controlled release formulation, including implants, transdermal patches, and microencapsulated delivery systems. Biodegradable, biocompatible polymers can be used, such as ethylene-vinyl acetate, polyanhydrides, polyglycolic acid, collagen, poly orthoesters, and polylactic acid. See, e.g., Sustained and Controlled Release Drug Delivery Systems, J. R. Robinson, ed., Marcel Dekker, Inc., New York, 1978.
[00366] Therapeutic compositions can be administered via medical devices such as (1) needleless hypodermic injection devices (e.g,, US 5,399,163; 5,383,851; 5,312,335; 5,064,413; 4,941,880; 4,790,824; and 4,596,556); (2) micro-infusion pumps (US 4,487,603); (3) transdermal devices (US 4,486,194); (4) infusion apparati (US 4,447,233 and 4,447,224); and (5) osmotic devices (US 4,439,196 and 4,475,196); the disclosures of which are incorporated herein byreference.
[00367] In some embodiments, the human monoclonal antibodies of the invention described herein can be formulated to ensure proper distribution in vivo. For example, to ensure that the therapeutic compounds of the invention cross the blood-brain barrier, they can be formulated in liposomes, which may additionally comprise targeting moieties to enhance selective transport to specific cells or organs. See, e.g., US 4,522,811; 5,374,548; 5,416,016; and 5,399,331; V.V. Ranade (1989) Clin. Pharmacol. 29:685; Urnezawa et al., (1988) Biochem. Biophys. Res. Cornmun. 153:1038; Bloeman et al., (1995) FEBS Lett. 357: 140; M. Owais et al., (1995) Antimicrob. Agents Chemother. 39:180; Briscoe et al. (1995) Am. Physiol. 1233: 134; Schreier et al., (1994). Biol. Chem. 269:9090; Keinanen and Laukkanen (1994) FEBS Lett. 346: 123; and Killion and Fidler (1994) Immunomethods 4:273. [00368] In some embodiments, the initial dose may be followed by administration of a second or a plurality of subsequent doses of the antibody or antigen-binding fragment thereof in an amount that can be approximately the same or less than that of the initial dose, wherein the subsequent doses are separated by at least 1 day to 3 days; at least one week, at least 2 weeks; at least 3 weeks; at least 4 weeks; at least 5 weeks; at least 6 weeks; at least 7 weeks; at least 8 weeks; at least 9 weeks; at least 10 weeks; at least 12 weeks; or at least 14 weeks. [00369] Various delivery systems are known and can be used to administer the pharmaceutical composition of the invention, e.g., encapsulation in liposomes, microparticles, microcapsules, recombinant cells capable of expressing the mutant viruses, receptor-mediated endocytosis (see, e.g., Wu et al., (1987) J. Biol. Chem. 262:4429-4432). Methods of introduction include, but are not limited to, intradermal, transdermal, intramuscular, intraperitoneal, intravenous, subcutaneous, intranasal, epidural, and oral routes. The composition may be administered by any convenient route, for example by infusion or bolus injection, by absorption through epithelial or mucocutaneous linings (e.g., oral mucosa, rectal and intestinal mucosa, etc.) and may be administered together with other biologically active agents. Administration can be systemic or local. The pharmaceutical composition can also be delivered in a vesicle, in particular, a liposome (see, for example, Langer (1990) Science 249: 1527-1533). [00370] The use of nanoparticles to deliver the antibodies of the present invention is also contemplated herein. Antibody-conjugated nanoparticles may be used both for therapeutic and diagnostic applications. Antibody-conjugated nanoparticles and methods of preparation and use are described in detail by Arruebo, M., et al., 2009 (“Antibody-conjugated nanoparticles for biomedical applications” in J. Nanomat. Volume 2009, Article ID 439389), incorporated herein by reference. Nanoparticles may be developed and conjugated to antibodies contained in pharmaceutical compositions to target cells. Nanoparticles for drug delivery have also been described in, for example, US 8257740, or US 8246995, each incorporated herein in its entirety. [00371] In certain situations, the pharmaceutical composition can be delivered in a controlled release system. In one embodiment, a pump may be used. In another embodiment, polymeric materials can be used. In yet another embodiment, a controlled release system can be placed in proximity of the composition's target, thus requiring only a fraction of the systemic dose. [00372] The injectable preparations may include dosage forms for intravenous, subcutaneous, intracutaneous, intracranial, intraperitoneal, intramuscular injections, drip infusions, etc. These injectable preparations may be prepared by methods publicly known. For example, the injectable preparations may be prepared, e.g., by dissolving, suspending, or emulsifying the antibody or its salt described herein in a sterile aqueous medium or an oily medium conventionally used for injections. As the aqueous medium for injections, there are, for example, physiological saline, an isotonic solution containing glucose and other auxiliary agents, etc., which may be used in combination with an appropriate solubilizing agent such as an alcohol (e.g., ethanol), a polyalcohol (e.g., propylene glycol, polyethylene glycol), a nonionic surfactant [e.g., polysorbate 80, HCO-50 (polyoxyethylene (50 mol) adduct of hydrogenated castor oil)], etc. As the oily medium, there are employed, e.g., sesame oil, soybean oil, etc., which may be used in combination with a solubilizing agent such as benzyl benzoate, benzyl alcohol, etc. The injection thus prepared is preferably filled in an appropriate ampoule. [00373] A pharmaceutical composition of the present invention can be delivered subcutaneously or intravenously with a standard needle and syringe. In addition, with respect to subcutaneous delivery, a pen delivery device readily has applications in delivering a pharmaceutical composition of the present invention. Such a pen delivery device can be reusable or disposable. A reusable pen delivery device generally utilizes a replaceable cartridge that contains a pharmaceutical composition. Once all of the pharmaceutical composition within the cartridge has been administered and the cartridge is empty, the empty cartridge can readily be discarded and replaced with a new cartridge that contains the pharmaceutical composition. The pen delivery device can then be reused. In a disposable pen delivery device, there is no replaceable cartridge. Rather, the disposable pen delivery device comes prefilled with the pharmaceutical composition held in a reservoir within the device. Once the reservoir is emptied of the pharmaceutical composition, the entire device is discarded. [00374] Numerous reusable pens and autoinjector delivery devices have applications in the subcutaneous delivery of a pharmaceutical composition of the present invention. Examples include, but certainly are not limited to AUTOPEN™ (Owen Mumford, Inc., Woodstock, UK), DISETRONIC™ pen (Disetronic Medical Systems, Burghdorf, Switzerland), HUMALOG MIX 75/25™ pen, HUMALOG™ pen, HUMALIN 70/30™ pen (Eli Lilly and Co., Indianapolis, IN), NOVOPEN™ I, II and III (Novo Nordisk, Copenhagen, Denmark), NOVOPEN JUNIOR™ (Novo Nordisk, Copenhagen, Denmark), BD™ pen (Becton Dickinson, Franklin Lakes, NJ), OPTIPEN™, OPTIPEN PRO™, OPTIPEN STARLET™, and OPTICLIK™ (Sanofi-Aventis, Frankfurt, Germany), to name only a few. Examples of disposable pen delivery devices having applications in subcutaneous delivery of a pharmaceutical composition of the present invention include, but certainly are not limited to, the SOLOSTAR™ pen (Sanofi- Aventis), the FLEXPEN™ (Novo Nordisk), and the KWIKPEN™ (Eli Lilly), the SURECLICK™ Autoinjector (Amgen, Thousand Oaks, CA), the PENLET™ (Haselmeier, Stuttgart, Germany), the EPIPEN® (Dey, L.P.) and the HUMIRA™ Pen (Abbott Labs, Abbott Park, IL), to name only a few. [00375] Advantageously, the pharmaceutical compositions for oral or parenteral use described herein are prepared into dosage forms in a unit dose suited to fit a dose of the active ingredients. Such dosage forms in a unit dose include, for example, tablets, pills, capsules, injections (ampoules), suppositories, etc. The amount of the antibody contained is generally about 5 to about 500 mg per dosage form in a unit dose; especially in the form of injection, it is preferred that the antibody is contained in about 5 to about 300 mg and in about 10 to about 300 mg for the other dosage forms. [00376] In another aspect, this disclosure provides a kit comprising a pharmaceutically acceptable dose unit of the antibody or antigen-binding fragment thereof or the pharmaceutical composition as described herein. Also within the scope of this disclosure is a kit for the diagnosis, prognosis, or monitoring of treatment of SARS-CoV-2 in a subject, comprising: the antibody or antigen-binding fragment thereof as described; and a least one detection reagent that binds specifically to the antibody or antigen-binding fragment thereof. [00377] In some embodiments, the kit also includes a container that contains the composition and optionally informational material. The informational material can be descriptive, instructional, marketing, or other material that relates to the methods described herein and/or the use of the agents for therapeutic benefit. In an embodiment, the kit also includes an additional therapeutic agent, as described herein. For example, the kit includes a first container that contains the composition and a second container for the additional therapeutic agent. [00378] The informational material of the kits is not limited in its form. In some embodiments, the informational material can include information about production of the composition, concentration, date of expiration, batch or production site information, and so forth. In one embodiment, the informational material relates to methods of administering the composition, e.g., in a suitable dose, dosage form, or mode of administration (e.g., a dose, dosage form, or mode of administration described herein), to treat a subject in need thereof. In one embodiment, the instructions provide a dosing regimen, dosing schedule, and/or route of administration of the composition or the additional therapeutic agent. The information can be provided in a variety of formats, including printed text, computer-readable material, video recording, audio recording, or information that contains a link or address to substantive material. [00379] The kit can include one or more containers for the composition. In some embodiments, the kit contains separate containers, dividers, or compartments for the composition and informational material. For example, the composition can be contained in a bottle or vial, and the informational material can be contained in a plastic sleeve or packet. In other embodiments, the separate elements of the kit are contained within a single, undivided container. For example, the composition is contained in a bottle or vial that has attached thereto the informational material in the form of a label. In some embodiments, the kit includes a plurality (e.g., a pack) of individual containers, each containing one or more unit dosage forms (e.g., a dosage form described herein) of the agents. [00380] The kit optionally includes a device suitable for administration of the composition or other suitable delivery device. The device can be provided pre-loaded with one or both of the agents or can be empty, but suitable for loading. Such a kit may optionally contain a syringe to allow for injection of the antibody contained within the kit into an animal, such as a human. 1.6. Therapeutic Methods and Applications Methods of Treatment [00381] Provided herein are methods of treating a SARS-CoV-2 infection in a subject, comprising administering to the subject a therapeutically effective amount of the antibodies, antigen-binding fragments thereof, or pharmaceutical compositions thereof disclosed herein. Also provided herein are methods of preventing a SARS-CoV-2 infection in a subject, comprising administering to the subject a therapeutically effective amount of the antibodies, antigen-binding fragments thereof, or pharmaceutical compositions thereof disclosed herein. Also provided herein are methods of neutralizing a SARS-CoV-2 infection in a subject, comprising administering to the subject a therapeutically effective amount of the antibodies, antigen-binding fragments thereof, or pharmaceutical compositions thereof disclosed herein. [00382] In some embodiments, the method described herein may be used in the treatment and/or prevention of SARS-CoV-2. Exemplary SARS-CoV-2 variants include WHO alpha variant, WHO beta variant, WHO gamma variant, WHO delta variant, WHO epsilon variant, WHO Eta variant, WHO iota variant, WHO kappa variant, WHO omicron variant, WHO zeta variant, WHO mu variant, and B.1.617.3. [00383] In some embodiments, the subject may be a neonate, a juvenile, or an adult. In some embodiments, the subject is human. In some embodiments, the subject is non-human primates (e.g., monkeys, baboons, and chimpanzees), mice, rats, bovines, horses, household cats, tigers and other large cats, dogs, pigs, rabbits, goats, deer, sheep, ferrets, gerbils, guinea pigs, hamsters, bats, and birds (e.g., chickens, turkeys, and ducks). A number of these household pets and farm animals are capable of carrying and transmitting SARS-CoV-2 viruses without themselves getting substantially sick or dying, thereby transmitting the disease to humans. Thus, in some embodiments, these animals are treated not because they are suffering from disease, but rather, because they can transmit virus to humans and cause human disease. [00384] In some embodiments, the antibodies, antigen-binding fragments thereof, or pharmaceutical compositions thereof are administered to a subject in order to prevent infection with a SARS-CoV-2 virus. In some embodiments, the antibodies, antigen-binding fragments thereof, or pharmaceutical compositions thereof disclosed herein are administered to a subject prior to exposure to or infection with SARS-CoV-2 viruses. In some embodiments, the antibodies, antigen-binding fragments thereof, or pharmaceutical compositions thereof disclosed herein are administered to a subject after the exposure to or infection with SARS-CoV-2 viruses. The neutralizing of the SARS-CoV-2 virus can be done via (i) inhibiting SARS-CoV-2 virus binding to a target cell; (ii) inhibiting SARS-CoV-2 virus uptake by a target cell; (iii) inhibiting SARS- CoV-2 virus replication; and (iv) inhibiting SARS-CoV-2 virus particles release from infected cells. One skilled in the art possesses the ability to perform any assay to assess neutralization of SARS-CoV-2 virus. [00385] Notably, the neutralizing properties of antibodies may be assessed by a variety of tests, which all may assess the consequences of (i) inhibition of SARS-CoV-2 virus binding to a target cell; (ii) inhibition of SARS-CoV-2 virus uptake by a target cell; (iii) inhibition of SARS-CoV-2 virus replication; and (iv) inhibition of SARS-CoV-2 virus particles release from infected cells. In other words, implementing different tests may lead to the observation of the same consequence, i.e., the loss of infectivity of the SARS-CoV-2 virus. Thus, in one embodiment, the present invention provides a method of neutralizing a SARS-CoV-2 virus in a subject comprising administering to the subject a therapeutically effective amount of the antibody of the present invention described herein. [00386] In some embodiments, the present disclosure provides methods of preventing infection with a SARS-CoV-2 virus in an immunocompromised subject. Immunocompromised subjects include subjects that suffer from an immune deficiency (e.g., a primary or acquired immune deficiency) or autoimmune disease, subjects that have undergone or are currently undergoing treatment with one or more immunosuppressive drugs (e.g., chemotherapy, glucocorticoids, protease inhibitors, immune cell depleting monoclonal antibodies, etc.), subjects that have recently received an organ transplant or hematopoietic stem cell transplant, subjects that have received a CAR-T therapy, and subjects that have undergone or are currently undergoing radiation treatment. Immunosuppressive drugs are known to those in the art. See e.g., Hussain Y, Khan H. Immunosuppressive Drugs. Encyclopedia of Infection and Immunity. 