SARS-CoV-2 Spike Glycoprotein Antibodies and Antigen-Binding Fragments

Abstract

The present disclosure provides antibodies and antigen-binding fragments thereof that bind specifically to a coronavirus spike protein and methods of using such antibodies and fragments for treating or preventing viral infections (e.g., coronavirus infections).

Description

SEQUENCE LISTING

[0001] An official copy of the sequence listing is submitted concurrently with the specification electronically via EFS-Web as an ASCII formatted sequence listing with a file name of "10753WO01-Sequence.txt", created on June 25, 2020, and having a size of 922,462 bytes. The sequence listing contained in this ASCII formatted document is part of the specification and is herein incorporated by reference in its entirety.FIELD OF THE INVENTION

[0002] The present invention relates to antibodies and antigen-binding fragments that bind specifically to coronavirus spike proteins and methods for treating or preventing coronavirus infections with said antibodies and fragments.BACKGROUND OF THE INVENTION

[0003] Newly identified viruses, such as coronaviruses, can be difficult to treat because they are not sufficiently characterized. The emergence of these newly identified viruses highlights the need for the development of novel antiviral strategies. Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a newly-emergent coronavirus which causes a severe acute respiratory disease, COVID-19. SARS-CoV-2 was first identified from an outbreak in Wuhan, China and as of March 20, 2020, the World Health Organization has reported 209,839 confirmed cases in 168 countries, areas, or territories, resulting in 8,778 deaths. Clinical features of COVID-19 include fever, dry cough, and fatigue, and the disease can cause respiratory failure resulting in death.

[0004] Thus far, there has been no vaccine or therapeutic agent to prevent or treat SARS-CoV-2 infection. In view of the continuing threat to human health, there is an urgent need for preventive and therapeutic antiviral therapies for SARS-CoV-2 control. Because this virus uses its spike glycoprotein for interaction with the cellular receptor ACE2 and the serine protease TMPRSS2 for entry into a target cell, this spike protein represents an attractive target for antibody therapeutics. In particular, fully human antibodies that specifically bind to the SARS-CoV-2-Spike protein (SARS-CoV-2-S) with high affinity and that inhibit virus infectivity could be important in the prevention and treatment of COVID-19.SUMMARY OF THE INVENTION

[0005] There is a need for neutralizing therapeutic anti-SARS-CoV-2-Spike protein (SARS-CoV-2-S) antibodies and their use for treating or preventing viral infection. The present disclosure addresses this need, in part, by providing human anti-SARS-CoV-2-S antibodies, such as those of Table 1, and combinations thereof including, for example, combinations with other therapeutics (e.g., anti-inflammatory agents, antimalarial agents, antiviral agents, or other antibodies or antigen-binding fragments), and methods of use thereof for treating viral infections.

[0006] The present disclosure provides neutralizing human antigen-binding proteins that specifically bind to SARS-CoV-2-S, for example, antibodies or antigen-binding fragments thereof.

[0007] In one aspect, the present disclosure provides an isolated recombinant antibody or antigen-binding fragment thereof that specifically binds to a coronavirus spike protein (CoV-S), wherein the antibody has one or more of the following characteristics: (a) binds to CoV-S with an EC 50 of less than about 10 -9< M; (b) demonstrates an increase in survival in a coronavirus-infected animal after administration to said coronavirus-infected animal, as compared to a comparable coronavirus-infected animal without said administration; and / or (c) comprises three heavy chain complementarity determining regions (CDRs) (CDR-H1, CDR-H2, and CDR-H3) contained within a heavy chain variable region (HCVR) comprising an amino acid sequence having at least about 90% sequence identity to an HCVR of Table 1; and three light chain CDRs (CDR-L1, CDR-L2, and CDR-L3) contained within a light chain variable region (LCVR) comprising an amino acid sequence having at least about 90% sequence identity to an LCVR Table 1.

[0008] In some embodiments, the antibody or antigen-binding fragment comprises: (a) an immunoglobulin heavy chain variable region comprising the CDR-H1, CDR-H2, and CDR-H3 of an antibody of Table 1; and / or (b) an immunoglobulin light chain variable region comprising the CDR-L1, CDR-L2, and CDR-L3 of an antibody of Table 1.

[0009] In some embodiments, the antibody or antigen-binding fragment comprises: (a) a heavy chain immunoglobulin variable region comprising an amino acid sequence having at least 90% amino acid sequence identity to an HCVR sequence of Table 1; and / or (b) a light chain immunoglobulin variable region comprising an amino acid sequence having at least 90% amino acid sequence identity to an LCVR sequence of Table 1.

[0010] In some embodiments, the antibody or antigen-binding fragment comprises the CDR-H1, CDR-H2, CDR-H3, CDR-L1, CDR-L2, and CDR-L3 of a single antibody of Table 1. In some embodiments, the antibody or antigen-binding fragment comprises an immunoglobulin that comprises the HCVR and the LCVR of a single antibody of Table 1.

[0011] In one aspect, the present disclosure provides an antigen-binding protein that competes with any one of the antibodies or antigen-binding fragments discussed above or herein for binding to CoV-S.

[0012] In one aspect, the present disclosure provides an antigen-binding protein that binds to the same epitope as, or to an overlapping epitope on, CoV-S as any one of the antibodies or antigen-binding fragments discussed above or herein.