2022:726–40. Vaccines, including vaccines against SARS-CoV-2 infections, are less effective in immunocompromised subjects. Furthermore, some immunocompromised subjects may not be able to receive a SARS- CoV-2 vaccine. The antibodies and antigen-binding fragments thereof provided herein therefore provide a therapeutic option for subjects who cannot receive a SARS-CoV-2 vaccine or who demonstrate reduced efficacy of a SARS-CoV-2 vaccine. [00387] Another aspect of the present invention provides a method of treating a SARS-CoV-2- related disease. Such a method includes therapeutic (following SARS-CoV-2 infection) and prophylactic (prior to SARS-CoV-2 exposure, infection, or pathology). For example, therapeutic and prophylactic methods of treating an individual for a SARS-CoV-2 infection include treatment of an individual having or at risk of having a SARS-CoV-2 infection or pathology, treating an individual with a SARS-CoV-2 infection, and methods of protecting an individual from a SARS- CoV-2 infection, to decrease or reduce the probability of a SARS-CoV-2 infection in an individual, to decrease or reduce susceptibility of an individual to a SARS-CoV-2 infection, or to inhibit or prevent a SARS-CoV-2 infection in an individual, and to decrease, reduce, inhibit or suppress transmission of a SARS-CoV-2 from an infected individual to an uninfected individual. Such methods include administering an antibody of the present invention or a composition comprising the antibody disclosed herein to therapeutically or prophylactically treat (vaccinate or immunize) an individual having or at risk of having a SARS-CoV-2 infection or pathology. Accordingly, methods can treat the SARS-CoV-2 infection or pathology, or provide the individual with protection from infection (e.g., prophylactic protection). [00388] In one embodiment, a method of treating a SARS-CoV-2-related disease comprises administering to an individual in need thereof an antibody or therapeutic composition disclosed herein in an amount sufficient to reduce one or more physiological conditions or symptoms associated with a SARS-CoV-2 infection or pathology, thereby treating the SARS-CoV-2 -related disease. [00389] In one embodiment, an antibody or therapeutic composition disclosed herein is used to treat a SARS-CoV-2-related disease. Use of an antibody or therapeutic composition disclosed herein treats a SARS-CoV-2-related disease by reducing one or more physiological conditions or symptoms associated with a SARS-CoV-2 infection or pathology. In aspects of this embodiment, administration of an antibody or therapeutic composition disclosed herein is in an amount sufficient to reduce one or more physiological conditions or symptoms associated with a SARS- CoV-2 infection or pathology, thereby treating the SARS-CoV-2-based disease. In other aspects of this embodiment, administration of an antibody or therapeutic composition disclosed herein is in an amount sufficient to increase, induce, enhance, augment, promote or stimulate SARS-CoV- 2 clearance or removal; or decrease, reduce, inhibit, suppress, prevent, control, or limit transmission of SARS-CoV-2 to another individual. [00390] In some embodiments, treating refers to the treatment of a disease in a mammal, e.g., in a human, including (a) inhibiting the disease, i.e., arresting disease development or preventing disease progression; (b) relieving the disease, i.e., causing regression of the disease state or relieving one or more symptoms of the disease; and (c) curing the disease, i.e., remission of one or more disease symptoms. In some embodiments, treatment results in an improvement or remediation of the symptoms of the disease. In some embodiments, treatment may refer to a short- term (e.g., temporary and/or acute) and/or a long-term (e.g., sustained) improvement or remediation in one or more disease symptoms. In some embodiments, the improvement is an observable or measurable improvement. In some embodiments, the improvement is an improvement in the general feeling of well-being of the subject. In some embodiments, administration of the pharmaceutical compositions disclosed herein may reduce one or more symptoms of the SARS-COV-2 infection, including but not limited to, death, incidence of emphysema, incidence of pneumonia, shortness of breath, racing heart, fever, cough, sore throat, congestion, muscle or body aches, headaches, fatigue, vomiting, diarrhea, loss of taste or smell, cognitive issues like “brain fog”, memory or attention problems, and Postural Orthostatic Tachycardia Syndrome (POTS). [00391] One or more physiological conditions or symptoms associated with a SARS-CoV-2 infection or pathology will respond to a method of treatment disclosed herein. The symptoms of SARS-CoV-2 infection or pathology vary, depending on the phase of infection. [00392] In some embodiments, the method of neutralizing SARS-CoV-2 in a subject comprises administering to a subject in need thereof a therapeutically effective amount of a first antibody or antigen-binding fragment thereof and a second antibody or antigen-binding fragment thereof of the antibody or antigen-binding fragment, as described herein, wherein the first antibody or antigen-binding fragment thereof and the second antibody or antigen-binding fragment thereof exhibit synergistic activity or a therapeutically effective amount of the pharmaceutical composition described herein. [00393] In some embodiments, the method of preventing or treating a SARS-CoV-2 infection, comprising administering to a subject in need thereof a therapeutically effective amount of a first antibody or antigen-binding fragment thereof and a second antibody or antigen-binding fragment thereof of the antibody or antigen-binding fragment, as described herein, wherein the first antibody or antigen-binding fragment thereof and the second antibody or antigen-binding fragment thereof exhibit synergistic activity or a therapeutically effective amount of the pharmaceutical composition described herein. In some embodiments, the first antibody or antigen-binding fragment thereof is administered before, after, or concurrently with the second antibody or antigen-binding fragment thereof. [00394] In some embodiments, the first antibody or antigen-binding fragment thereof and the second antibody or antigen-binding fragment thereof can be any combinations of the antibody or antigen-binding fragment thereof comprising a heavy chain and a light chain that comprise the respective amino acid sequences described herein. [00395] In some embodiments, the first antibody or antigen-binding fragment thereof comprises a heavy chain variable region and a light chain variable region comprising a HCVR/LCVR amino acid sequence pair of SEQ ID NOs: 1-2, 3-4, 5-6, 7-8, 9-10, 11-12, 13-14, 15-16, 17-18, 19-20, 21-22, 23-24, 25-26, 27-28, 29-30, 31-32, 33-34, 35-36, 37-38, 39-40, 41-42, 43-44, 45-46, 47-48, 49-50, 51-52, 53-54, 55-56, 57-58, 59-60, 61-62, 63-64, 65-66, 67-68, 69-70, 71-72, 73-74, 75-76, 77-78, 79-80, 81-82, 83-84, 85-86, 87-88, 89-90, 91-92, 93-94, 95-96, 97-98, 99-100, 101-102, 103-104, 105-106, 107-108, 109-110, 111-112, 113-114, 115-116, 117-118, 119-120, 121-122, 123-124, 125-126, 127-128, 129-130, 131-132, 133-134, 135-136, 137-138, 139-140, 141-142, 143-144, 145-146, 147-148, 149-150, 151-152, 153-154, 155-156, 157-158, 159-160, 161-162, 163-164, 165-166, 167-168, 169-170, 171-172, 173-174, 175-176, 177-178, 179-180, 181-182, 183-184, 185-186, 187-188, 189-190, 191-192, 193-194, 195-196, 197-198, 199-200, 201-202, 203-204, 205-206, 207-208, 209-210, 211-212, 213-214, 215-216, 217-218, 219-220, 221-222, 223-224, 225-226, 227-228, 229-230, 231-232, 233-234, 235-236, 237-238, 239-240, 241-242, 243-244, 245-246, 247-248, 249-250, 251-252, 253-254, 255-256, 257-258, 259-260, 261-262, 263-264, 265-266, 267-268, 269-270, 271-272, 273-274, 275-276, 277-278, 279-280, 281-282, 283-284, 285-286, 287-288, 289-290, 291-292, 293-294, 295-296, 297-298, 299-300, 301-302, 303-304, 305-306, 307-308, 309-310, 311-312, 313-314, 315-316, 317-318, 319-320, 321-322, 323-324, 325-326, 327-328, 329-330, 331-332, 333-334, 335-336, 337-338, 339-340, 341-342, 343-344, 345-346, 347-348, 349-350, 351-352, 353-354, 355-356, 357-358, 359-360, 361-362, 363-364, 365-366, 367-368, 369-370, 371-372, 373-374, 375-376, 377-378, 379-380, 381-382, 383-384, 385-386, 387-388, 389-390, 391-392, 393-394, 395-396, 397-398, 399-400, 401-402, 403-404, 405-406, 407-408, 409-410, 411-412, 413-414, 415-416, 417-418, 419-420, 421-422, 423-424, 425-426, 427-428, 429-430, 431-432, 433-434, 435-436, 437-438, 439-440, 441-442, 443-444, 445-446, 447-448, 449-450, 451-452, 453-454, 455-456, 457-458, 459-460, 461-462, 463-464, 465-466, 467-468, 469-470, 471-472, 473-474, 475-476, 477-478, 479-480, 481-482, 483-484, 485-486, 487-488, 489-490, 491-492, 493-494, 495-496, 497-498, 499-500, 501-502, 503-504, 505-506, 507-508, 509-510, 511-512, 513-514, 515-516, 517-518, 519-520, 521-522, 523-524, 525-526, 527-528, 529-530, 531-532, 533-534, 535-536, 537-538, 539-540, 541-542, 543-544, 545-546, 547-548, 549-550, 551-552, 553-554, 555-556, 557-558, 559-560, 561-562, 563-564, 565-566, 567-568, 569-570, 571-572, 573-574, 575-576, 577-578, 579-580, 581-582, 583-584, 585-586, 587-588, 589-590, 591-592, 593-594, 595-596, 597-598, 599-600, 601-602, 603-604, 605-606, 607-608, 609-610, 611-612, 613-614, 615-616, 617-618, 619-620, 621-622, 623-624, 625-626, 627-628, 629-630, 631-632, 633-634, 635-636, 637-638, 639-640, 641-642, 643-644, 645-646, 647-648, 649-650, 651-652, 653-654, 655-656, 657-658, 659-660, 661-662, 663-664, 665-666, 667-668, 669-670, 671-672, 673-674, or 675-676. [00396] In some embodiments, the first antibody or antigen-binding fragment thereof comprises a heavy chain variable region and a light chain variable region comprising a HCVR/LCVR amino acid sequence pair of SEQ ID NOs: 25-26, 191-192, 219-220, 395-396, or 397-398. [00397] In some embodiments, the second antibody or antigen-binding fragment thereof comprising a HCVR/LCVR amino acid sequence pair of SEQ ID NOs: 1-2, 3-4, 5-6, 7-8, 9-10, 11-12, 13-14, 15-16, 17-18, 19-20, 21-22, 23-24, 25-26, 27-28, 29-30, 31-32, 33-34, 35-36, 37-38, 39-40, 41-42, 43-44, 45-46, 47-48, 49-50, 51-52, 53-54, 55-56, 57-58, 59-60, 61-62, 63-64, 65-66, 67-68, 69-70, 71-72, 73-74, 75-76, 77-78, 79-80, 81-82, 83-84, 85-86, 87-88, 89-90, 91-92, 93-94, 95-96, 97-98, 99-100, 101-102, 103-104, 105-106, 107-108, 109-110, 111-112, 113-114, 115-116, 117-118, 119-120, 121-122, 123-124, 125-126, 127-128, 129-130, 131-132, 133-134, 135-136, 137-138, 139-140, 141-142, 143-144, 145-146, 147-148, 149-150, 151-152, 153-154, 155-156, 157-158, 159-160, 161-162, 163-164, 165-166, 167-168, 169-170, 171-172, 173-174, 175-176, 177-178, 179-180, 181-182, 183-184, 185-186, 187-188, 189-190, 191-192, 193-194, 195-196, 197-198, 199-200, 201-202, 203-204, 205-206, 207-208, 209-210, 211-212, 213-214, 215-216, 217-218, 219-220, 221-222, 223-224, 225-226, 227-228, 229-230, 231-232, 233-234, 235-236, 237-238, 239-240, 241-242, 243-244, 245-246, 247-248, 249-250, 251-252, 253-254, 255-256, 257-258, 259-260, 261-262, 263-264, 265-266, 267-268, 269-270, 271-272, 273-274, 275-276, 277-278, 279-280, 281-282, 283-284, 285-286, 287-288, 289-290, 291-292, 293-294, 295-296, 297-298, 299-300, 301-302, 303-304, 305-306, 307-308, 309-310, 311-312, 313-314, 315-316, 317-318, 319-320, 321-322, 323-324, 325-326, 327-328, 329-330, 331-332, 333-334, 335-336, 337-338, 339-340, 341-342, 343-344, 345-346, 347-348, 349-350, 351-352, 353-354, 355-356, 357-358, 359-360, 361-362, 363-364, 365-366, 367-368, 369-370, 371-372, 373-374, 375-376, 377-378, 379-380, 381-382, 383-384, 385-386, 387-388, 389-390, 391-392, 393-394, 395-396, 397-398, 399-400, 401-402, 403-404, 405-406, 407-408, 409-410, 411-412, 413-414, 415-416, 417-418, 419-420, 421-422, 423-424, 425-426, 427-428, 429-430, 431-432, 433-434, 435-436, 437-438, 439-440, 441-442, 443-444, 445-446, 447-448, 449-450, 451-452, 453-454, 455-456, 457-458, 459-460, 461-462, 463-464, 465-466, 467-468, 469-470, 471-472, 473-474, 475-476, 477-478, 479-480, 481-482, 483-484, 485-486, 487-488, 489-490, 491-492, 493-494, 495-496, 497-498, 499-500, 501-502, 503-504, 505-506, 507-508, 509-510, 511-512, 513-514, 515-516, 517-518, 519-520, 521-522, 523-524, 525-526, 527-528, 529-530, 531-532, 533-534, 535-536, 537-538, 539-540, 541-542, 543-544, 545-546, 547-548, 549-550, 551-552, 553-554, 555-556, 557-558, 559-560, 561-562, 563-564, 565-566, 567-568, 569-570, 571-572, 573-574, 575-576, 577-578, 579-580, 581-582, 583-584, 585-586, 587-588, 589-590, 591-592, 593-594, 595-596, 597-598, 599-600, 601-602, 603-604, 605-606, 607-608, 609-610, 611-612, 613-614, 615-616, 617-618, 619-620, 621-622, 623-624, 625-626, 627-628, 629-630, 631-632, 633-634, 635-636, 637-638, 639-640, 641-642, 643-644, 645-646, 647-648, 649-650, 651-652, 653-654, 655-656, 657-658, 659-660, 661-662, 663-664, 665-666, 667-668, 669-670, 671-672, 673-674, or 675-676; wherein the first antibody or antigen-binding fragment thereof is different from the second antibody or antigen-binding fragment thereof. [00398] In some embodiments, the second antibody or antigen-binding fragment thereof comprises a HCVR/LCVR amino acid sequence pair of SEQ ID NOs: 25-26, 191-192, 219-220, 395-396, or 397-398, wherein the first antibody or antigen-binding fragment thereof is different from the second antibody or antigen-binding fragment thereof. [00399] In some embodiments, the second therapeutic agent comprises an anti-inflammatory drug or an antiviral compound. In some embodiments, the antiviral compound comprises: a nucleoside analog, a peptoid, an oligopeptide, a polypeptide, a protease inhibitor, a 3C-like protease inhibitor, a papain-like protease inhibitor, or an inhibitor of an RNA dependent RNA polymerase. In some embodiments, the antiviral compound may include: acyclovir, ganciclovir, vidarabine, foscarnet, cidofovir, amantadine, ribavirin, trifluorothymidine, zidovudine, didanosine, zalcitabine, or an interferon. In some embodiments, the interferon is an interferon-Į^ RU^ DQ^ interferon-ȕ^ [00400] In some embodiments, the antibody or antigen-binding fragment thereof is administered before, after, or concurrently with the second therapeutic agent or therapy. In some embodiments, the antibody or antigen-binding fragment thereof is administered to the subject intravenously, subcutaneously, or intraperitoneally. In some embodiments, the antibody or antigen-binding fragment thereof is administered prophylactically or therapeutically. [00401] The antibodies described herein can be used together with one or more of other anti- SARS-CoV-2 virus antibodies to neutralize SARS-CoV-2 virus, thereby treating SARS-CoV-2 infections. [00402] Administration of the antibodies, antigen-binding fragments thereof, or pharmaceutical compositions thereof can occur by infusion (e.g., continuous or bolus), injection, irrigation, inhalation, consumption, electro-osmosis, hemodialysis, iontophoresis, and other methods known in the art. [00403] In some embodiments administration route is intraarterial, intracranial, intradermal, intraduodenal, intrammamary, intrameningeal, intraperitoneal, intrathecal, intratumoral, intravenous, intravitreal, ophthalmic, parenteral, spinal, subcutaneous, ureteral, urethral, vaginal, or intrauterine. In some embodiments, administration route is local or systemic. [00404] The effective amount of the antibodies, antigen-binding fragments thereof, or pharmaceutical compositions thereof administered to a particular subject will depend on a variety of factors, several of which will differ from patient to patient including the disorder being treated and the severity of the disorder; activity of the specific agent(s) employed; the age, body weight, general health, sex and diet of the patient; the timing of administration, route of administration; the duration of the treatment; drugs used in combination; the judgment of the prescribing physician; and like factors known in the medical arts. Dosage amount and interval can be adjusted individually to provide plasma levels of the compound(s) which are sufficient to maintain therapeutic or prophylactic effect. In cases of local administration or selective uptake, such as local topical administration, the effective local concentration of active compound(s) cannot be related to plasma concentration. Skilled artisans will be able to optimize effective local dosages without undue experimentation. [00405] Dosage amounts of the pharmaceutical compositions disclosed herein can typically be in the range of from about 0.0001 mg/kg/day to about 1000 mg/kg/day, but can be higher or lower, depending upon, among other factors, the activity of the compound, its bioavailability, and various factors discussed above. In some embodiments, the dose is from about 0.0001 mg/kg to about 1000 mg/kg of body weight per day. In some embodiments, the dose is from about 0.001 mg/kg to about 1000 mg/kg of body weight per day. In some embodiments, the dose is from about 0.01 mg/kg to about 1000 mg/kg of body weight per day. In some embodiments, the dose is from about 0.1 mg/kg to about 100 mg/kg of body weight per day. In some embodiments, the dose is from about 0.5 mg/kg to about 50 mg/kg of body weight per day. In some embodiments, the dose is from about 1 mg/kg to about 25 mg/kg of body weight per day. In some embodiments, the dose is from about 5 mg/kg to about 15 mg/kg of body weight per day. In some embodiments, the dose is about 1 mg/kg, about 5 mg/kg, about 10 mg/kg, about 15 mg/kg, about 20 mg/kg, about 25 mg/kg, about 30 mg/kg, about 35 mg/kg, about 40 mg/kg, about 45 mg/kg, about 50 mg/kg, about 55 mg/kg, about 60 mg/kg, about 65 mg/kg, about 70 mg/kg, about 75 mg/kg, about 80 mg/kg, about 85 mg/kg, about 90 mg/kg, about 95 mg/kg, or about 100 mg/kg. [00406] The number of administrations of treatment to a subject may vary. In some embodiments, introducing the antibodies, antigen-binding fragments thereof, or pharmaceutical compositions thereof into the subject may be a one-time event. In some embodiments, such treatment may require an on-going series of repeated treatments (e.g., once per day, once per week, or multiple times per day or week). In some embodiments, multiple administrations of the pharmaceutical compositions may be required before an effect is observed. The exact protocols depend upon the disease or condition, the stage of the disease and parameters of the individual subject being treated. Combination Therapies [00407] In some embodiments, the antibodies, antigen-binding fragments thereof, or pharmaceutical compositions thereof disclosed herein are administered in combination with one or more additional therapeutic composition(s). In some embodiments, the additional therapeutic composition is an anti-viral drug. In some embodiments, the additional therapeutic composition is a viral entry inhibitor. In some embodiments, the additional therapeutic composition is a viral attachment inhibitor. [00408] In some embodiments, the antibodies, antigen-binding fragments thereof, or pharmaceutical compositions thereof disclosed herein and the additional therapeutic composition(s) are administered simultaneously. In some embodiments, the antibodies, antigen-binding fragments thereof, or pharmaceutical compositions thereof disclosed herein is administered before the additional therapeutic composition(s). In some embodiments, the antibodies, antigen-binding fragments thereof, or pharmaceutical compositions thereof disclosed herein is administered after the additional therapeutic composition(s). [00409] In some embodiments, administration of the antibodies, antigen-binding fragments thereof, or pharmaceutical compositions thereof disclosed herein in combination with the additional therapeutic composition(s) results in an enhanced therapeutic effect in a subject infected with SARS-CoV-2 viruses than is observed by treatment with either the antibodies, antigen- binding fragments thereof, or pharmaceutical compositions thereof disclosed herein or the additional therapeutic composition(s) alone. [00410] Combination therapies may include an anti- SARS-CoV-2 antibody of the invention and any additional therapeutic agent that may be advantageously combined with an antibody of the invention or with a biologically active fragment of an antibody of the invention. The antibodies of the present invention may be combined synergistically with one or more drugs or therapy used to treat a disease or disorder associated with a viral infection, such as a SARS-CoV-2 infection. In some embodiments, the antibodies of the invention may be combined with a second therapeutic agent to ameliorate one or more symptoms of said disease. In some embodiments, the antibodies of the invention may be combined with a second antibody to provide synergistic activity in ameliorating one or more symptoms of said disease. In some embodiments, the first antibody or antigen-binding fragment thereof is administered before, after, or concurrently with the second antibody or antigen-binding fragment thereof. [00411] For example, the antibody described herein can be used in various detection methods for use in, e.g., monitoring the progression of a SARS-CoV-2 infection; monitoring patient response to treatment for such an infection, etc. The present disclosure provides methods of detecting a neuraminidase polypeptide in a biological sample obtained from an individual. The methods generally involve: a) contacting the biological sample with a subject anti- neuraminidase antibody; and b) detecting binding, if any, of the antibody to an epitope present in the sample. In some instances, the antibody comprises a detectable label. The level of neuraminidase polypeptide detected in the biological sample can provide an indication of the stage, degree, or severity of a SARS-CoV-2 infection. The level of the neuraminidase polypeptide detected in the biological sample can provide an indication of the individual's response to treatment for a SARS-CoV-2 infection. [00412] In some embodiments, the second therapeutic agent is another antibody to a SARS- COV-2 protein or a fragment thereof. It is contemplated herein to use a combination (“cocktail”) of antibodies with broad neutralization or inhibitory activity against SARS-COV-2. In some embodiments, non-competing antibodies may be combined and administered to a subject in need thereof. In some embodiments, the antibodies comprising the combination bind to distinct non- overlapping epitopes on the protein. In some embodiments, the second antibody may possess a longer half-life in human serum. [00413] As used herein, the term “in combination with” means that additional therapeutically active component(s) may be administered prior to, concurrent with, or after the administration of the anti-SARS-COV-2 antibody of the present invention. The term “in combination with” also includes sequential or concomitant administration of an anti-SARS-COV-2 antibody and a second therapeutic agent. [00414] The additional therapeutically active component(s) may be administered to a subject prior to administration of an anti-SARS-COV-2 antibody of the present invention. For example, a first component may be deemed to be administered “prior to” a second component if the first component is administered 1 week before, 72 hours before, 60 hours before, 48 hours before, 36 hours before, 24 hours before, 12 hours before, 6 hours before, 5 hours before, 4 hours before, 3 hours before, 2 hours before, 1 hour before, 30 minutes before, 15 minutes before, 10 minutes before, 5 minutes before, or less than 1 minute before administration of the second component. In other embodiments, the additional therapeutically active component(s) may be administered to a subject after administration of an anti-SARS-COV-2 antibody of the present invention. For example, a first component may be deemed to be administered “after” a second component if the first component is administered 1 minute after, 5 minutes after, 10 minutes after, 15 minutes after, 30 minutes after, 1 hour after, 2 hours after, 3 hours after, 4 hours after, 5 hours after, 6 hours after, 12 hours after, 24 hours after, 36 hours after, 48 hours after, 60 hours after, 72 hours after administration of the second component. In yet other embodiments, the additional therapeutically active component(s) may be administered to a subject concurrent with administration of an anti- SARS-COV-2 antibody of the present invention. “Concurrent” administration, for purposes of the present invention, includes, e.g., administration of an anti-SARS-COV-2 antibody and an additional therapeutically active component to a subject in a single dosage form, or in separate dosage forms administered to the subject within about 30 minutes or less of each other. If administered in separate dosage forms, each dosage form may be administered via the same route (e.g., both the anti-SARS-COV-2 antibody and the additional therapeutically active component may be administered intravenously, etc.); alternatively, each dosage form may be administered via a different route (e.g., the anti-SARS-COV-2 antibody may be administered intravenously, and the additional therapeutically active component may be administered orally). In any event, administering the components in a single dosage form, in separate dosage forms by the same route, or in separate dosage forms by different routes are all considered “concurrent administration,” for purposes of the present disclosure. For purposes of the present disclosure, administration of an anti-SARS-COV-2 antibody “prior to,” “concurrent with,” or “after” (as those terms are defined hereinabove) administration of an additional therapeutically active component is considered administration of an anti-SARS-COV-2 antibody “in combination with” an additional therapeutically active component. [00415] The present invention includes pharmaceutical compositions in which an anti-SARS- COV-2 antibody of the present invention is co-formulated with one or more of the additional therapeutically active component(s) as described elsewhere herein. Administration Regimens [00416] According to certain embodiments, a single dose of an anti-SARS-COV-2 antibody of the invention (or a pharmaceutical composition comprising a combination of an anti-SARS-COV- 2 antibody and any of the additional therapeutically active agents mentioned herein) may be administered to a subject in need thereof. According to certain embodiments of the present invention, multiple doses of an anti-SARS-COV-2 antibody (or a pharmaceutical composition comprising a combination of an anti-SARS-COV-2 antibody and any of the additional therapeutically active agents mentioned herein) may be administered to a subject over a defined time course. The methods according to this aspect of the invention comprise sequentially administering to a subject multiple doses of an anti-SARS-COV-2 antibody of the invention. As used herein, “sequentially administering” means that each dose of anti- SARS-COV-2 antibody is administered to the subject at a different point in time, e.g., on different days separated by a predetermined interval (e.g., hours, days, weeks, or months). The present invention includes methods that comprise sequentially administering to the patient a single initial dose of an anti- SARS-COV-2 antibody, followed by one or more secondary doses of the anti-SARS-COV-2 antibody, and optionally followed by one or more tertiary doses of the anti-SARS-COV-2 antibody. [00417] The terms “initial dose,” “secondary doses,” and “tertiary doses,” refer to the temporal sequence of administration of the anti-SARS-COV-2 antibody of the invention. Thus, the “initial dose” is the dose, which is administered at the beginning of the treatment regimen (also referred to as the “baseline dose”); the “secondary doses” are the doses, which are administered after the initial dose; and the “tertiary doses” are the doses which are administered after the secondary doses. The initial, secondary, and tertiary doses may all contain the same amount of anti-SARS-COV-2 antibody, but generally may differ from one another in terms of frequency of administration. In some embodiments, however, the amount of anti-SARS-COV-2 antibody contained in the initial, secondary and/or tertiary doses varies from one another (e.g., adjusted up or down as appropriate) during the course of treatment. In some embodiments, two or more (e.g., 2, 3, 4, or 5) doses are administered at the beginning of the treatment regimen as “loading doses” followed by subsequent doses that are administered on a less frequent basis (e.g., “maintenance doses”). [00418] In certain exemplary embodiments of the present invention, each secondary and/or tertiary dose is administered 1 to 48 hours (e.g., 1, 1 ½, 2, 2½, 3, 3½, 4, 4½, 5, 5½, 6, 6½, 7, 7½, 8, 8½, 9, 9½, 10, 10½, 11, 11 ½, 12, 12½, 13, 13½, 14, 14½, 15, 15½, 16, 16½, 17, 17½, 18, 18½, 19, 19½, 20, 20½, 21, 21 ½, 22, 22½, 23, 23 ½, 24, 24½, 25, 25 ½, 26, 26½, or more) after the immediately preceding dose. The phrase “the immediately preceding dose,” as used herein, means, in a sequence of multiple administrations, the dose of anti-SARS-COV-2 antibody, which is administered to a patient prior to the administration of the very next dose in the sequence with no intervening doses. [00419] The methods, according to this aspect of the invention, may comprise administering to a patient any number of secondary and/or tertiary doses of an anti-SARS-COV-2 antibody. For example, In some embodiments, only a single secondary dose is administered to the patient. In other embodiments, two or more (e.g., 2, 3, 4, 5, 6, 7, 8, or more) secondary doses are administered to the patient. Likewise, In some embodiments, only a single tertiary dose is administered to the patient. In other embodiments, two or more (e.g., 2, 3, 4, 5, 6, 7, 8, or more) tertiary doses are administered to the patient. [00420] In some embodiments of the invention, the frequency at which the secondary and/or tertiary doses are administered to a patient can vary over the course of the treatment regimen. The frequency of administration may also be adjusted during the course of treatment by a physician depending on the needs of the individual patient following clinical examination. Diagnostic Uses of the Antibodies [00421] The anti-SARS-COV-2 antibodies of the present invention may be used to detect and/or measure SARS-COV-2 in a sample, e.g., for diagnostic purposes. Some embodiments contemplate the use of one or more antibodies of the present invention in assays to detect a SARS-COV-2- associated disease or disorder. Exemplary diagnostic assays for SARS-COV-2 may comprise, e.g., contacting a sample obtained from a patient with an anti-SARS-COV-2 antibody of this disclosure, wherein the anti-SARS-COV-2 antibody is labeled with a detectable label or reporter molecule or used as a capture ligand to selectively isolate SARS-COV-2 from patient samples. Alternatively, an unlabeled anti-SARS-COV-2 antibody can be used in diagnostic applications in combination with a secondary antibody, which is itself detectably labeled. The detectable label or reporter molecule can be a radioisotope, such as H, C, P, S, or I: a fluorescent or chemiluminescent moiety such as fluorescein isothiocyanate, or rhodamine; or an enzyme such as alkaline phosphatase, [3- galactosidase, horseradish peroxidase, or luciferase. Specific exemplary assays that can be used to detect or measure SARS-COV-2 in a sample include enzyme- linked immunosorbent assay (ELISA), radioimmunoassay (RIA), and fluorescence-activated cell sorting (FACS).
[00422] In another aspect, this disclosure further provides a method for detecting the presence of SARS CoV-2 in a sample comprising the steps of: (i) contacting a sample with the antibody or antigen-binding fragment thereof described herein; and (ii) determining binding of the antibody or antigen-binding fragment to one or more SARS CoV-2 antigens, wherein binding of the antibody to the one or more SARS CoV-2 antigens is indicative of the presence of SARS CoV-2 in the sample.
[00423] In some embodiments, the SARS-CoV-2 antigen comprises the receptor binding domain (RBD) of the S polypeptide.
[00424] In some embodiments, the SARS-CoV-2 antigen comprises the N-terminal domain (NTD) of the S polypeptide. In some embodiments, the SARS-CoV-2 antigen comprises the supersite b-hairpin (residues 152-158), the |38-strand (residue 97-102), N4-loop (residues 178-188), and/or N-linked glycans at positions N122 and N149 of the S polypeptide. In some embodiments, the SARS-CoV-2 antigen comprises residues 27-32, 57-60, 210-218, and/or 286-303 of the S polypeptide. In some embodiments, the SARS-CoV-2 antigen comprises residues 600-606 of the S polypeptide.
[00425] In some embodiments, the antibody or antigen-binding fragment thereof is conjugated to a label. In some embodiments, the step of detecting comprises contacting a secondary' antibody with the antibody or antigen-binding fragment thereof and wherein the secondary/ antibody comprises a label. In some embodiments, the label includes a fluorescent label, a chemiluminescent label, a radiolabel, and an enzyme.
[00426] In some embodiments, the step of detecting comprises detecting fluorescence or chemiluminescence. In some embodiments, the step of detecting comprises a competitive binding assay or ELISA.