[0013] In any of the various embodiments, the antibody or antigen-binding fragment may be multispecific.

[0014] In any of the various embodiments, the antibody or antigen-binding fragment may comprise one or more of the following properties: a) inhibits growth of coronavirus; b) binds to the surface of a coronavirus; c) limits spread of coronavirus infection of cells in vitro; and d) protects mice engineered to express the human ACE2 or TMPRSS2 protein from death and / or weight loss caused by coronavirus infection.

[0015] In any of the various embodiments, CoV-S is SARS-CoV-2-S.

[0016] In one aspect, the present disclosure provides a complex comprising an antibody or antigen-binding fragment as discussed above or herein bound to a CoV-S polypeptide. In some embodiments, the CoV-S is SARS-CoV-2-S.

[0017] In one aspect, the present disclosure provides a method for making an antibody or antigen-binding fragment as discussed above or herein, comprising: (a) introducing into a host cell one or more polynucleotides encoding said antibody or antigen-binding fragment; (b) culturing the host cell under conditions favorable to expression of the one or more polynucleotides; and (c) optionally, isolating the antibody or antigen-binding fragment from the host cell and / or a medium in which the host cell is grown. In some embodiments, the host cell is a Chinese hamster ovary cell.

[0018] In one aspect, the present disclosure provides an antibody or antigen-binding fragment that is a product of the method discussed above.

[0019] In one aspect, the present disclosure provides a polypeptide comprising: (a) CDR-H1, CDR-H2, and CDR-H3 of an HCVR domain of an antibody or antigen-binding fragment that comprises an HCVR amino acid sequence set forth in Table 1; or (b) CDR-L1, CDR-L2, and CDR-L3 of an LCVR domain of an immunoglobulin chain that comprises an LCVR amino acid sequence set forth in Table 1.

[0020] In one aspect, the present disclosure provides a polynucleotide encoding the polypeptide discussed above.

[0021] In one aspect, the present disclosure provides a vector comprising the polynucleotide discussed above.

[0022] In one aspect, the present disclosure provides a host cell comprising the antibody or antigen-binding fragment or polypeptide or polynucleotide or vector as discussed above or herein.

[0023] In one aspect, the present disclosure provides a composition or kit comprising the antibody or antigen-binding fragment discussed above or herein in association with a further therapeutic agent.

[0024] In one aspect, the present disclosure provides a pharmaceutical composition comprising the antigen-binding protein, antibody or antigen-binding fragment discussed above or herein and a pharmaceutically acceptable carrier and, optionally, a further therapeutic agent. In some embodiments, the further therapeutic agent is an anti-viral drug or a vaccine. In some embodiments, the further therapeutic agent is selected from the group consisting of: an anti-inflammatory agent, an antimalarial agent, an antibody or antigen-binding fragment thereof that specifically binds TMPRSS2, and an antibody or antigen-binding fragment thereof that specifically binds to CoV-S. In some cases, the antimalarial agent is chloroquine or hydroxychloroquine. In some cases, the anti-inflammatory agent is an antibody, such as sarilumab, tocilizumab, or gimsilumab. In some embodiments, the further therapeutic agent is a second antibody or antigen-binding fragment comprising HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3 sequences of Table 1.

[0025] In one aspect, the present disclosure provides a vessel or injection device comprising the antigen-binding protein, antibody or antigen-binding fragment, or composition as discussed above or herein.

[0026] In one aspect, the present disclosure provides a method for treating or preventing infection with a coronavirus, in a subject in need thereof, comprising administering a therapeutically effective amount of an antigen-binding protein, antibody or antigen-binding fragment as discussed above or herein. In some embodiments, the coronavirus is selected from the group consisting of SARS-CoV-2, SARS-CoV, and MERS-CoV.

[0027] In some embodiments of the method for treating or preventing infection with a coronavirus, the subject is administered one or more further therapeutic agents. In some cases, the one or more further therapeutic agents is an anti-viral drug or a vaccine. In some cases, the one or more further therapeutic agents is selected from the group consisting of: an anti-inflammatory agent, an antimalarial agent, an antibody or antigen-binding fragment thereof that specifically binds TMPRSS2, and an antibody or antigen-binding fragment thereof that specifically binds to CoV-S. In some cases, the antimalarial agent is chloroquine or hydroxychloroquine. In some cases, the anti-inflammatory agent is an antibody, such as for example, sarilumab, tocilizumab, or gimsilumab. In some embodiments, the further therapeutic agent is a second antibody or antigen-binding fragment comprising HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3 sequences of Table 1. Other antibodies that can be used alone or in combination with one another or with one or more of the antibodies disclosed herein for use in the context of the methods of the present disclosure include, e.g., LY-CoV555 (Eli Lilly); 47D11 (Wang et al Nature Communications Article No. 2251); B38, H4, B5 and / or H2 (Wu et al., 10.1126 / science.abc2241 (2020); STI-1499 (Sorrento Therapeutics); VIR-7831 and VIR-7832 (Vir Biotherapeutics).

[0028] In one aspect, the present disclosure provides a method for administering an antibody or antigen-binding fragment discussed above or herein into the body of a subject comprising injecting the antibody or antigen-binding fragment into the body of the subject. In some embodiments, the antibody or antigen-binding fragment is injected into the body of the subject subcutaneously, intravenously or intramuscularly.