[00427] In some embodiments, the method further comprises binding the sample to a solid support. In some embodiments, the solid support includes microparticles, microbeads, magnetic beads, and an affinity purification column. [00428] Samples that can be used in SARS-COV-2 diagnostic assays according to the present invention include any tissue or fluid sample obtainable from a patient, which contains detectable quantities of either SARS-COV-2 protein, or fragments thereof, under normal or pathological conditions. Generally, levels of SARS-COV-2 protein in a particular sample obtained from a healthy patient (e.g., a patient not afflicted with a disease associated with SARS-COV-2) will be measured to initially establish a baseline, or standard, level of SARS-COV-2. This baseline level of SARS-COV-2 can then be compared against the levels of SARS-COV-2 measured in samples obtained from individuals suspected of having a SARS-COV-2-associated condition or symptoms associated with such condition. [00429] The antibodies specific for SARS-COV-2 protein may contain no additional labels or moieties, or they may contain an N-terminal or C-terminal label or moiety. In one embodiment, the label or moiety is biotin. In a binding assay, the location of a label (if any) may determine the orientation of the peptide relative to the surface upon which the peptide is bound. For example, if a surface is coated with avidin, a peptide containing a N-terminal biotin will be oriented such that the C-terminal portion of the peptide will be distal to the surface. 2. FURTHER NUMBERED EMBODIMENTS [00430] Further numbered embodiments of the present disclosure are as follows: [00431] Embodiment 1. An isolated anti-SARS-CoV-2 antibody or antigen-binding fragment thereof that binds specifically to a SARS-CoV-2 antigen. [00432] Embodiment 2. The antibody or antigen-binding fragment thereof of Embodiment 1, wherein the SARS-CoV-2 antigen comprises a spike polypeptide, and preferably wherein the SARS-CoV-2 antigen comprises a spike polypeptide of a human or an animal SARS-CoV-2. [00433] Embodiment 3. The antibody or antigen-binding fragment thereof of any one of the preceding Embodiments, wherein the SARS-CoV-2 antigen comprises the receptor binding domain (RBD) or the N-terminal domain (NTD) of the spike polypeptide. [00434] Embodiment 4. The antibody or antigen-binding fragment thereof of any one of the preceding Embodiments, wherein: (a) the SARS-CoV-2 antigen comprises the supersite b-hairpin (residues 152-^^^^^^WKH^ȕ^-strand (residue 97-102), N4-loop (residues 178-188), and/or N-linked glycans at positions N122 and N149 of the spike polypeptide; or (b) wherein the SARS-CoV-2 antigen comprises residues 27-32, 57-60, 210-218, and/or 286-303 of the spike polypeptide, and preferably wherein the SARS-CoV-2 antigen comprises residues 600-606 of the spike polypeptide. [00435] Embodiment 5. The antibody or antigen-binding fragment thereof of any one of the preceding Embodiments, comprising: a. three heavy chain complementarity determining regions (HCDRs) (HCDR1, HCDR2, and HCDR3) of a heavy chain variable region (HCVR) having the amino acid sequence of SEQ ID NO: 25; and three light chain CDRs (LCDR1, LCDR2, and LCDR3) of a light chain variable region (LCVR) having the amino acid sequence of SEQ ID NO: 26; b. HCDR1, HCDR2, and HCDR3 of a HCVR having the amino acid sequence of SEQ ID NO: 191; and LCDR1, LCDR2, and LCDR3 of a LCVR having the amino acid sequence of SEQ ID NO: 192; c. HCDR1, HCDR2, and HCDR3 of a HCVR having the amino acid sequence of SEQ ID NO: 219; and LCDR1, LCDR2, and LCDR3 of a LCVR having the amino acid sequence of SEQ ID NO: 220; d. HCDR1, HCDR2, and HCDR3 of a HCVR having the amino acid sequence of SEQ ID NO: 395; and LCDR1, LCDR2, and LCDR3 of a LCVR having the amino acid sequence of SEQ ID NO: 396; or e. HCDR1, HCDR2, and HCDR3 of a HCVR having the amino acid sequence of SEQ ID NO: 397; and LCDR1, LCDR2, and LCDR3 of a LCVR having the amino acid sequence of SEQ ID NO: 398. [00436] Embodiment 6. The antibody or antigen-binding fragment thereof of any one of the preceding Embodiments, comprising: a. a HCVR comprising an amino acid sequence with at least 75% identity to the amino acid sequence of SEQ ID NO: 25 or comprising the amino acid sequence of SEQ ID NO: 25; and a LCVR comprising an amino acid sequence with at least 75% identity to the amino acid sequence of SEQ ID NO: 26 or comprising the amino acid sequence of SEQ ID NO: 26; b. a HCVR comprising an amino acid sequence with at least 75% identity to the amino acid sequence of SEQ ID NO: 191 or comprising the amino acid sequence of SEQ ID NO: 191; and a LCVR comprising an amino acid sequence with at least 75% identity to the amino acid sequence of SEQ ID NO: 192 or comprising the amino acid sequence of SEQ ID NO: 192; c. a HCVR comprising an amino acid sequence with at least 75% identity to the amino acid sequence of SEQ ID NO: 219 or comprising the amino acid sequence of SEQ ID NO: 219; and a LCVR comprising an amino acid sequence with at least 75% identity to the amino acid sequence of SEQ ID NO: 220 or comprising the amino acid sequence of SEQ ID NO: 220; d. a HCVR comprising an amino acid sequence with at least 75% identity to the amino acid sequence of SEQ ID NO: 395 or comprising the amino acid sequence of SEQ ID NO: 395; and a LCVR comprising an amino acid sequence with at least 75% identity to the amino acid sequence of SEQ ID NO; 396 or comprising the amino acid sequence of SEQ ID NO: 396; or e. a HCVR comprising an amino acid sequence with at least 75% identity to the amino acid sequence of SEQ ID NO: 397 or comprising the amino acid sequence of SEQ ID NO: 397; and a LCVR comprising an amino acid sequence with at least 75% identity to the amino acid sequence of SEQ ID NO; 398 or comprising the amino acid sequence of SEQ ID NO: 398. [00437] Embodiment 7. The antibody or antigen-binding fragment thereof of any one of the preceding Embodiments, comprising a HCVR and a LCVR that comprise a HCVR/LCVR amino acid sequence pair of SEQ ID NOs: 25-26, 191-192, 219-220, 395-396, or 397-398. [00438] Embodiment 8. The antibody or antigen-binding fragment thereof of any one of the preceding Embodiments, comprising a HCVR and a LCVR that comprise a HCVR/LCVR amino acid sequence pair with at least 75% identity to the amino acid sequence of SEQ ID NOs: 25-26, 191-192, 219-220, 395-396, or 397-398. [00439] Embodiment 9. The antibody or antigen-binding fragment thereof of any one of the preceding Embodiments, wherein the antibody or antigen-binding fragment thereof is capable of neutralizing a plurality of SARS-CoV-2 strains. [00440] Embodiment 10. The antibody or antigen-binding fragment thereof of any one of the preceding Embodiments, wherein the antibody or antigen-binding fragment thereof is a multivalent antibody comprising (a) a first target binding site that specifically binds to an epitope within the spike polypeptide, and (b) a second target binding site that binds to an epitope on a different epitope on the spike polypeptide or a different molecule. [00441] Embodiment 11. The antibody or antigen-binding fragment thereof of Embodiment 10, wherein the multivalent antibody is a bivalent or bispecific antibody. [00442] Embodiment 12. The antibody or the antigen-binding fragment thereof of any one of the preceding Embodiments, further comprising a variant Fc constant region. [00443] Embodiment 13. The antibody or antigen-binding fragment thereof of any one of the preceding Embodiments, wherein the antibody is a monoclonal antibody. [00444] Embodiment 14. The antibody or antigen-binding fragment thereof of any one of the preceding Embodiments, wherein the antibody or antigen-binding fragment thereof is a chimeric antibody, a humanized antibody, or humanized monoclonal antibody. [00445] Embodiment 15. The antibody or antigen-binding fragment thereof of any one of the preceding Embodiments, wherein the antibody or antigen-binding fragment thereof is a single- chain antibody, Fab or Fab2 fragment. [00446] Embodiment 16. The antibody or antigen-binding fragment thereof of any one of the preceding Embodiments, wherein the antibody or antigen-binding fragment thereof is detectably labeled or conjugated to a toxin, a therapeutic agent, a polymer, a receptor, an enzyme, or a receptor ligand, preferably wherein the polymer is polyethylene glycol (PEG). [00447] Embodiment 17. A pharmaceutical composition comprising the antibody or antigen- binding fragment thereof of any one of the preceding Embodiments and optionally a pharmaceutically acceptable carrier or excipient. [00448] Embodiment 18. The pharmaceutical composition of Embodiment 16, wherein the pharmaceutical comprises two or more of the antibody or antigen-binding fragment thereof of any one of Embodiments 1 to 16. [00449] Embodiment 19. The pharmaceutical composition of Embodiment 18, wherein the two or more of the antibody or antigen-binding fragment thereof comprise: a. a first antibody or antigen-binding fragment thereof comprising a HCVR/LCVR amino acid sequence pair of SEQ ID NOs: 25-26, 191-192, 219-220, 395-396, or 397-398, b. wherein the first antibody or antigen-binding fragment thereof is different from the second antibody or antigen-binding fragment thereof. [00450] Embodiment 20. The pharmaceutical composition of Embodiment 18, wherein the two or more of the antibody or antigen-binding fragment thereof comprise: a. a first antibody or antigen-binding fragment thereof comprising a HCVR/LCVR amino acid sequence pair with at least 75% identity to the amino acid sequence of SEQ ID NOs: 25-26, 191-192, 219-220, 395-396, or 397-398, b. wherein the first antibody or antigen-binding fragment thereof is different from the second antibody or antigen-binding fragment thereof. [00451] Embodiment 21. The pharmaceutical composition of any one of Embodiments 17 to 20, further comprising a second therapeutic agent, preferably wherein the second therapeutic agent comprises an anti-inflammatory drug or an antiviral compound. [00452] Embodiment 22. The pharmaceutical composition of Embodiment 21, wherein: (a) the antiviral compound comprises: a nucleoside analog, a peptoid, an oligopeptide, a polypeptide, a protease inhibitor, a 3C-like protease inhibitor, a papain-like protease inhibitor, or an inhibitor of an RNA dependent RNA polymerase; or (b) the antiviral compound is selected from: acyclovir, ganciclovir, vidarabine, foscarnet, cidofovir, amantadine, ribavirin, trifluorothymidine, zidovudine, didanosine, zalcitabine, and an interferon, and preferably wherein the interferon is an interferon-Į^ or an interferon-ȕ^ [00453] Embodiment 23. Use of the antibody or antigen-binding fragment thereof of any one of Embodiments 1 to 16 or the pharmaceutical composition of any one of Embodiments 17 to 22 in the manufacture of a medicament for the diagnosis, prophylaxis, treatment, or combination thereof of a condition resulting from a SARS-CoV-2. [00454] Embodiment 24. A nucleic acid molecule encoding a polypeptide chain of the antibody or antigen-binding fragment thereof of any one of Embodiments 1 to 16. [00455] Embodiment 25. A vector comprising the nucleic acid molecule of Embodiment 24. [00456] Embodiment 26. A cultured host cell comprising the vector of Embodiment 25. [00457] Embodiment 27. A method of preparing an antibody or antigen-binding fragment thereof, comprising: a. obtaining the cultured host cell of Embodiment 26; b. culturing the cultured host cell in a medium under conditions permitting expression of a polypeptide encoded by the vector and assembling of an antibody or fragment thereof; and c. purifying the antibody or fragment from the cultured cell or the medium of the cell. [00458] Embodiment 28. A kit comprising a pharmaceutically acceptable dose unit of the antibody or antigen-binding fragment thereof of any one of Embodiments 1 to 16 or the pharmaceutical composition of any one of Embodiments 17 to 22. [00459] Embodiment 29. A kit for the diagnosis, prognosis or monitoring treatment of SARS- CoV-2 in a subject, comprising: the antibody or antigen-binding fragment thereof of any one of Embodiments 1 to 16; and a least one detection reagent that binds specifically to the antibody or antigen-binding fragment thereof. [00460] Embodiment 30. A method of neutralizing SARS-CoV-2 in a subject, comprising administering to a subject in need thereof a therapeutically effective amount of the antibody or antigen-binding fragment thereof of any one of Embodiments 1 to 16 or a therapeutically effective amount of the pharmaceutical composition of any one of Embodiments 17 to 22. [00461] Embodiment 31. A method of preventing or treating a SARS-CoV-2 infection, comprising administering to a subject in need thereof a therapeutically effective amount of the antibody or antigen-binding fragment thereof of any one of Embodiments 1 to 16 or a therapeutically effective amount of the pharmaceutical composition of any one of Embodiments 17 to 22. [00462] Embodiment 32. A method of neutralizing SARS-CoV-2 in a subject, comprising administering to a subject in need thereof a therapeutically effective amount of a first antibody or antigen-binding fragment thereof and a second antibody or antigen-binding fragment thereof of any one of Embodiments 1 to 16 or a therapeutically effective amount of the pharmaceutical composition of any one of Embodiments 17 to 22, wherein the first antibody or antigen-binding fragment thereof and the second antibody or antigen-binding fragment thereof exhibit synergistic activity. [00463] Embodiment 33. A method of preventing or treating a SARS-CoV-2 infection, comprising administering to a subject in need thereof a therapeutically effective amount of a first antibody or antigen-binding fragment thereof of any one of Embodiments 1 to 16 and a second antibody or antigen-binding fragment thereof of any one of Embodiments 1 to 16 or a therapeutically effective amount of the pharmaceutical composition of any one of Embodiments 17 to 22, wherein the first antibody or antigen-binding fragment thereof and the second antibody or antigen-binding fragment thereof exhibit synergistic activity. [00464] Embodiment 34. The method of Embodiment 30 or 31, further comprising administering to the subject a therapeutically effective amount of a second therapeutic agent or therapy. [00465] Embodiment 35. The method of Embodiment 34, wherein the antibody or antigen- binding fragment thereof comprises a heavy chain variable region and a light chain variable region comprising a HCVR/LCVR amino acid sequence pair of SEQ ID NOs: 25-26, 191-192, 219-220, 395-396, or 397-398, wherein the antibody or antigen-binding fragment thereof is different from the second therapeutic agent or therapy. [00466] Embodiment 36. The method of Embodiment 34, wherein the antibody or antigen- binding fragment thereof comprises a heavy chain variable region and a light chain variable region comprising a HCVR/LCVR amino acid sequence pair with at least 75% identity to the amino acid sequence of SEQ ID NOs: 25-26, 191-192, 219-220, 395-396, or 397-398, wherein the antibody or antigen-binding fragment thereof is different from the second therapeutic agent or therapy. [00467] Embodiment 37. The method of Embodiment 34 or 35, wherein the antibody or antigen- binding fragment thereof is administered before, after, or concurrently with the second therapeutic agent or therapy. [00468] Embodiment 38. The method of any one of Embodiments 34 to 37, wherein the second therapeutic agent comprises an anti-inflammatory drug or an antiviral compound. [00469] Embodiment 39. The method of Embodiment 38, wherein: (a) the antiviral compound comprises: a nucleoside analog, a peptoid, an oligopeptide, a polypeptide, a protease inhibitor, a 3C-like protease inhibitor, a papain-like protease inhibitor, or an inhibitor of an RNA dependent RNA polymerase; or (b) the antiviral compound is selected from: acyclovir, ganciclovir, vidarabine, foscarnet, cidofovir, amantadine, ribavirin, trifluorothymidine, zidovudine, didanosine, zalcitabine, and an interferon, and preferably wherein the interferon is an interferon-Į^ RU^ DQ^ interferon-ȕ^ [00470] Embodiment 40. The method of Embodiment 32 or 33, wherein the first antibody or antigen-binding fragment thereof comprises a heavy chain variable region and a light chain variable region comprising a HCVR/LCVR amino acid sequence pair of SEQ ID NOs: 25-26, 191- 192, 219-220, 395-396, or 397-398, wherein the first antibody or antigen-binding fragment thereof is different from the second antibody or antigen-binding fragment thereof. [00471] Embodiment 41. The method of Embodiment 32 or 33, wherein the first antibody or antigen-binding fragment thereof comprises a heavy chain variable region and a light chain variable region comprising a HCVR/LCVR amino acid sequence pair with at least 75% identity to the amino acid sequence of SEQ ID NOs: 25-26, 191-192, 219-220, 395-396, or 397-398, wherein the first antibody or antigen-binding fragment thereof is different from the second antibody or antigen-binding fragment thereof. [00472] Embodiment 42. The method of any one of Embodiments 32,33 and 40, wherein the first antibody or antigen-binding fragment thereof is administered before, after, or concurrently with the second antibody or antigen-binding fragment thereof. [00473] Embodiment 43. The method of any one of Embodiments 30 to 42, wherein the antibody or antigen-binding fragment thereof is administered to the subject intravenously, subcutaneously, or intraperitoneally. [00474] Embodiment 44. The method of Embodiment 32 or 33, wherein the antibody or antigen- binding fragment thereof is administered prophylactically or therapeutically. [00475] Embodiment 45. A method for detecting the presence of SARS CoV-2 in a sample comprising the steps of: a. contacting a sample with the antibody or antigen-binding fragment thereof of any one of Embodiments 1 to 16; and b. determining binding of the antibody or antigen-binding fragment to one or more SARS CoV-2 antigens, c. wherein binding of the antibody to the one or more SARS CoV-2 antigens is indicative of the presence of SARS CoV-2 in the sample. [00476] Embodiment 46. The method of Embodiment 45, wherein the SARS-CoV-2 antigen comprises a spike polypeptide of a human or an animal SARS-CoV-2, and preferably wherein the SARS-CoV-2 antigen comprises the receptor binding domain (RBD) or the N-terminal domain (NTD) of the S polypeptide. [00477] Embodiment 47. The method of any one of Embodiments 45 to 46, wherein the antibody or antigen-binding fragment thereof is conjugated to a label, preferably wherein the label is selected from a fluorescent label, a chemiluminescent label, a radiolabel, and an enzyme. [00478] Embodiment 48. The method of any one of Embodiments 45 to 47, further comprising contacting a secondary antibody with the antibody or antigen-binding fragment thereof, wherein the secondary antibody comprises a label. [00479] Embodiment 49. The method of Embodiment 48, further comprising detecting fluorescence or chemiluminescence of the label, preferably wherein the step of detecting comprises a competitive binding assay or ELISA. [00480] Embodiment 50. The method of any one of Embodiments 45 to 49, wherein the sample is a blood sample. [00481] Embodiment 51. The method of any one of Embodiment 45 to 50, further comprising binding the sample to a solid support, preferably wherein the solid support is selected from microparticles, microbeads, magnetic beads, and an affinity purification column. EXAMPLES [00482] The following is a description of various methods and materials used in the studies. They are put forth so as to provide those of ordinary skill in the art with a complete disclosure and description of how to make and use the present invention, and are not intended to limit the scope of what the inventors regard as their invention, nor are they intended to represent that the experiments below were performed and are all of the experiments that may be performed. It is to be understood that exemplary descriptions written in the present tense were not necessarily performed, but rather that the descriptions can be performed to generate the data and the like associated with the teachings of the present invention. Efforts have been made to ensure accuracy with respect to numbers used (e.g., amounts, percentages, etc.), but some experimental errors and deviations should be accounted for. EXAMPLE 1 [00483] This example describes the materials, methods, and instrumentation used in EXAMPLE 2. Study participants [00484] Participants were healthy adults that had been vaccinated with 2 or 3 doses of an mRNA vaccine (mRNA-1273 (Moderna) or BNT162b2 (Pfizer)) and reported breakthrough SARS-CoV- 2 infection diagnosed by PCR or antigen testing. Breakthrough infection with delta or omicron variants was deduced based on the prevalent variant circulating in New York City at the time of infection (Gaebler et al., 2022). All participants were provided written informed consent before participating in the study, and the study was conducted in accordance with Good Clinical Practice. The study was performed in compliance with all relevant ethical regulations, and the protocol (DRO-1006) for studies with human participants was approved by the Institutional Review Board of the Rockefeller University. For detailed participant characteristics, see Table 1.