[0029] In any of the various embodiments discussed above or herein, the antibody or antigen-binding binding fragment comprises a VH3-66 or Vk1-33 variable domain sequence.

[0030] In one aspect, the present disclosure provides an isolated antibody or antigen-binding fragment thereof that binds a SARS-CoV-2 spike protein comprising the amino acid sequence set forth in SEQ ID NO: 832, wherein said isolated antibody or antigen-binding fragment comprises three heavy chain complementarity determining regions (CDRs) (HCDR1, HCDR2 and HCDR3) contained within a heavy chain variable region (HCVR) comprising the amino acid sequence set forth in SEQ ID NO: 202, and three light chain complementarity determining regions (CDRs) (LCDR1, LCDR2 and LCDR3) contained within a light chain variable region (LCVR) comprising the amino acid sequence set forth in SEQ ID NO: 210.

[0031] In some embodiments, the HCDR1 comprises the amino acid sequence set forth in SEQ ID NO: 204, the HCDR2 comprises the amino acid sequence set forth in SEQ ID NO: 206, the HCDR3 comprises the amino acid sequence set forth in SEQ ID NO: 208, the LCDR1 comprises the amino acid sequence set forth in SEQ ID NO: 212, the LCDR2 comprises the amino acid sequence set forth in SEQ ID NO: 55, and the LCDR3 comprises the amino acid sequence set forth in SEQ ID NO: 214. In some embodiments, the isolated antibody or antigen-binding fragment thereof comprises an HCVR that comprises the amino acid sequence set forth in SEQ ID NO: 202. In some embodiments, the isolated antibody or antigen-binding fragment thereof comprises an LCVR that comprises the amino acid sequence set forth in SEQ ID NO: 210. In some embodiments, the isolated antibody or antigen-binding fragment thereof comprises an HCVR that comprises the amino acid sequence set forth in SEQ ID NO: 202 and an LCVR that comprises the amino acid sequence set forth in SEQ ID NO: 210.

[0032] In one aspect, the present disclosure provides an isolated antibody that binds a SARS-CoV-2 spike protein comprising the amino acid sequence set forth in SEQ ID NO: 832, wherein said isolated antibody comprises an immunoglobulin constant region, three heavy chain complementarity determining regions (CDRs) (HCDR1, HCDR2 and HCDR3) contained within a heavy chain variable region (HCVR) comprising the amino acid sequence set forth in SEQ ID NO: 202, and three light chain complementarity determining regions (CDRs) (LCDR1, LCDR2 and LCDR3) contained within a light chain variable region (LCVR) comprising the amino acid sequence set forth in SEQ ID NO: 210.

[0033] In some embodiments, the HCDR1 comprises the amino acid sequence set forth in SEQ ID NO: 204, the HCDR2 comprises the amino acid sequence set forth in SEQ ID NO: 206, the HCDR3 comprises the amino acid sequence set forth in SEQ ID NO: 208, the LCDR1 comprises the amino acid sequence set forth in SEQ ID NO: 212, the LCDR2 comprises the amino acid sequence set forth in SEQ ID NO: 55, and the LCDR3 comprises the amino acid sequence set forth in SEQ ID NO: 214. In some embodiments, the isolated antibody comprises an HCVR that comprises the amino acid sequence set forth in SEQ ID NO: 202 and an LCVR that comprises the amino acid sequence set forth in SEQ ID NO: 210. In some embodiments, the isolated antibody comprises a heavy chain comprising the amino acid sequence set forth in SEQ ID NO: 216 and a light chain comprising the amino acid sequence set forth in SEQ ID NO: 218. In some cases, the immunoglobulin constant region is an IgG1 constant region. In some cases, the isolated antibody is a recombinant antibody. In some cases, the isolated antibody is multispecific.

[0034] In one aspect, the present disclosure provides a pharmaceutical composition comprising an isolated antibody as discussed above or herein, and a pharmaceutically acceptable carrier or diluent.

[0035] In some embodiments, the pharmaceutical composition further comprises a second therapeutic agent. In some cases, the second therapeutic agent is selected from the group consisting of: a second antibody, or an antigen-binding fragment thereof, that binds a SARS-CoV-2 spike protein comprising the amino acid sequence set forth in SEQ ID NO: 832, an anti-inflammatory agent, an antimalarial agent, and an antibody or antigen-binding fragment thereof that binds TMPRSS2.

[0036] In some embodiments, the second therapeutic agent is a second antibody, or an antigen-binding fragment thereof, that binds a SARS-CoV-2 spike protein comprising the amino acid sequence set forth in SEQ ID NO: 832. In some cases, the second antibody or antigen-binding fragment thereof comprises three heavy chain CDRs (HCDR1, HCDR2 and HCDR3) contained within an HCVR comprising the amino acid sequence set forth in SEQ ID NO: 640, and three light chain CDRs (LCDR1, LCDR2 and LCDR3) contained within an LCVR comprising the amino acid sequence set forth in SEQ ID NO: 646. In some cases, the second antibody or antigen-binding fragment thereof comprises: HCDR1, comprising the amino acid sequence set forth in SEQ ID NO: 642; HCDR2, comprising the amino acid sequence set forth in SEQ ID NO: 499; HCDR3, comprising the amino acid sequence set forth in SEQ ID NO: 644; LCDR1, comprising the amino acid sequence set forth in SEQ ID NO: 648; LCDR2, comprising the amino acid sequence set forth in SEQ ID NO: 650; and LCDR3, comprising the amino acid sequence set forth in SEQ ID NO: 652. In some cases, the second antibody or antigen-binding fragment thereof comprises an HCVR comprising the amino acid sequence set forth in SEQ ID NO: 640 and an LCVR comprising the amino acid sequence set forth in SEQ ID NO: 646. In some cases, the second antibody or antigen-binding fragment thereof comprises a heavy chain comprising the amino acid sequence set forth in SEQ ID NO: 654 and a light chain comprising the amino acid sequence set forth in SEQ ID NO: 656.