Blood samples processing and storage
[00485] V enous blood samples were collected into Heparin and Serum-gel monovette tubes by standard phlebotomy at the Rockefeller University. Peripheral Blood Mononuclear Cells (PBMCs) obtained from samples collected were further purified as previously reported by gradient centrifugation and stored in liquid nitrogen in the presence of Fetal Calf Serum (FCS) and Dimethylsulfoxide (DMSO) (Gaebler et al., 2021; Robbiani et al., 2020). Heparinized serum and plasma samples were aliquoted and stored at -20°C or less. Prior to experiments, aliquots of plasma samples were heat-inactivated (56°C for 1 hour) and then stored at 4°C.
ELISAs
[00486] Enzyme-Linked Immunosorbent Assays (ELISAs)(Amanat et al., 2020; Grifoni. et al,, 2020) were performed to evaluate antibodies binding to SARS-CoV-2 wild-type (Wuhan-Hu- 1) RBD, and variants of concern Delta (B.1 ,617.2) RBD, and Omicron (BA.1) spike protein by coating of high-binding 96-half-well plates (Corning 3690) with 50 μl per well of a 1 ug/ml indicated protein solution in Phosphate-buffered Saline (PBS) overnight at 4°C. Plates were washed 6 times with washing buffer (1 X PBS with 0.05% Tween-20 (Sigma-Aldrich)) and incubated with 170 μl per well blocking buffer (1 - PBS with 2% BSA and 0.05% Tween-20 (Sigma)) for 1 hour at room temperature. Immediately after blocking, plasma samples or monoclonal antibodies were added in PBS and incubated for 1 hour at room temperature. Plasma samples were assayed at. a 1:66 starting dilution and 10 additional 3 -fold serial dilutions.
[00487] 10 μg/ml starting concentration was used to test monoclonal antibodies, followed by 10 additional 4-fold serial dilutions. Plates were washed 6 times with washing buffer and then incubated with anti-human IgG secondary antibody conjugated to horseradish peroxidase (HRP) (Jackson Immuno Research 109-036-088 109-035-129 and Sigma A0295) in blocking buffer at a 1:5,000 dilution. Plates were developed by addition of the HRP substrate, 3, 3’, 5,5’- Tetramethyl benzidine (TMB) (ThermoFisher) for 10 minutes (plasma samples and monoclonal antibodies). 50 pl of 1 M H2SO4 was used to stop the reaction, and absorbance was measured at 450 nm with an ELISA microplate reader (FluoStar Omega, BMG Labtech) with Omega and Omega MARS software for analysis. A positive control (For anti-RBD ELISA, plasma from participant COV72, diluted 66.6-fold and ten additional threefold serial dilutions in PBS; for anti- Omicron spike ELISA, plasma from B039 was used as a control) was added to every assay plate for normalization for plasma samples. The average of its signal was used for normalization of all the other values on the same plate with Excel software before calculating the area under the curve using Prism V9.1 (GraphPad). Negative controls of pre-pandemic plasma samples from healthy donors were used for validation (for more details, please see Robbiani et al., 2020). For monoclonal antibodies, the ELISA half-maximal concentration (EC50) was determined using four-parameter nonlinear regression (GraphPad Prism V9. 1). EC50s above 1000 ng/mL were considered nonbinders.
Proteins
[00488] The mammalian expression vector encoding the Receptor Binding-Domain (RBD) of SARS-CoV-2 (GenBank MN985325.1; Spike (S) protein residues 319-539) was previously described (Barnes et al., 2020b).
SARS-CoV-2 pseudotyped reporter virus
[00489] A panel of plasmids expressing RBD-mutant SARS-CoV -2 spike proteins in the context of pSARS-CoV-2-SA19 has been described (Cho et al,, 2021 ; Muecksch et al., 2021 ; Wang et al., 2021c; Weisblum et al,, 2020). Variant pseudoviruses resembling SARS-CoV-2 variants Delta (B. 1.617.2) and Omicron BA. l (B.1,1 .529) have been described before (Cho et al., 2021; Schmidt et al., 2022; Wang et. al., 2021b) and were generated by introduction of substitutions using synthetic gene fragments (IDT) or overlap extension PCR mediated mutagenesis and Gibson assembly. Specifically, the variant-specific deletions and substitutions introduced were:
[00490] Delta: T19R, Al 56-158, L452R, T478K, D614G, P681R, D950N
[00491] Omicron BA.l : A67V, A69-70, T95I, G142D, A143-145, A211, L212I, ins214EPE, G339D, S371L, S373P, S375F, K417N, N440K, G446S, S477N, T478K, E484A, Q493K, G496S, Q498R, N501Y, Y505H, T547K, D614G, H655Y, H679K, P681H, N764K, D796Y, N856K, Q954H, N969H, N969K, L981F
[00492] Omicron BA.2: T19I, L24S, de!25-27, G142D, V213G, G339D, S371F, S373P, S375F, T376A, D405N, R408S, K417N, N440K, S477N, T478K, E484A, Q493R, Q498R, N501Y, Y5O5H, D614G, H655Y, N679K, P681H, N764K, D796Y, Q954H, N969K [00493] Omicron BA.4/5: T19I, L24S, del25-27, del69-70, G142D, V213G, G339D, S371F, S373P, S375F, T376A, D405N, R408S, K417N, N440K, L452R, S477N, T478K, E484A, F486V, Q498R, N501Y, Y505H, D614G, H655Y, N679K, P681H, N764K, D796Y, Q954H, N969K [00494] Deletions/substitutions corresponding to variants of concern listed above were incorporated into a spike protein that also includes the R683G substitution, which disrupts the furin cleavage site and increases particle infectivity. Neutralizing activity against mutant pseudoviruses was compared to a wild-type (WT) SARS-CoV-2 spike sequence (NC_045512), carrying R683G where appropriate. [00495] SARS-CoV-2 pseudotyped particles were generated as previously described (Robbiani et al., 2020; Schmidt et al., 2020a). Briefly, 293T (CRL-11268) cells were obtained from ATCC, and the cells were transfected with pNL4-^ǻ(QY-nanoluc and pSARS-CoV-2-Sǻ^^. Particles were harvested 48 hours post-transfection, filtered, and stored at -80°C. Pseudotyped virus neutralization assay [00496] Pre-pandemic negative control plasma from healthy donors, plasma from individuals who received mRNA vaccines and had Delta or Omicron BA.1 breakthrough infection, or monoclonal antibodies were five-fold serially diluted and incubated with SARS-CoV-2 pseudotyped virus for 1 hour at 37 °C. The mixture was subsequently incubated with 293TAce2 cells (Robbiani et al., 2020) (for all WT neutralization assays) or HT1080/Ace2 cl14 cells (for all variant neutralization assays) for 48 hours, after which cells were washed with PBS and lysed with Luciferase Cell Culture Lysis 5× reagent (Promega). Nanoluc Luciferase activity in lysates was measured using the Nano-Glo Luciferase Assay System (Promega) with the ClarioStar Microplate Multimode Reader (BMG). The relative luminescence units were normalized to those derived from cells infected with SARS-CoV-2 pseudotyped virus(Wang et al., 2021c) in the absence of plasma or monoclonal antibodies. The half-maximal neutralization titers for plasma (NT50) or half- maximal and 90% inhibitory concentrations for monoclonal antibodies (IC50 and IC90) were determined using four-parameter nonlinear regression (least squares regression method without weighting; constraints: top=1, bottom=0) (GraphPad Prism). Biotinylation of viral protein for use in flow cytometry [00497] Purified and Avi-tagged SARS-CoV-2 WT and Delta RBD were biotinylated using the Biotin-Protein Ligase-BIRA kit according to manufacturer’s instructions (Avidity) as described before (Robbiani et al., 2020). Ovalbumin (Sigma, A5503-1G) was biotinylated using the EZ-Link Sulfo-NHS-LC-Biotinylation kit according to the manufacturer’s instructions (Thermo Scientific). Biotinylated ovalbumin was conjugated to streptavidin-BV711 for single-cell sorts (BD biosciences, 563262) or to streptavidin-BB515 for phenotyping panel (BD, 564453). WT RBD was conjugated to streptavidin-PE (BD Biosciences, 554061) and streptavidin-AF647 (Biolegend, 405237) for single-cell sorts, or streptavidin- BV421 (Biolegend, 405225) and streptavidin-BV711 (BD biosciences, 563262) for phenotyping. Delta RBD was conjugated to streptavidin-PE (BD Biosciences, 554061) and Omicron BA. 1 RBD (ACROBiosystems, SPD-C82E4) was conjugated to streptavidin-AF647 (Biolegend, 405237).
Flow cytometry and single cell sorting
[00498] Single-cell sorting by flow cytometry was described previously (Robbiani et al., 2020). Simply, peripheral blood mononuclear cells (PBMC) were enriched for B cells by negative selection using a pan-B-cell isolation kit according to the manufacturer’s instructions (Miltenyi Biotec, 130-101-638). The enriched B cells were incubated in Fluorescence- Activated Cell-sorting (FACS) buffer (1 x PBS, 2% FCS, 1 mM ethylenediaminetetraacetic acid (EDTA)) with the following anti-human antibodies (all at 1 :200 dilution): anti-CD20-PECy7 (BD Biosciences, 335793), anti-CD3-APC-eFluro 780 (Invitrogen, 47-0037-41), anti-CD8-APC-eFluor 780 (Invitrogen, 47-0086-42), anti-CDl 6-APC-eFluor 780 (Invitrogen, 47-0168-41), anti-CDl 4-APC- eFluor 780 (Invitrogen, 47-0149-42), as well as Zombie NIR (BioLegend, 423105) and fluorophore-labeled RBD and ovalbumin (Ova) for 30 min on ice. Single CD3-CD8-CD14-CD16- CD20+Ova------\VT RBD-PE+- WT RBD-AF647+ B cells were sorted into individual wells of 96- well plates containing 4 pl of lysis buffer (0.5 x PBS, 10 mM Dithiothreitol (DTT), 3,000 units/ml RNasin Ribonuclease Inhibitors (Promega, N2615) per well using a FACS Aria III and FACSDiva software (Becton Dickinson) for acquisition and FlowJo for analysis. The sorted cells were frozen on dry ice, and then stored at -80 °C or immediately used for subsequent RNA reverse transcription. For B cell phenotype analysis, in addition to the above antibodies, B cells were also stained with the following anti-human antibodies (all at 1:200 dilution): anti-IgD-BV650 (BD, 740594), anti- CD27-BV786 (BD biosciences, 563327), anti-CDl 9-BV605 (Biolegend, 302244), anti-CD71 - PerCP-Cy5.5 (Biolegend, 334114), anti-IgG-PECF594 (BD, 562538), anti-IgM-AF700 (Biolegend, 314538), anti-IgA-Viogreen (Miltenyi Biotec, 130-113-481).
Antibody sequencing, cloning, and expression [00499] Antibodies were identified and sequenced as described previously (Robbiani et aL, 2020; Wang et aL, 2021a). In brief, RNA from single cells was reverse-transcribed (SuperScript III Reverse Transcriptase, Invitrogen, 18080-044), and the cDNA was stored at ---20 °C or used for subsequent amplification of the variable IGH, IGL, and IGK genes by nested PCR and Sanger sequencing. Sequence analysis was performed using Mac Vector. Amplicons from the first PCR reaction were used as templates for sequence- and ligation-independent cloning into antibody expression vectors. Recombinant monoclonal antibodies were produced and purified as previously described (Robbiani et al., 2020).
[00500] During this process, several non-native mutations were introduced into the heavy and light chain fragments. Principally, the kappa (light) chain fragments were amplified using primers that reached into the framework region of the antibodies. The kappa primer introduces non-native ammo acids into the kappa chain. In addition, the heavy chain was truncated at the C-termmus for the purpose of antibody production, being one amino acid shorter than what usually is present in IgG heavy chain constant regions.