[0037] In one aspect, the present disclosure provides an isolated antibody or antigen-binding fragment thereof that binds a SARS-CoV-2 spike protein comprising the amino acid sequence set forth in SEQ ID NO: 832, wherein said isolated antibody or antigen-binding fragment comprises three heavy chain complementarity determining regions (CDRs) (HCDR1, HCDR2 and HCDR3) contained within a heavy chain variable region (HCVR) comprising the amino acid sequence set forth in SEQ ID NO: 640, and three light chain complementarity determining regions (CDRs) (LCDR1, LCDR2 and LCDR3) contained within a light chain variable region (LCVR) comprising the amino acid sequence set forth in SEQ ID NO: 646.

[0038] In some embodiments, the HCDR1 comprises the amino acid sequence set forth in SEQ ID NO: 642, the HCDR2 comprises the amino acid sequence set forth in SEQ ID NO: 499, the HCDR3 comprises the amino acid sequence set forth in SEQ ID NO: 644, the LCDR1 comprises the amino acid sequence set forth in SEQ ID NO: 648, the LCDR2 comprises the amino acid sequence set forth in SEQ ID NO: 650, and the LCDR3 comprises the amino acid sequence set forth in SEQ ID NO: 652. In some embodiments, the isolated antibody or antigen-binding fragment thereof comprises an HCVR that comprises an amino acid sequence set forth in SEQ ID NO: 640. In some embodiments, the isolated antibody or antigen-binding fragment thereof comprises an LCVR that comprises an amino acid sequence set forth in SEQ ID NO: 646. In some embodiments, the isolated antibody or antigen-binding fragment thereof comprises an HCVR that comprises an amino acid sequence set forth in SEQ ID NO: 640 and an LCVR that comprises an amino acid sequence set forth in SEQ ID NO: 646.

[0039] In one aspect, the present disclosure provides an isolated antibody that binds a SARS-CoV-2 spike protein comprising the amino acid sequence set forth in SEQ ID NO: 832, wherein said isolated antibody comprises an immunoglobulin constant region, three heavy chain complementarity determining regions (CDRs) (HCDR1, HCDR2 and HCDR3) contained within a heavy chain variable region (HCVR) comprising the amino acid sequence set forth in SEQ ID NO: 640, and three light chain complementarity determining regions (CDRs) (LCDR1, LCDR2 and LCDR3) contained within a light chain variable region (LCVR) comprising the amino acid sequence set forth in SEQ ID NO: 646.

[0040] In some embodiments, the HCDR1 comprises the amino acid sequence set forth in SEQ ID NO: 642, the HCDR2 comprises the amino acid sequence set forth in SEQ ID NO: 499, the HCDR3 comprises the amino acid sequence set forth in SEQ ID NO: 644, the LCDR1 comprises the amino acid sequence set forth in SEQ ID NO: 648, the LCDR2 comprises the amino acid sequence set forth in SEQ ID NO: 650, and the LCDR3 comprises the amino acid sequence set forth in SEQ ID NO: 652. In some embodiments, the isolated antibody comprises an HCVR that comprises the amino acid sequence set forth in SEQ ID NO: 640 and an LCVR that comprises the amino acid sequence set forth in SEQ ID NO: 646. In some embodiments, the isolated antibody comprises a heavy chain comprising the amino acid sequence set forth in SEQ ID NO: 654 and a light chain comprising the amino acid sequence set forth in SEQ ID NO: 656. In some cases, the immunoglobulin constant region is an IgG1 constant region. In some cases, the isolated antibody is a recombinant antibody. In some cases, the isolated antibody is multispecific.

[0041] In one aspect, the present disclosure provides a pharmaceutical composition comprising an isolated antibody, as discussed above or herein, and a pharmaceutically acceptable carrier or diluent.

[0042] In some embodiments, the pharmaceutical composition further comprising a second therapeutic agent. In some cases, the second therapeutic agent is selected from the group consisting of: a second antibody, or an antigen-binding fragment thereof, that binds a SARS-CoV-2 spike protein comprising the amino acid sequence set forth in SEQ ID NO: 832, an anti-inflammatory agent, an antimalarial agent, and an antibody or antigen-binding fragment thereof that binds TMPRSS2.