Biolayer interferometry
[00501] Biolayer interferometry assays were performed as previously described (Robbiani et al,, 2020). In brief, the Octet Red instrument (ForteBio) was used at 30 °C with shaking at 1 ,000 r.p.m. Epitope binding assays were performed with protein A biosensor (ForteBio 18-5010), following the manufacturer’s protocol “classical sandwich assay” as follows: (1 ) Sensor check: sensors immersed 30 sec in buffer alone (buffer ForteBio 18-1105), (2) Capture 1st Ab: sensors immersed 10 mm with Abl at 10 μg/mL, (3) Baseline: sensors immersed 30 sec in buffer alone, (4) Blocking: sensors immersed 5 min with IgG isotype control at 10 μg/mL. (5) Baseline: sensors immersed 30 sec in buffer alone, (6) Antigen association: sensors immersed 5 min with RBD at 10 μg/mL. (7) Baseline: sensors immersed 30 sec in buffer alone. (8) Association Ab2: sensors immersed 5 min with Ab2 at 10 μg/mL. Curve fitting was performed using the Fortebio Octet Data analysis software (ForteBio). Affinity measurements of anti-SARS-CoV-2 IgGs binding were corrected by subtracting the signal obtained from traces performed with IgGs in the absence of WT RBD. The kinetic analysis using protein A biosensor (as above) was performed as follows: (1) baseline: 60 sec immersion in buffer. (2) loading: 200 sec immersion in a solution with IgGs 10 μg/ml. (3) baseline: 200 sec immersion in buffer. (4) Association: 300 sec immersion in solution with WT RBD at 20, 10 or 5 μg/ml (5) dissociation: 600 sec immersion in buffer. Curve fitting was performed using a fast 1:1 binding model and the Data analysis software (ForteBio). Mean KD values were determined by averaging all binding curves that matched the theoretical fit with an R2 value ı 0.8. Computational analyses of antibody sequences [00502] Antibody sequences were trimmed based on quality and annotated using Igblastn v.1.14. with IMGT domain delineation system. Annotation was performed systematically using Change- O toolkit v.0.4.540 (Gupta et al., 2015). Clonality of heavy and light chains was determined using DefineClones.py implemented by Change-O v0.4.5 (Gupta et al., 2015). The script calculates the Hamming distance between each sequence in the data set and its nearest neighbor. Distances are subsequently normalized and to account for differences in junction sequence length, and clonality is determined based on a cut-off threshold of 0.15. Heavy and light chains derived from the same cell were subsequently paired, and clonotypes were assigned based on their V and J genes using in-house R and Perl scripts. All scripts and the data used to process antibody sequences are publicly available on GitHub (github.com/stratust/igpipeline/tree/igpipeline2_timepoint_v2). [00503] The frequency distributions of human V genes in anti-SARS-CoV-2 antibodies from this study were compared to 131,284,220 IgH and IgL sequences generated by (Soto et al., 2019) and downloaded from cAb-Rep (Guo et al., 2019), a database of human shared BCR clonotypes available at cab-rep.c2b2.columbia.edu/. The IgH and IgL sequences were selected from the database that are partially coded by the same V genes and counted them according to the constant region. The frequencies shown in FIG.8 are relative to the source and isotype analyzed. The two- sided binomial test was used to check whether the number of sequences belonging to a specific IGHV or IGLV gene in the repertoire is different according to the frequency of the same IgV gene in the database. Adjusted p-values were calculated using the false discovery rate (FDR) correction. Significant differences are denoted with stars. [00504] Nucleotide somatic hypermutation and Complementarity-Determining Region (CDR3) length were determined using in-house R and Perl scripts. For somatic hypermutations, IGHV and IGLV nucleotide sequences were aligned against their closest germlines using Igblastn, and the number of differences was considered to correspond to nucleotide mutations. The average number of mutations for V genes was calculated by dividing the sum of all nucleotide mutations across all participants by the number of sequences used for the analysis. Code availability statement: [00505] Computer code to process the antibody sequences is available at GitHub (github.com/stratust/igpipeline/tree/igpipeline2_time- point_v2). EXAMPLE 2 Plasma binding and neutralization [00506] Plasma IgG antibody titers against SARS-CoV-2 Wuhan-Hu-1-(wild-type, WT), or Delta-receptor binding domain (RBD), and Omicron BA.1-Spike were measured by enzyme- linked immunosorbent assays (ELISA) (Wang et al., 2021c). Anti-WT-RBD IgG titers were significantly increased after Delta breakthrough infection in individuals who received 2 doses of mRNA vaccine (Delta BT), compared to vaccinated individuals who did not experience infection (5m-Vax2) (p<0.0001, Vax2 (Cho et al., 2021) vs. Delta, FIG. 1b and Table 1). Similarly, there was a 2-fold increase in geometric mean IgG-binding titers against WT-RBD after Omicron BA.1 breakthrough infection (Omicron BT) in individuals who received 3 doses of mRNA vaccine, compared to vaccinated individuals who were not infected after the 3rd vaccine dose (1m-Vax3) (p=0.033, Vax3 (Muecksch et al., 2022) vs. Omicron, FIG. 1b and Table 1). Individuals who experienced Omicron BA.1 infection exhibited higher anti-Delta-RBD and anti-Omicron BA.1- Spike IgG binding titers than individuals with Delta breakthrough infection or those receiving 3 mRNA vaccine doses (FIG. 4, anti-Delta RBD: p<0.0001, Delta BT vs. Omicron BT, p=0.047, Vax3 vs. Omicron BT; anti-Omicron BA.1 Spike: p<0.0001, Delta BT vs. Omicron BT, p=0.021, Vax3 vs. Omicron BT). [00507] Plasma neutralizing activity in 49 participants was measured using HIV-1 pseudotyped with the WT SARS-CoV-2 spike protein (Cho et al., 2021; Wang et al., 2021c) (FIG.1c and Table 1). Compared to individuals that received 2 mRNA vaccine doses(Cho et al., 2021), Delta breakthrough infection resulted in 11-fold increased geometric mean half-maximal neutralizing titer (NT50) (p=0.0003, Vax2 vs. Delta, FIG. 1c). However, the resulting geometric mean NT50 was lower than after the 3rd mRNA vaccine dose (p=0.03, Delta vs. Vax3, FIG. 1c). Notably, the NT50 against WT after Omicron breakthrough was not significantly different from individuals that received a 3rd vaccine dose (Muecksch et al., 2022) (p>0.99, Vax3 vs. Omicron, Fig. 1c). [00508] Plasma neutralizing activity was also assessed against SARS-CoV-2 Delta, Omicron BA.1, BA.2, and BA.4/5 variants using viruses pseudotyped with appropriate variant spike proteins. [00509] Delta breakthrough infection resulted in 15-fold increased neutralizing titers against Delta compared to 2-dose vaccinated-only individuals (p<0.0001, FIG. 1c) with resulting titers being comparable to 3-dose vaccinated individuals before and after Omicron breakthrough infection (p>0.99, FIG. 1c). While Delta breakthrough infection also increased neutralizing titers against Omicron BA.1, BA.2, and BA4/5 (p=0.0003, p=0.002 and p<0.0001, respectively, FIG. 1c), the titers were not significantly different from titers observed in 3-dose vaccinated individuals (Muecksch et al., 2022)(p>0.99, p>0.99 and p=0.61, respectively, FIG.1c). Conversely, Omicron breakthrough infection after 3-dose vaccination resulted in a further 3.5-fold and 2.9-fold increase of Omicron BA.1 and BA.2 neutralizing titers, respectively, when compared to 3-dose vaccinated only individuals (Muecksch et al., 2022)(p=0.005 and p=0.019, respectively. FIG. 1c). Omicron BA.4/5 showed the highest neutralization resistance of all variants tested, resulting in low geometric mean neutralizing titers in plasma samples obtained after the second vaccine dose (NT50=72, FIG. 1c). Nevertheless, individuals who had at least 3 antigen exposures (Delta breakthrough, Vax3 and Omicron breakthrough) were able to neutralize Omicron BA.4/5 with NT50s of 2173, 1311 and 2476, respectively at the time points assayed. Memory B cells [00510] mRNA vaccines elicit memory B cells (MBCs) that can contribute to durable immune protection from serious disease by mediating rapid and anamnestic antibody response (Muecksch et al., 2022; Victora and Nussenzweig, 2022). To better understand the MBC compartment after Delta or Omicron BA.1 breakthrough infection in vaccinated individuals, RBD-specific MBCs were enumerated using Alexa Fluor 647 (AF647)- and phycoerythrin (PE)-labeled WT RBD of the SARS-CoV-2 spike protein by flow cytometry (FIG. 2a, FIG. 5). The number of WT RBD- specific MBCs after Delta breakthrough infection was significantly higher than after the 2nd or 3rd vaccine dose (Delta vs. Vax2, p<0.0001, and Delta vs. Vax3, p=0.011, FIG. 2b). Omicron BA.1 breakthrough infection elicited a 1.7-fold increase in the number of MBCs compared to individuals that received 3 vaccine doses (Vax3 vs. Omicron, p=0.013, FIG.2b). Consistent with previous reports (Goel et al., 2022; Kaku et al., 2022; Nutalai et al., 2022; Park et al., 2022), flow cytometry showed that a larger fraction of the MBCs developing after the 3rd vaccine dose or Omicron BA.1 breakthrough infection were cross-reactive with WT-, Delta- and Omicron BA.1- RBDs than after Delta breakthrough infection (FIG. 2c). Additional phenotyping indicated that RBD-specific memory B cells elicited by Vax3 or Delta or Omicron BA.1 breakthrough infection showed higher frequencies of IgG than IgM and IgA expression (FIGs.5c-e). [00511] To examine the specificity and neutralizing activity of the antibodies produced by MBCs, antibody genes were purified and sequenced in individual WT-RBD-specific B cells from 10 individuals that experienced Delta or Omicron BA.1 breakthrough infection, following the 2nd or 3rd vaccine dose, respectively (FIG. 2d, FIG. 5f, Table 1), including 2 participants for whom paired samples were collected shortly after their 3rd vaccine dose and after subsequent Omicron BA.1 breakthrough infection. [00512] 686 paired heavy and light chain anti-RBD antibody sequences were obtained (FIG.2d). Clonally expanded WT-RBD-specific B cells represented 9% of all memory B cells after Delta breakthrough infection and 28% of the repertoire after Omicron BA.1 breakthrough infection (FIG. 2d). Similar to mRNA vaccinees (Cho et al., 2021; Muecksch et al., 2022; Wang et al., 2021c), several sets of VH genes including VH3-30 and VH3-53 were over-represented in Delta- or Omicron BA.1 breakthrough infection (FIGs. 6a-f). In addition, VH3-49, VH4-38, and VH1-24 were exclusively over-represented after Delta breakthrough infection (FIG. 6a), while VH1-69, VH1-58, VH4-61, and VH4-38 were specifically over-represented after Omicron BA.1 breakthrough infection (FIG. 6d). These results suggest that Delta and Omicron BA.1 breakthrough infections elicit variant-specific memory antibody responses. While levels of somatic mutation in memory B cells emerging after Delta breakthrough infection were comparable to those after the 2nd vaccine dose, significantly higher numbers of somatic mutations were noted following Omicron BA.1 breakthrough infection compared to the 3rd vaccine dose (p<0.0001) (FIG. 2e, FIG. 6g). Monoclonal antibodies [00513] 338 anti-RBD monoclonal antibodies were expressed and tested for binding by ELISA, including 115 antibodies obtained after Delta breakthrough infection (Delta BT), 40 isolated from 2 longitudinal samples after their 3rd vaccine dose in individuals that were subsequently infected (Vax3), and 183 antibodies obtained from 6 individuals after Omicron BA.1 breakthrough infection (Omicron). 85% (n=288) of the antibodies bound to the WT RBD with an EC50 of less than 1000 ng/mL (Table 2). The geometric mean ELISA half-maximal concentration (EC50) against WT RBD for the monoclonal antibodies obtained from Vax3 was comparable to those found after Delta or Omicron BA.1 breakthrough infections (FIG. 3a). In addition, antibodies isolated after both Delta and Omicron-breakthrough infection showed comparable affinity for WT RBD to antibodies obtained from Vax3 when measured by biolayer interferometry (BLI, FIG.7a). However, when tested against Delta-RBD antibodies obtained after Delta breakthrough infection showed increased binding compared to those after Vax3. In contrast, there was no statistically significant difference in binding to Omicron BA.1-Spike by Omicron and Vax3 antibodies (FIG. 3a). [00514] Anti-RBD antibodies elicited by mRNA vaccination target 4 structurally defined classes of epitopes on the SARS-CoV-2 RBD (Barnes et al., 2020a; Cho et al., 2021; Muecksch et al., 2022; Yuan et al., 2020). To compare the epitopes recognized by anti-RBD memory antibodies elicited by mRNA vaccination (Muecksch et al., 2022) and breakthrough infection, BLI competition experiments were performed. A preformed antibody-RBD-complex was exposed to a second antibody recognizing one of four classes of structurally defined antigenic sites (C105 as Class 1; C144 as Class 2, C135 as Class 3, and C2172 as Class 4 (Barnes et al., 2020a)). Antibodies obtained after Delta (n=48) or Omicron BA.1 (n=49) breakthrough infection were examined, including 30 of 48 from Delta Breakthrough and 30 of 49 from Omicron BA.1 breakthrough with IC50s lower than 1000 ng/mL (neutralizing) against WT (FIG. 7b). In general, there was no significant difference in the distribution of targeted epitopes among antibodies obtained following breakthrough infection as compared to those obtained after mRNA vaccination (FIG.7b). [00515] All 288 WT RBD-binding antibodies were tested for neutralization in a SARS-CoV-2 pseudotype neutralization assay based on the WT SARS-CoV-2 spike (Robbiani et al., 2020; Schmidt et al., 2020b). For comparison, a previously characterized set of antibodies isolated after the 2nd (Cho et al., 2021) or the 3rd vaccine dose (Muecksch et al., 2022) were used. Potency against WT was considerably improved after Delta breakthrough infection compared to the 2nd vaccine dose (Vax2) (IC50=182 ng/ml vs IC50=50ng/ml, p=0.0013, FIG.3b) but not compared to the 3rd vaccine dose (IC50=98 ng/ml, p=0.62, FIG. 3b). In addition, there was no further improvement of neutralizing activity following Omicron BA.1 breakthrough infection compared to the 3rd dose (IC50=73 ng/ml) (p>0.99, FIG.3b). [00516] To examine whether and how neutralizing breadth evolves in vaccinees after Delta or Omicron BA.1 breakthrough infection, the 288 newly expressed antibodies obtained from breakthrough individuals and 45 previously described antibodies obtained from Vax2 individuals (Cho et al., 2021) were analyzed, and their neutralizing activity against SARS-CoV-2 pseudoviruses carrying amino acid substitutions found in the Delta-RBD and Omicron BA.1 variant was measured. In addition, 105 randomly selected antibodies from all four groups were tested against an Omicron BA.4/5 pseudovirus (FIG. 3c). Neutralizing potency was generally lower against Omicron BA.1 compared to Delta pseudovirus. However, while antibodies obtained 5 months after the 2nd vaccine dose were not significantly more potent against Delta (IC50=181ng/ml) vs. BA.1 pseudovirus (IC50=405ng/ml) (p=0.20, FIG.3c), those obtained after subsequent Delta breakthrough infection neutralize Delta with 6.8-fold increased potency compared to BA.1 (p<0.0001, FIG. 3c). In contrast, the ratio of Delta vs. BA.1 IC50 in Vax3 antibodies was only 2.2 (p=0.07, FIG. 3c), while antibodies recovered after subsequent omicron breakthrough neutralized Delta and Omicron with similar potencies IC50=122 ng/ml for Delta vs. 148 ng/ml for Omicron (p=0.92, FIG. 3c). [00517] Compared to the 2nd vaccine dose, antibodies from Delta breakthrough infection showed increased potency against Delta pseudovirus (181 vs. 61 ng/ml, p=0.047, FIG. 3d). However, there was no significant improvement of antibody potency against Delta, Omicron BA.1 or Omicron BA.4/5 pseudovirus comparing 5m-Vax2 versus Delta breakthrough antibodies. Moreover, there were only 2 fold differences that did not reach statistical significance when comparing Vax3 and Omicron breakthrough antibodies for Omicron BA.1 and BA.4/5 neutralization (FIG. 3d). Notably, Omicron BA.4/5 showed the highest degree of neutralization resistance for all tested antibody groups (FIGs. 3c-d). [00518] When comparing the fraction of antibodies showing neutralizing activity against Delta or Delta+WT, or Omicron BA.1 or Omicron BA.1+WT, or all three viruses (WT+Delta+Omicron BA.1), it became apparent that antibodies isolated after 2 vaccine doses and subsequent Delta breakthrough infection show the largest proportion of Delta-neutralizing antibodies. Conversely, antibodies isolated after the 3rd vaccine dose and subsequent Omicron BA.1 breakthrough infection show the largest number of antibodies that neutralized all three pseudoviruses (FIG. 3e, FIG. 8). Vax3 antibodies and Omicron BA.1 breakthrough antibodies were enriched for those neutralizing BA.4/5 with IC50 values of less than 1000 ng/ml with 37% and 38% of all tested antibodies neutralizing BA.4/5, respectively, while only 17% and 27% of Vax2 and Delta breakthrough antibodies, respectively neutralized BA.4/5 (FIG. 3f). Thus, in both cases tested a 3rd exposure to antigen increases memory antibody potency and breadth but a 4th exposure with Omicron BA.1 does little more when it occurs in the time frame measured in this study. Discussion [00519] Individuals that receive a 3rd mRNA vaccine dose show enhanced protection against severe COVID19 but little is known about the impact of breakthrough infections on memory responses. Here, the memory antibodies that develop after a 3rd or 4th antigenic exposure by Delta or Omicron BA.1 infection were examine, respectively. A 3rd exposure to antigen by Delta breakthrough increases the number of memory B cells that produce antibodies with comparable potency and breadth to a 3rd mRNA vaccine dose. A 4th antigenic exposure with Omicron BA.1 infection increased variant specific plasma antibody and memory B cell responses. However, the 4th exposure did not increase the overall frequency of memory B cells or their general potency or breadth compared to a 3rd mRNA vaccine dose. In conclusion, a 3rd antigenic exposure by Delta infection elicits strain-specific memory responses and increases in the overall potency and breadth of the memory B cells. In contrast, the effects of a 4th antigenic exposure with Omicron BA.1 is limited to increased strain specific memory with little effect on the potency or breadth of memory B cell antibodies. The results suggest that the effect of strain-specific boosting on memory B cell compartment may be limited. [00520] Omicron and its subvariants are reported to be more transmissible than any prior VoC and have spurred a resurgence of new cases worldwide (Mallapaty, 2022). While early reports suggested that Omicron may cause less severe illness, recent studies show variant-specific symptoms but similar virulence (Whitaker et al., 2022), and increased resistance to approved vaccine regimens (Nealon and Cowling, 2022). [00521] It was shown that a 3rd mRNA vaccine dose boosts plasma antibody responses to SARS-CoV-2 variants, including Omicron BA.1, and increases the number, potency, and breadth of the antibodies found in the memory B cell compartment (Goel et al., 2022; Muecksch et al., 2022). Although the antibodies in plasma are generally not sufficient to prevent breakthrough infection boosted individuals are protected against serious disease upon breakthrough infection (Kuhlmann et al., 2022; Nemet et al., 2022). The present study shows that a 3rd exposure to antigen in the form of Delta breakthrough infection produces similar effects on the overall size of the memory compartment to a 3rd mRNA vaccine dose, and specifically boosts strain-specific responses. In contrast, while a 4th antigen exposure by infection with Omicron elicits strain- specific memory, it has far more modest effects on the overall potency and breadth of memory B cell antibodies. The data indicate that a variant-specific mRNA vaccine boost will increase plasma neutralizing activity and memory B-cells that are specific to the variant and closely related strains but may not elicit memory B cells with better general potency or breadth than the Wuhan-Hu-1- based mRNA vaccine. The data highlights the challenges involved in selecting variant-specific vaccines in the absence of reliable information on the nature of the next emerging variant and suggests that a focus should be on designing vaccines with broader general activity against coronaviruses. [00522] The foregoing examples and description of the preferred embodiments should be taken as illustrating, rather than as limiting the present invention as defined by the claims. As will be readily appreciated, numerous variations and combinations of the features set forth above can be utilized without departing from the present invention as set forth in the claims. Such variations are not regarded as a departure from the scope of the invention, and all such variations are intended to be included within the scope of the following claims. All references cited herein are incorporated by reference in their entireties. * * * * * [00523] It will be appreciated by those skilled in the art that changes could be made to the embodiments described above without departing from the broad inventive concept thereof. It is understood, therefore, that this invention is not limited to the particular embodiments disclosed, but it is intended to cover modifications within the spirit and scope of the present invention as defined by the present description. [00524] Various publications, articles and patents are cited or described in the background and throughout the specification; each of these references is herein incorporated by reference in its entirety. Discussion of documents, acts, materials, devices, articles or the like which has been included in the present specification is for the purpose of providing context for the invention. 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TABLES AND SEQUENCES
Table 1. Individual participant characteristics
Study subgroup
Previously
Age published Breakthrough
ID (years) Sex Race Ethnicity cohort infection VOC 1st dose 2nd dose 3rd dose
Figure imgf000167_0001
Figure imgf000168_0001
Figure imgf000169_0001
V = published as Gaebler. C,., DaSilva, J., Bednardski, E.. et al. SARS-CoV-2 neutralization after mRNA vaccination and variant breakthrough infection. Open Forum Infectious Diseases (2022). doi: doi.org/10.1093/ofid/otac227
= published as Cho, A., Muecksch, F., Schaefer-Babajew, D., et al. Anti-SARS-CoV-2 receptor binding domain antibody evolution after mRNA vaccination. Nature 1-9 (2021) doi: 10. 1038/s41586-021-04060-7.