[0043] In some embodiments, the second therapeutic agent is a second antibody, or an antigen-binding fragment thereof, that binds a SARS-CoV-2 spike protein comprising the amino acid sequence set forth in SEQ ID NO: 832. In some cases, the second antibody or antigen-binding fragment thereof comprises three heavy chain CDRs (HCDR1, HCDR2 and HCDR3) contained within an HCVR comprising the amino acid sequence set forth in SEQ ID NO: 202, and three light chain CDRs (LCDR1, LCDR2 and LCDR3) contained within an LCVR comprising the amino acid sequence set forth in SEQ ID NO: 210. In some cases, the second antibody or antigen-binding fragment thereof comprises: HCDR1, comprising the amino acid sequence set forth in SEQ ID NO: 204; HCDR2, comprising the amino acid sequence set forth in SEQ ID NO: 206; HCDR3, comprising the amino acid sequence set forth in SEQ ID NO: 208 LCDR1, comprising the amino acid sequence set forth in SEQ ID NO: 212; LCDR2, comprising the amino acid sequence set forth in SEQ ID NO: 55; and LCDR3, comprising the amino acid sequence set forth in SEQ ID NO: 214. In some cases, the second antibody or antigen-binding fragment thereof comprises an HCVR comprising the amino acid sequence set forth in SEQ ID NO: 202 and an LCVR comprising the amino acid sequence set forth in SEQ ID NO: 210. In some cases, the second antibody or antigen-binding fragment thereof comprises a heavy chain comprising the amino acid sequence set forth in SEQ ID NO: 216 and a light chain comprising the amino acid sequence set forth in SEQ ID NO: 218.

[0044] In various embodiments, any of the features or components of embodiments discussed above or herein may be combined, and such combinations are encompassed within the scope of the present disclosure. Any specific value discussed above or herein may be combined with another related value discussed above or herein to recite a range with the values representing the upper and lower ends of the range, and such ranges are encompassed within the scope of the present disclosure.BRIEF DESCRIPTION OF THE FIGURES

[0045] Figure 1 shows ELISA blocking data for selected anti-SARS-CoV-2-S antibodies against SARS-CoV-2 spike protein, preventing the spike protein from binding to its receptor ACE2. Figure 2 shows ELISA blocking data for selected anti-SARS-CoV-2-S antibodies against SARS-CoV-2 spike protein, preventing the spike protein from binding to its receptor ACE2. Figure 3 shows ELISA blocking data for selected anti-SARS-CoV-2-S antibodies against SARS-CoV-2 spike protein, preventing the spike protein from binding to its receptor ACE2. Figure 4 shows ELISA blocking data for selected anti-SARS-CoV-2-S antibodies against SARS-CoV-2 spike protein, preventing the spike protein from binding to its receptor ACE2. Figure 5 shows ELISA blocking data for selected anti-SARS-CoV-2-S antibodies against SARS-CoV-2 spike protein, preventing the spike protein from binding to its receptor ACE2. Figure 6 shows ELISA blocking data for selected anti-SARS-CoV-2-S antibodies against SARS-CoV-2 spike protein, preventing the spike protein from binding to its receptor ACE2. Figure 7 shows ELISA blocking data for selected anti-SARS-CoV-2-S antibodies against SARS-CoV-2 spike protein, preventing the spike protein from binding to its receptor ACE2. Figure 8 shows ELISA blocking data for selected anti-SARS-CoV-2-S antibodies against SARS-CoV-2 spike protein, preventing the spike protein from binding to its receptor ACE2. Figure 9A and Figure 9B display V gene frequencies for paired Heavy (X-axis) and Light (Y-axis) chains of isolated neutralizing antibodies to SARS-CoV-2 for VelocImmune® mice (Figure 9A; N=185) and convalescent human donors (Figure 9B; N=68). The shade and size of the circle corresponds to the number of Heavy and Light chain pairs present in the repertoires of isolated neutralizing antibodies. Neutralization is defined as >70% with 1:4 dilution of antibody (∼2 µg / ml) in VSV pseudoparticle neutralization assay. Figure 10A and Figure 10B display neutralization potency. Figure 10A displays the neutralization potency of anti-SARS-CoV-2 Spike mAbs. Serial dilutions of anti-Spike mAbs, IgG1 isotype control, and recombinant dimeric ACE2 (hACE2.hFc) were added with pVSV-SARS-CoV-2-S-mNeon to Vero cells and mNeon expression was measured 24 hours post-infection as a read-out for virus infectivity. Data is graphed as percent neutralization relative to virus only infection control. Figure 10B displays neutralization potency of individual anti-Spike mAbs and combinations of mAbs against SARS-CoV-2-S virus in VeroE6 cells. Figure 11 displays epitope bin analysis from a matrix of pre-mix binding assays for different anti-SARS-CoV-2 mAbs. Epitope binning was performed against nine anti-SARS-CoV-2 mAb as described. There were three phases (I, II, and III) for each graph. In phase I anti-SARS-CoV-2 mAb (20ug / ml) was loaded to the anti-human Fc probe. In phase II human IgG1 blocking mAb solution (100ug / ml). In phase III a solution of 100nM SARS CoV-2 RBD-MMH pre-mix complex of each 600 nM anti-SARS-CoV-2 mAb binding site flowed over the mAb capture probe. Figure 12 displays a 3D surface model for the structure of the Spike protein RBD domain showing the ACE2 interface and HDX-MS epitope mapping results. RBD residues protected by anti-SARS-CoV2-Spike antibodies are indicated with shading that represent the extent of protection as determined by HDX-MS experiments. The RBD structure is reproduced from PDB 6M17. Figure 13A and Figure 13B display a complex of mAb10933 and mAb10987 with the SARS-CoV-2 RBD. Figure 13A displays a 3.9 Å cryoEM map of mAb10933 + RBD + mAb10987 complex, shaded according to the chains in the refined model of Figure 13B.RBD, mAb 10933 heavy and light chains, and mAb10987 heavy and light chain are identified. Figure 14 displays cryoEM data statistics. Data collection and refinement statistics are reported for the mAb10987 + mAb10933 + SARS-CoV-2 RBD complex structure shown in Figure 13A and Figure 13B. DETAILED DESCRIPTION OF THE INVENTION

[0046] Before the present methods are described, it is to be understood that this invention is not limited to particular methods, and experimental conditions described, as such methods and conditions may vary. It is also to be understood that the terminology used herein is for the purpose of describing particular embodiments only, and is not intended to be limiting, since the scope of the present invention will be limited only by the appended claims.

[0047] Unless defined otherwise, 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 invention belongs. Although any methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present invention, preferred methods and materials are now described. All publications mentioned herein are incorporated herein by reference in their entirety.

[0048] The term "coronavirus" or "CoV" refers to any virus of the coronavirus family, including but not limited to SARS-CoV-2, MERS-CoV, and SARS-CoV. SARS-CoV-2 refers to the newly-emerged coronavirus which was identified as the cause of a serious outbreak starting in Wuhan, China, and which is rapidly spreading to other areas of the globe. SARS-CoV-2 has also been known as 2019-nCoV and Wuhan coronavirus. It binds via the viral spike protein to human host cell receptor angiotensin-converting enzyme 2 (ACE2). The spike protein also binds to and is cleaved by TMPRSS2, which activates the spike protein for membrane fusion of the virus.

[0049] The term "CoV-S", also called "S" or "S protein" refers to the spike protein of a coronavirus, and can refer to specific S proteins such as SARS-CoV-2-S, MERS-CoV S, and SARS-CoV S. The SARS-CoV-2-Spike protein is a 1273 amino acid type I membrane glycoprotein which assembles into trimers that constitute the spikes or peplomers on the surface of the enveloped coronavirus particle. The protein has two essential functions, host receptor binding and membrane fusion, which are attributed to the N-terminal (S1) and C-terminal (S2) halves of the S protein. CoV-S binds to its cognate receptor via a receptor binding domain (RBD) present in the S1 subunit. The amino acid sequence of full-length SARS-CoV-2 spike protein is exemplified by the amino acid sequence provided in SEQ ID NO: 832. The term "CoV-S" includes protein variants of CoV spike protein isolated from different CoV isolates as well as recombinant CoV spike protein or a fragment thereof. The term also encompasses CoV spike protein or a fragment thereof coupled to, for example, a histidine tag, mouse or human Fc, or a signal sequence such as ROR1.

[0050] The term "coronavirus infection" or "CoV infection," as used herein, refers to infection with a coronavirus such as SARS-CoV-2, MERS-CoV, or SARS-CoV. The term includes coronavirus respiratory tract infections, often in the lower respiratory tract. Symptoms can include high fever, dry cough, shortness of breath, pneumonia, gastro-intestinal symptoms such as diarrhea, organ failure (kidney failure and renal dysfunction), septic shock, and death in severe cases.Viruses

[0051] The present invention includes methods for treating or preventing a viral infection in a subject. The term "virus" includes any virus whose infection in the body of a subject is treatable or preventable by administration of an anti-CoV-S antibody or antigen-binding fragment thereof (e.g., wherein infectivity of the virus is at least partially dependent on CoV-S). In an embodiment of the invention, a "virus" is any virus that expresses spike protein (e.g., CoV-S). The term "virus" also includes a CoV-S-dependent respiratory virus which is a virus that infects the respiratory tissue of a subject (e.g., upper and / or lower respiratory tract, trachea, bronchi, lungs) and is treatable or preventable by administration of an anti-CoV-S antibody or antigen-binding fragment thereof. For example, in an embodiment of the invention, virus includes coronavirus, SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2), SARS-CoV (severe acute respiratory syndrome coronavirus), and MERS-CoV (Middle East respiratory syndrome (MERS) coronavirus). Coronaviruses can include the genera of alphacoronaviruses, betacoronaviruses, gammacoronaviruses, and deltacoronaviruses. In some embodiments, the antibodies or antigen-binding fragments provided herein can bind to and / or neutralize an alphacoronavirus, a betacoronavirus, a gammacoronavirus, and / or a deltacoronavirus. In certain embodiments, this binding and / or neutralization can be specific for a particular genus of coronavirus or for a particular subgroup of a genus. "Viral infection" refers to the invasion and multiplication of a virus in the body of a subject.