§ = published as Muecksch, F., Wang, Z., Cho, A., et al. Increased Memory B Cell Potency and Breadth After a SARS-CoV -2 mRNA Boost. Nature (2022). doi: doi.org/10.1038/s41586-022-04778-y
Table 1 (continued)
Figure imgf000170_0001
Figure imgf000171_0001
Figure imgf000172_0001
Figure imgf000173_0002
Figure imgf000173_0001
Figure imgf000174_0001
Table 2. Sequences, RBD binding and neutralization nf representative cloned recombinant antibodies.
Figure imgf000175_0001
Figure imgf000176_0001
Figure imgf000177_0001
Figure imgf000178_0001
Figure imgf000179_0001
Figure imgf000180_0001
Figure imgf000181_0001
Figure imgf000182_0001
Figure imgf000183_0001
Figure imgf000184_0001
Figure imgf000185_0001
Figure imgf000186_0001
Figure imgf000187_0001
Figure imgf000188_0001
Figure imgf000189_0001
Figure imgf000190_0001
Figure imgf000191_0001
Figure imgf000192_0001
Figure imgf000193_0001
Figure imgf000194_0001
Figure imgf000195_0001
Figure imgf000196_0001
Figure imgf000197_0001
Figure imgf000198_0001
Figure imgf000199_0001
Figure imgf000200_0001
Figure imgf000201_0001
Figure imgf000202_0001
Figure imgf000203_0001
Figure imgf000204_0001
Figure imgf000205_0001
Figure imgf000206_0001
Figure imgf000207_0001
Figure imgf000208_0001
Figure imgf000209_0001
Figure imgf000210_0001
Figure imgf000211_0001
Figure imgf000212_0001
Figure imgf000213_0001
Figure imgf000214_0001
Figure imgf000215_0001
Figure imgf000216_0001
Figure imgf000217_0001
Figure imgf000218_0001
Figure imgf000219_0001
Figure imgf000220_0001
Figure imgf000221_0001
Figure imgf000222_0001
Figure imgf000223_0001
Figure imgf000224_0001
Figure imgf000225_0001
Figure imgf000226_0001
Figure imgf000227_0001
Figure imgf000228_0001
Figure imgf000229_0001
Figure imgf000230_0001
Figure imgf000231_0001
Figure imgf000232_0001
Figure imgf000233_0001
Figure imgf000234_0001
Figure imgf000235_0001
Figure imgf000236_0001
Figure imgf000237_0001
Figure imgf000238_0001
Figure imgf000239_0001
Figure imgf000240_0001
Figure imgf000241_0001
Figure imgf000242_0001
Figure imgf000243_0001
Figure imgf000244_0001
Figure imgf000245_0001
Table 2 (continued)
Figure imgf000247_0001
Figure imgf000248_0001
Figure imgf000249_0001
Figure imgf000250_0001
Figure imgf000251_0001
Figure imgf000252_0001
Figure imgf000253_0001
Figure imgf000254_0001
Figure imgf000255_0001
Figure imgf000256_0001
Figure imgf000257_0001
Figure imgf000258_0001
Figure imgf000259_0001
Figure imgf000260_0001
Figure imgf000261_0001
Figure imgf000262_0001
Figure imgf000263_0001
Figure imgf000264_0001
Figure imgf000265_0001
Figure imgf000266_0001
Figure imgf000267_0001
Figure imgf000268_0001
Figure imgf000269_0001
Figure imgf000270_0001
Figure imgf000271_0001
Figure imgf000272_0001
Figure imgf000273_0001
Figure imgf000274_0001
Figure imgf000275_0001
Figure imgf000276_0001
Figure imgf000277_0001
Figure imgf000278_0001
Figure imgf000279_0001
Figure imgf000280_0001
Figure imgf000281_0001
Figure imgf000282_0001
Figure imgf000283_0001
Figure imgf000284_0001
Figure imgf000285_0001
Figure imgf000286_0001
Figure imgf000287_0001
Figure imgf000288_0001
Figure imgf000289_0001
Figure imgf000290_0001
Figure imgf000291_0001
Figure imgf000292_0001
Figure imgf000293_0001
Figure imgf000294_0001
Figure imgf000295_0001
Figure imgf000296_0001
Figure imgf000297_0001
Figure imgf000298_0001
Figure imgf000299_0001
Figure imgf000300_0001
Figure imgf000301_0001
Figure imgf000302_0001
Figure imgf000303_0001
Figure imgf000304_0001
Figure imgf000305_0001
Figure imgf000306_0001
Figure imgf000307_0001
Figure imgf000308_0001
Figure imgf000309_0001
Figure imgf000310_0001
Figure imgf000311_0001
Figure imgf000312_0001
Figure imgf000313_0001
Figure imgf000314_0001
Figure imgf000315_0001
Figure imgf000316_0001
Figure imgf000317_0001
Figure imgf000318_0001
Figure imgf000319_0001
Figure imgf000320_0001
Figure imgf000321_0001
Figure imgf000322_0001
Figure imgf000323_0001
Figure imgf000324_0001
Figure imgf000325_0001
Figure imgf000326_0001
Figure imgf000327_0001
Figure imgf000328_0001
Figure imgf000329_0001
Figure imgf000330_0001
Figure imgf000331_0001
Figure imgf000332_0001
Figure imgf000333_0001
Figure imgf000334_0001
Figure imgf000335_0001
Figure imgf000336_0001
Figure imgf000337_0001
Figure imgf000338_0001
Figure imgf000339_0001
Figure imgf000340_0001
Figure imgf000341_0001
Figure imgf000342_0001
Figure imgf000343_0001
Figure imgf000344_0001
Figure imgf000345_0001
Figure imgf000346_0001
Figure imgf000347_0001
Figure imgf000348_0001
Figure imgf000349_0001
Figure imgf000350_0001
Figure imgf000351_0001
Figure imgf000352_0001
Figure imgf000353_0001
Figure imgf000354_0001
Figure imgf000355_0001
Figure imgf000356_0001
Figure imgf000357_0001
Figure imgf000358_0001
Figure imgf000359_0001
Figure imgf000360_0001
Figure imgf000361_0001
Figure imgf000362_0001
Figure imgf000363_0001
Figure imgf000364_0001
Figure imgf000365_0001
Figure imgf000366_0001
Figure imgf000367_0001
Figure imgf000368_0001
Figure imgf000369_0001
Figure imgf000370_0001
Figure imgf000371_0001
Figure imgf000372_0001
Figure imgf000373_0001
Figure imgf000374_0001
Figure imgf000375_0001
Figure imgf000376_0001
Figure imgf000377_0001
Figure imgf000378_0001
Figure imgf000379_0001
Figure imgf000380_0001
Figure imgf000381_0001
Figure imgf000382_0001
Figure imgf000383_0001
Figure imgf000384_0001
Figure imgf000385_0001
Figure imgf000386_0001
Figure imgf000387_0001
Figure imgf000388_0001
Figure imgf000389_0001
Figure imgf000390_0001
Figure imgf000391_0001
Figure imgf000392_0001
Figure imgf000393_0001
Figure imgf000394_0001
Figure imgf000395_0001
Figure imgf000396_0001
Figure imgf000397_0001
Figure imgf000398_0001
Figure imgf000399_0001
Figure imgf000400_0001
Figure imgf000401_0001
Figure imgf000402_0002
Figure imgf000402_0001
Figure imgf000403_0001
Figure imgf000404_0001
Figure imgf000405_0001
Figure imgf000406_0001
Figure imgf000407_0001
Figure imgf000408_0001
Figure imgf000409_0001
Figure imgf000410_0001
Figure imgf000411_0001
Figure imgf000412_0001
Figure imgf000413_0001
Figure imgf000414_0001
Figure imgf000415_0001
Figure imgf000416_0001
Figure imgf000417_0001
Figure imgf000418_0001
Figure imgf000419_0001
Figure imgf000420_0001
Figure imgf000421_0001
Figure imgf000422_0001
Figure imgf000423_0001
Figure imgf000424_0001
Figure imgf000425_0001
Figure imgf000426_0001
Figure imgf000427_0001
Figure imgf000428_0001
Figure imgf000429_0001
Figure imgf000430_0001
Figure imgf000431_0001
Figure imgf000432_0001
Figure imgf000433_0001
Figure imgf000434_0001
Figure imgf000435_0001
Figure imgf000436_0001
Figure imgf000437_0001
Figure imgf000438_0001
Figure imgf000439_0001
Figure imgf000440_0001
Figure imgf000441_0001
Figure imgf000442_0001
Figure imgf000443_0001
Figure imgf000444_0001
Figure imgf000445_0001
Figure imgf000446_0001
Figure imgf000447_0001
Figure imgf000448_0001
Figure imgf000449_0001
Figure imgf000450_0001
Figure imgf000451_0001
Figure imgf000452_0001
Figure imgf000453_0001
Figure imgf000454_0001
Figure imgf000455_0001
Figure imgf000456_0001
Figure imgf000457_0001
Figure imgf000458_0001
Figure imgf000459_0001
Figure imgf000460_0001
Figure imgf000461_0001
Figure imgf000462_0001
Figure imgf000463_0001
Figure imgf000464_0001
Figure imgf000465_0001
Figure imgf000466_0001
Figure imgf000467_0001
Figure imgf000468_0001
Figure imgf000469_0001
Figure imgf000470_0001
Figure imgf000471_0001
Figure imgf000472_0001
Figure imgf000473_0001
Figure imgf000474_0001
Figure imgf000475_0001
Figure imgf000476_0001
Figure imgf000477_0001
Figure imgf000478_0001
Figure imgf000479_0001
Figure imgf000480_0001
Figure imgf000481_0001
Figure imgf000482_0001
Figure imgf000483_0001
Figure imgf000484_0001
Figure imgf000485_0001
Figure imgf000486_0001
Figure imgf000487_0001
Figure imgf000488_0001
Figure imgf000489_0001
Figure imgf000490_0001
Figure imgf000491_0001
Figure imgf000492_0001
Figure imgf000493_0001
Figure imgf000494_0001
Figure imgf000495_0001
Figure imgf000496_0001
Figure imgf000497_0001
Figure imgf000498_0001
Figure imgf000499_0001
Figure imgf000500_0001
Figure imgf000501_0001
Figure imgf000502_0001
Figure imgf000503_0001
Figure imgf000504_0001
Figure imgf000505_0001
Figure imgf000506_0001
Figure imgf000507_0001
Figure imgf000508_0001
Table 2 (continued)
Figure imgf000509_0001
Figure imgf000510_0001
Figure imgf000511_0001
Figure imgf000512_0001
Figure imgf000513_0001
Figure imgf000514_0001
Figure imgf000515_0001
Figure imgf000516_0001
Figure imgf000517_0001
Figure imgf000518_0001
Figure imgf000519_0001

Claims

CLAIMS 1. An antibody or antigen-binding fragment thereof that binds to the S polypeptide of a Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), wherein the antibody or antigen- binding fragment thereof comprises: a. a variable heavy chain CDR1 (HCDR1) comprising an amino acid sequence selected from SEQ ID NO: 1354, 1360, 1366, 1372, and 1378; b. a variable heavy chain CDR2 (HCDR2) comprising an amino acid sequence of SEQ ID NO: 1355, 1361, 1367, 1373, and 1379; c. a variable heavy chain CDR3 (HCDR3) comprising an amino acid sequence selected from SEQ ID NO: 1356, 1362, 1368, 1374, and 1380; d. a variable light chain CDR1 (LCDR1) comprising an amino acid sequence selected from SEQ ID NO: 1357, 1363, 1369, 1375, and 1381; e. a variable light chain CDR2 (LCDR2) comprising an amino acid sequence selected from SEQ ID NO: 1358, 1364, 1370, 1376, and 1382; and f. a variable light chain CDR3 (LCDR3) comprising an amino acid sequence selected from SEQ ID NO: 1359, 1365, 1371, 1377, and 1383.
2. The antibody or antigen-binding fragment thereof of claim 1, wherein the antibody or antigen-binding fragment thereof comprises: a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1354, a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1355, a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1356, a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1357, a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1358, and a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1359.
3. The antibody or antigen-binding fragment thereof of claim 2, wherein the antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising an amino acid sequence that is at least about 75%, at least 85%, at least 90%, at least 95%, at least 99%, or 100% identical to SEQ ID NO: 25, and a light chain variable domain (VL) comprising an amino acid sequence that is at least about 75%, at least 85%, at least 90%, at least 95%, at least 99%, or 100% identical to SEQ ID NO: 26.
4. The antibody or antigen-binding fragment thereof of claim 2 or claim 3, wherein the antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising or consisting of the amino acid sequence of SEQ ID NO: 25, and a light chain variable domain (VL) comprising or consisting of the amino acid sequence of SEQ ID NO: 26.
5. The antibody or antigen-binding fragment thereof of claim 1, wherein the antibody or antigen-binding fragment thereof comprises a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1360, a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1361, a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1362, a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1363, a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1364, and a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1365.
6. The antibody or antigen-binding fragment thereof of claim 5, wherein the antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising an amino acid sequence that is at least about 75%, at least 85%, at least 90%, at least 95%, at least 99%, or 100% identical to SEQ ID NO: 395, and a light chain variable domain (VL) comprising an amino acid sequence that is at least about 75%, at least 85%, at least 90%, at least 95%, at least 99%, or 100% identical to SEQ ID NO: 396.
7. The antibody or antigen-binding fragment thereof of claim 5 or claim 6, wherein the antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising or consisting of the amino acid sequence of SEQ ID NO: 395, and a light chain variable domain (VL) comprising or consisting of the amino acid sequence of SEQ ID NO: 396.
8. The antibody or antigen-binding fragment thereof of claim 1, wherein the antibody or antigen-binding fragment thereof comprises a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1366, a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1367, a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1368, a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1369, a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1370, and a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1371.
9. The antibody or antigen-binding fragment thereof of claim 8, wherein the antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising an amino acid sequence that is at least about 75%, at least 85%, at least 90%, at least 95%, at least 99%, or 100% identical to SEQ ID NO: 397, and a light chain variable domain (VL) comprising an amino acid sequence that is at least about 75%, at least 85%, at least 90%, at least 95%, at least 99%, or 100% identical to SEQ ID NO: 398.
10. The antibody or antigen-binding fragment thereof of claim 8 or claim 9, wherein the antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising or consisting of the amino acid sequence of SEQ ID NO: 397, and a light chain variable domain (VL) comprising or consisting of the amino acid sequence of SEQ ID NO: 398.