[0052] Coronavirus virions are spherical with diameters of approximately 125 nm. The most prominent feature of coronaviruses is the club-shape spike projections emanating from the surface of the virion. These spikes are a defining feature of the virion and give them the appearance of a solar corona, prompting the name, coronaviruses. Within the envelope of the virion is the nucleocapsid. Coronaviruses have helically symmetrical nucleocapsids, which is uncommon among positive-sense RNA viruses, but far more common for negative-sense RNA viruses. SARS-CoV-2, MERS-CoV, and SARS-CoV belong to the coronavirus family. The initial attachment of the virion to the host cell is initiated by interactions between the S protein and its receptor. The sites of receptor binding domains (RBD) within the S1 region of a coronavirus S protein vary depending on the virus, with some having the RBD at the C-terminus of S1. The S-protein / receptor interaction is the primary determinant for a coronavirus to infect a host species and also governs the tissue tropism of the virus. Many coronaviruses utilize peptidases as their cellular receptor. Following receptor binding, the virus must next gain access to the host cell cytosol. This is generally accomplished by acid-dependent proteolytic cleavage of S protein by a cathepsin, TMPRRS2 or another protease, followed by fusion of the viral and cellular membranes.Anti-CoV-S Antibodies and Antigen-Binding Fragments

[0053] The present invention provides antigen-binding proteins, such as antibodies and antigen-binding fragments thereof, that specifically bind to CoV spike protein or an antigenic fragment thereof.

[0054] The term "antibody", as used herein, refers to immunoglobulin molecules comprising four polypeptide chains, two heavy chains (HCs) and two light chains (LCs) inter-connected by disulfide bonds (i.e., "full antibody molecules"), as well as multimers thereof (e.g. IgM). Exemplary antibodies include, for example, those listed in Table 1. Each heavy chain comprises a heavy chain variable region ("HCVR" or "V H ") and a heavy chain constant region (comprised of domains C H 1, C H 2 and C H 3). Each light chain is comprised of a light chain variable region ("LCVR or "V L ") and a light chain constant region (C L ). The V H and V L 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 V H and V L comprises three CDRs and four FRs, arranged from amino-terminus to carboxy-terminus in the following order: FR1, CDR1, FR2, CDR2, FR3, CDR3, FR4. Heavy chain CDRs can also be referred to as HCDRs or CDR-Hs, and numbered as described above (e.g., HCDR1, HCDR2, and HCDR3 or CDR-H1, CDR-H2, and CDR-H3). Likewise, light chain CDRs can be referred to as LCDRs or CDR-Ls, and numbered LCDR1, LCDR2, and LCDR3, or CDR-L1, CDR-L2, and CDR-L3. In certain embodiments of the invention, the FRs of the antibody (or antigen binding fragment thereof) are identical to the human germline sequences, or are naturally or artificially modified. Exemplary human germline sequences include, but are not limited to, VH3-66 and Vk1-33. Thus, the present disclosure provides anti-CoV-S antibodies or antigen-binding fragments thereof (e.g., anti-SARS-CoV-2-S antibodies or antigen-binding fragments thereof) comprising HCDR and LCDR sequences of Table 1 within a VH3-66 or Vk1-33 variable heavy chain or light chain region. The present disclosure further provides anti-CoV-S antibodies or antigen-binding fragments thereof (e.g., anti-SARS-CoV-2-S antibodies or antigen-binding fragments thereof) comprising HCDR and LCDR sequences of Table 1 within a combination of a light chain selected from IgKV4-1, IgKV 1-5, IgKV1-9, IgKV1-12, IgKV3-15, IgKV1-16, IgKV1-17, IgKV3-20, IgLV3-21, IgKV2-24, IgKV1-33, IgKV1-39, IgLV1-40, IgLV1-44, IgLV1-51, IgLV3-1, IgKV1-6, IgLV2-8, IgKV3-11, IgLV2-11, IgLV2-14, IgLV2-23, or IgLV6-57, and a heavy chain selected from IgHV1-69, IgHV3-64, IgHV4-59, IgHV3-53, IgHV3-48, IgHV4-34, IgHV3-33, IgHV3-30, IgHV3-23, IgHV3-20, IgHV1-18, IgHV3-15, IgHV3-11, IgHV3-9, IgHV1-8, IgHV3-7, IgHV2-5, IgHV1-2, IgHV2-70, IgHV3-66, IgHV5-51, IgHV1-46, IgHV4-39, IgHV4-31, IgHV3-30-3, IgHV2-26, or IgHV7-4-1. The present disclosure further provides anti-CoV-S antibodies or antigen-binding fragments thereof (e.g., anti-SARS-CoV-2-S antibodies or antigen-binding fragments thereof) comprising HCVR and LCVR sequences of Table 1 within a combination of a light chain selected from IgKV4-1, IgKV 1-5, IgKV1-9, IgKV1-12, IgKV3-15, IgKV1-16, IgKV1-17, IgKV3-20, IgLV3-21, IgKV2-24, IgKV1-33, IgKV1-39, IgLV1-40, IgLV1-44, IgLV1-51, IgLV3-1, IgKV1-6, IgLV2-8, IgKV3-11, IgLV2-11, IgLV2-14, IgLV2-23, or IgLV6-57, and a heavy chain selected from IgHV1-69, IgHV3-64, IgHV4-59, IgHV3-53, IgHV3-48, IgHV4-34, IgHV3-33, IgHV3-30, IgHV3-23, IgHV3-20, IgHV1-18, IgHV3-15, IgHV3-11, IgHV3-9, IgHV1-8, IgHV3-7, IgHV2-5, IgHV1-2, IgHV2-70, IgHV3-66, IgHV5-51, IgHV1-46, IgHV4-39, IgHV4-31, IgHV3-30-3, IgHV2-26, or IgHV7-4-1.