11. The antibody or antigen-binding fragment thereof of claim 1, wherein the antibody or antigen-binding fragment thereof comprises a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1372, a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1373, a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1374, a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1375, a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1376, and a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1377.
12. The antibody or antigen-binding fragment thereof of claim 11, wherein the antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising an amino acid sequence that is at least about 75%, at least 85%, at least 90%, at least 95%, at least 99%, or 100% identical to SEQ ID NO: 191, and a light chain variable domain (VL) comprising an amino acid sequence that is at least about 75%, at least 85%, at least 90%, at least 95%, at least 99%, or 100% identical to SEQ ID NO: 192.
13. The antibody or antigen-binding fragment thereof of claim 11 or claim 12, wherein the antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising or consisting of the amino acid sequence of SEQ ID NO: 191, and a light chain variable domain (VL) comprising or consisting of the amino acid sequence of SEQ ID NO: 192.
14. The antibody or antigen-binding fragment thereof of claim 1, wherein the antibody or antigen-binding fragment thereof comprises a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1378, a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1379, a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1380, a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1381, a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1382, and a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1383.
15. The antibody or antigen-binding fragment thereof of claim 14, wherein the antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising an amino acid sequence that is at least about 75%, at least 85%, at least 90%, at least 95%, at least 99%, or 100% identical to SEQ ID NO: 219, and a light chain variable domain (VL) comprising an amino acid sequence that is at least about 75%, at least 85%, at least 90%, at least 95%, at least 99%, or 100% identical to SEQ ID NO: 220.
16. The antibody or antigen-binding fragment thereof of claim 14 or claim 15, wherein the antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising or consisting of the amino acid sequence of SEQ ID NO: 219, and a light chain variable domain (VL) comprising or consisting of the amino acid sequence of SEQ ID NO: 220.
17. The antibody or antigen-binding fragment thereof of any one of claims 1-16, wherein the antibody or antigen-binding fragment thereof is a fully human antibody or antigen-binding fragment thereof.
18. The antibody or antigen-binding fragment thereof of any one of claims 1-17, wherein the antibody or antigen-binding fragment thereof binds to an epitope within the receptor binding domain (RBD) or the N-terminal domain (NTD) of the S polypeptide.
19. The antibody or antigen-binding fragment thereof of any one of claims 1-16, wherein the antibody or antigen-binding fragment thereof binds to an epitope selected from the following: a. the supersite b-hairpin (residues 152-158 of SEQ ID NO: 1353); b. WKH^ȕ^-strand (residue 97-102 of SEQ ID NO: 1353); c. N4-loop (residues 178-188 of SEQ ID NO: 1353) d. N-linked glycans at positions N122 and N149 of the spike polypeptide of SEQ ID NO: 1353; e. residues 27-32, 57-60, 210-218, and/or 286-303 of the spike polypeptide of SEQ ID NO: 1353; and/or f. residues 600-606 of the spike polypeptide of SEQ ID NO: 1353.
20. An antibody or antigen-binding fragment thereof, wherein the antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising an amino acid sequence that is at least about 75%, at least 85%, at least 90%, at least 95%, at least 99%, or 100% identical to a SEQ ID NO selected from column 7 of Table 2, and a light chain variable domain (VL) comprising an amino acid sequence that is at least about 75%, at least 85%, at least 90%, at least 95%, at least 99%, or 100% identical to a SEQ ID NO selected from column 11 of Table 2, wherein the VH SEQ ID No and the VL SEQ ID No are selected from the same row.
21. The antibody or antigen-binding fragment thereof of claim 20, wherein the antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising or consisting of a SEQ ID NO selected from column 7 of Table 2, and a light chain variable domain (VL) comprising or consisting of a SEQ ID NO selected from column 11 of Table 2, wherein the VH SEQ ID No and the VL SEQ ID No are selected from the same row.
22. The antibody or antigen-binding fragment thereof of any one of claims 1-19, wherein the antibody or antigen-binding fragment thereof neutralizes SARS-CoV-2.
23. The antibody or antigen-binding fragment thereof of claim 22, wherein the antibody or antigen-binding fragment thereof inhibits the fusion of SARS-CoV-2 and a host cell membrane.
24. The antibody or antigen-binding fragment thereof of any one of claims 1-23, wherein the antibody or antigen-binding fragment thereof is cytotoxic to a SARS-CoV-2 infected host cell.
25. The antibody or antigen-binding fragment thereof of any one of the preceding claims, wherein the antibody or antigen-binding fragment thereof is a multivalent antibody or antigen- binding fragment comprising (a) a first target binding site that specifically binds to an epitope within the spike polypeptide, and (b) a second target binding site that binds to an epitope on a different epitope on the spike polypeptide or a different molecule.
26. The antibody or the antigen-binding fragment thereof of any one of the preceding claims, further comprising a variant Fc constant region.
27. The antibody or antigen-binding fragment thereof of any one of the preceding claims, wherein the antibody or antigen-binding fragment thereof is a chimeric antibody, a humanized antibody, or humanized antibody.
28. The antibody or antigen-binding fragment thereof of any one of the preceding claims, wherein the antibody or antigen-binding fragment thereof is a single-chain antibody, Fab, or Fab2 fragment.
29. The antibody or antigen-binding fragment thereof of any one of the preceding claims, wherein the antibody or antigen-binding fragment thereof is detectably labeled or conjugated to a toxin, a therapeutic agent, a polymer, a receptor, an enzyme, or a receptor ligand, preferably wherein the polymer is polyethylene glycol (PEG).
30. A polynucleotide encoding the antibody or antigen-binding fragment thereof of any one of claims 1-29.
31. A vector, comprising the polynucleotide of claim 30.
32. A cultured host cell comprising the vector of claim 31
33. A pharmaceutical composition comprising the antibody or antigen-binding fragment thereof of any one of claims 1-29, the polynucleotide of claim 30, or the vector of claim 31.
34. A pharmaceutical composition comprising two or more of monoclonal antibodies or antigen-binding fragments thereof, wherein the two or more monoclonal antibodies or antigen- binding fragments thereof are selected from: a. an antibody or antigen-binding fragment thereof comprising: a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1354, a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1355, a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1356, a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1357, a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1358, and a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1359; b. an antibody or antigen-binding fragment thereof comprising: a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1360, a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1361, a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1362, a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1363, a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1364, and a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1365; c. an antibody or antigen-binding fragment thereof comprising: a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1366, a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1367, a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1368, a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1369, a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1370, and a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1371; d. an antibody or antigen-binding fragment thereof comprising: a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1372, a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1373, a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1374, a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1375, a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1376, and a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1377; and e. an antibody or antigen-binding fragment thereof comprising: a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1378, a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1379, a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1380, a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1381, a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1382, and a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1383.
35. The pharmaceutical compositions of claim 34, wherein the two or more monoclonal antibodies or antigen-binding fragments thereof are selected from: a. an antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising or consisting of the amino acid sequence of SEQ ID NO: 25, and a light chain variable domain (VL) comprising or consisting of the amino acid sequence of SEQ ID NO: 26; b. an antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising or consisting of the amino acid sequence of SEQ ID NO: 395, and a light chain variable domain (VL) comprising or consisting of the amino acid sequence of SEQ ID NO: 396; c. an antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising or consisting of the amino acid sequence of SEQ ID NO: 397, and a light chain variable domain (VL) comprising or consisting of the amino acid sequence of SEQ ID NO: 398; d. an antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising or consisting of the amino acid sequence of SEQ ID NO: 191, and a light chain variable domain (VL) comprising or consisting of the amino acid sequence of SEQ ID NO: 192; and e. an antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising or consisting of the amino acid sequence of SEQ ID NO: 219, and a light chain variable domain (VL) comprising or consisting of the amino acid sequence of SEQ ID NO: 220.
36. The pharmaceutical composition of any one of claims 33-35, wherein the pharmaceutical composition further comprises a pharmaceutically acceptable carrier.
37. The pharmaceutical composition of any one of claims 33-36, for use in treating a SARS- CoV-2 infection in a subject.
38. A method of treating a SARS-CoV-2 infection in a subject, comprising administering to the subject therapeutically effective amount of the antibody or antigen-binding fragment thereof of any one of claims 1-29 or a therapeutically effective amount of the pharmaceutical composition of any one of claims 33-36.
39. A method of neutralizing SARS-CoV-2 in a subject, comprising administering to a subject in need thereof a therapeutically effective amount of the antibody or antigen-binding fragment thereof of any one of claims 1-29 or a therapeutically effective amount of the pharmaceutical composition of any one of claims 33-36.
40. A method of preventing a SARS-CoV-2 infection in a subject, comprising administering to a subject in need thereof a therapeutically effective amount of the antibody or antigen-binding fragment thereof of any one of claims 1-29 or a therapeutically effective amount of the pharmaceutical composition of any one of claims 33-36.
41. The method of any one of claims 38-40, wherein the subject is immunocompromised.
42. The method of any one of claims 38-41, wherein the method further comprises administering a second therapeutic agent, preferably wherein the second therapeutic agent comprises an anti-inflammatory drug or an antiviral compound.
43. The method of claim 42, wherein: (a) the antiviral compound comprises: a nucleoside analog, a peptoid, an oligopeptide, a polypeptide, a protease inhibitor, a 3C-like protease inhibitor, a papain-like protease inhibitor, or an inhibitor of an RNA dependent RNA polymerase; or (b) the antiviral compound is selected from: acyclovir, ganciclovir, vidarabine, foscarnet, cidofovir, amantadine, ribavirin, trifluorothymidine, zidovudine, didanosine, zalcitabine, and an interferon, and preferably wherein the interferon is an interferon-Į^RU^DQ^LQWHUIHURQ-ȕ^
44. The method of claim 42 or claim 43, wherein the second therapeutic agent is administered before, after, or concurrently with the antibody or antigen-binding fragment thereof or the pharmaceutical composition thereof.
45. The method of any one of claims 38-40, wherein the pharmaceutical composition is administered to the subject after an exposure to SARS-CoV-2.
46. A kit for detecting a SARS-CoV-2 infection in a subject, comprising the antibody or antigen-binding fragment thereof of any one of claims 1-29.
47. A method of detecting the presence of a SARS-CoV-2 in a sample, comprising (1) contacting the sample with the antibody or antigen-binding fragment thereof of any one of claims 1-29, and (2) detecting the presence of an antibody-antigen complex, wherein the presence of the antibody-antigen complex indicates the presence of SARS-CoV-2.
48. The method of claim 47, wherein the antibody or antigen-binding fragment thereof is conjugated to a label, preferably wherein the label is selected from a fluorescent label, a chemiluminescent label, a radiolabel, and an enzyme.
49. The method of claim 47 or claim 48, further comprising contacting a secondary antibody with the antibody or antigen-binding fragment thereof, wherein the secondary antibody comprises a label.
50. The method of claim 49, further comprising detecting fluorescence or chemiluminescence of the label, preferably wherein the step of detecting the presence of an antibody-antigen complex comprises a competitive binding assay or ELISA.
51. The method of any one of claims 47-50, further comprising binding the sample to a solid support, preferably wherein the solid support is selected from microparticles, microbeads, magnetic beads, and an affinity purification column.
52. The method of any one of claims 47-51, wherein the sample is a blood sample, a nasal swab, or a throat swab.
53. A method of preparing an antibody or antigen-binding fragment thereof, comprising: a. obtaining the cultured host cell of claim 32; b. culturing the cultured host cell in a medium under conditions permitting expression of a polypeptide encoded by the vector and assembling of an antibody or antigen- binding fragment thereof; and c. purifying the antibody or fragment from the cultured cell or the medium of the cell.
54. A kit comprising a pharmaceutically acceptable dose unit of the antibody or antigen- binding fragment thereof of any one of claims 1-29 or the pharmaceutical composition of any one of claims 33-36.
55. A kit for the diagnosis, prognosis or monitoring treatment of SARS-CoV-2 in a subject, comprising: the antibody or antigen-binding fragment thereof of any one of claims 1-29; and a least one detection reagent that binds specifically to the antibody or antigen-binding fragment thereof.
56. A method of neutralizing SARS-CoV-2 in a subject, comprising administering to a subject in need thereof a therapeutically effective amount of two or more monoclonal antibodies or antigen-binding fragments thereof of any one of claims 1-29.
57. A method of treating a SARS-CoV-2 in a subject in need thereof, comprising administering to a subject in need thereof a therapeutically effective amount of two or more monoclonal antibodies or antigen-binding fragments thereof of any one of claims 1-29.
58. A method of preventing a SARS-CoV-2 in a subject in need thereof, comprising administering to a subject in need thereof a therapeutically effective amount of two or more monoclonal antibodies or antigen-binding fragments thereof of any one of claims 1-29.
59. The method of any one of claims 56-58, wherein the two or more monoclonal antibodies or antigen-binding fragments thereof are selected from: a. an antibody or antigen-binding fragment thereof comprising: a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1354, a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1355, a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1356, a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1357, a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1358, and a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1359; b. an antibody or antigen-binding fragment thereof comprising: a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1360, a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1361, a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1362, a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1363, a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1364, and a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1365; c. an antibody or antigen-binding fragment thereof comprising: a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1366, a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1367, a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1368, a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1369, a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1370, and a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1371; d. an antibody or antigen-binding fragment thereof comprising: a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1372, a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1373, a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1374, a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1375, a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1376, and a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1377; and e. an antibody or antigen-binding fragment thereof comprising: a HCDR1 comprising an amino acid sequence of SEQ ID NO: 1378, a HCDR2 comprising an amino acid sequence of SEQ ID NO: 1379, a HCDR3 comprising an amino acid sequence of SEQ ID NO: 1380, a LCDR1 comprising an amino acid sequence of SEQ ID NO: 1381, a LCDR2 comprising an amino acid sequence of SEQ ID NO: 1382, and a LCDR3 comprising an amino acid sequence of SEQ ID NO: 1383.
60. The method of claim 59, wherein the two or more monoclonal antibodies or antigen- binding fragments thereof are selected from: a. an antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising or consisting of the amino acid sequence of SEQ ID NO: 25, and a light chain variable domain (VL) comprising or consisting of the amino acid sequence of SEQ ID NO: 26; b. an antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising or consisting of the amino acid sequence of SEQ ID NO: 395, and a light chain variable domain (VL) comprising or consisting of the amino acid sequence of SEQ ID NO: 396; c. an antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising or consisting of the amino acid sequence of SEQ ID NO: 397, and a light chain variable domain (VL) comprising or consisting of the amino acid sequence of SEQ ID NO: 398; d. an antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising or consisting of the amino acid sequence of SEQ ID NO: 191, and a light chain variable domain (VL) comprising or consisting of the amino acid sequence of SEQ ID NO: 192; and e. an antibody or antigen-binding fragment thereof comprises a heavy chain variable domain (VH) comprising or consisting of the amino acid sequence of SEQ ID NO: 219, and a light chain variable domain (VL) comprising or consisting of the amino acid sequence of SEQ ID NO: 220.
61. The method of any one of claims 56-60, wherein the first antibody or antigen-binding fragment thereof and the second antibody or antigen-binding fragment thereof exhibit synergistic activity.
62. The method of any one of claims 56-61, further comprising administering to the subject a therapeutically effective amount of a second therapeutic agent or therapy.
63. The method of claim 62, wherein the second therapeutic agent comprises an anti- inflammatory drug or an antiviral compound.
64. The method of claim 63, wherein: (a) the antiviral compound comprises: a nucleoside analog, a peptoid, an oligopeptide, a polypeptide, a protease inhibitor, a 3C-like protease inhibitor, a papain-like protease inhibitor, or an inhibitor of an RNA dependent RNA polymerase; or (b) the antiviral compound is selected from: acyclovir, ganciclovir, vidarabine, foscarnet, cidofovir, amantadine, ribavirin, trifluorothymidine, zidovudine, didanosine, zalcitabine, and an interferon, and preferably wherein the interferon is an interferon-Į^RU^DQ^LQWHUIHURQ-ȕ^ 65. The method of any one of claims 56-64, wherein the first antibody or antigen-binding fragment thereof is administered before, after, or concurrently with the second antibody or antigen- binding fragment thereof. 66. The method of any one of claims 38-45 or 56-65, wherein the antibody or antigen-binding fragment thereof is administered to the subject intravenously, subcutaneously, or intraperitoneally.
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