[0055] Typically, the variable domains of both the heavy and light immunoglobulin chains comprise three hypervariable regions, also called complementarity determining regions (CDRs), located within relatively conserved framework regions (FR). In general, from N-terminal to C-terminal, both light and heavy chains variable domains comprise FR1, CDR1, FR2, CDR2, FR3, CDR3 and FR4. In an embodiment of the invention, the assignment of amino acids to each domain is in accordance with the definitions of Sequences of Proteins of Immunological Interest, Kabat, et al.; National Institutes of Health, Bethesda, Md.; 5th ed.; NIH Publ. No. 91-3242 (1991); Kabat (1978) Adv. Prot. Chem. 32:1-75; Kabat, et al., (1977) J. Biol. Chem. 252:6609-6616; Chothia, et al., (1987) J Mol. Biol. 196:901-917 or Chothia, et al., (1989) Nature 342:878-883.

[0056] The present invention includes monoclonal anti-CoV-S antigen-binding proteins, e.g., antibodies and antigen-binding fragments thereof, as well as monoclonal compositions comprising a plurality of isolated monoclonal antigen-binding proteins. The term "monoclonal antibody", as used herein, refers to a population of substantially homogeneous antibodies, i.e., the antibody molecules comprising the population are identical in amino acid sequence except for possible naturally occurring mutations that may be present in minor amounts. A "plurality" of such monoclonal antibodies and fragments in a composition refers to a concentration of identical (i.e., as discussed above, in amino acid sequence except for possible naturally occurring mutations that may be present in minor amounts) antibodies and fragments which is above that which would normally occur in nature, e.g., in the blood of a host organism such as a mouse or a human.

[0057] In an embodiment of the invention, an anti-CoV-S antigen-binding protein, e.g., antibody or antigen-binding fragment comprises a heavy chain constant domain, e.g., of the type IgA (e.g., IgA1 or IgA2), IgD, IgE, IgG (e.g., IgG1, IgG2, IgG3 and IgG4) or IgM. In an embodiment of the invention, an antigen-binding protein, e.g., antibody or antigen-binding fragment comprises a light chain constant domain, e.g., of the type kappa or lambda.

[0058] The term "human" antigen-binding protein, such as an antibody, as used herein, includes antibodies having variable and constant regions derived from human germline immunoglobulin sequences whether in a human cell or grafted into a non-human cell, e.g., a mouse cell. See e.g., US8502018, US6596541 or US5789215. The human mAbs of the invention may 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), for example in the CDRs and, in particular, CDR3. However, the term "human antibody", as used herein, is not intended to include mAbs in which CDR sequences derived from the germline of another mammalian species (e.g., mouse) have been grafted onto human FR sequences. The term includes antibodies recombinantly produced in a non-human mammal or in cells of a non-human mammal. The term is not intended to include antibodies isolated from or generated in a human subject. See below.

[0059] The present invention includes anti-CoV-S chimeric antigen-binding proteins, e.g., antibodies and antigen-binding fragments thereof, and methods of use thereof. As used herein, a "chimeric antibody" is an antibody having the variable domain from a first antibody and the constant domain from a second antibody, where the first and second antibodies are from different species. (US4816567; and Morrison et al., (1984) Proc. Natl. Acad. Sci. USA 81: 6851-6855).

[0060] The present invention includes anti-CoV-S hybrid antigen-binding proteins, e.g., antibodies and antigen-binding fragments thereof, and methods of use thereof. As used herein, a "hybrid antibody" is an antibody having the variable domain from a first antibody and the constant domain from a second antibody, wherein the first and second antibodies are from different animals, or wherein the variable domain, but not the constant region, is from a first animal. For example, a variable domain can be taken from an antibody isolated from a human and expressed with a fixed constant region not isolated from that antibody. Exemplary hybrid antibodies are described in Example 1, which refers to antibody heavy chain variable region and light chain variable region derived PCR products that were cloned into expression vectors containing a heavy constant region and a light constant region, respectively. Hybrid antibodies are synthetic and non-natrually occurring because the variable and constant regions they contain are not isolated from a single natural source.

[0061] The term "recombinant" antigen-binding proteins, such as antibodies or antigen-binding fragments thereof, refers to such molecules 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 includes 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 a non-human cell expression system, or isolated from a recombinant combinatorial human antibody library. In some embodiments, a recombinant antibody shares a sequence with an antibody isolated from an organism (e.g., a mouse or a human), but has been expressed via recombinant DNA technology. Such antibodies may have post-translational modifications (e.g., glycosylation) that differ from the antibody as isolated from the organism.

[0062] Recombinant anti-CoV-S antigen-binding proteins, e.g., antibodies and antigen-binding fragments, disclosed herein may also be produced in an E. coli / T7 expression system. In this embodiment, nucleic acids encoding the anti-CoV-S antibody immunoglobulin molecules of the invention (e.g., as found in Table 1) may be inserted into a pET-based plasmid and expressed in the E. coli / T7 system. For example, the present invention includes methods for expressing an antibody or antigen-binding fragment thereof or immunoglobulin chain thereof in a host cell (e.g., bacterial host cell such as E. coli such as BL21 or BL21DE3) comprising expressing T7 RNA polymerase in the cell which also includes a polynucleotide encoding an immunoglobulin chain that is operably linked to a T7 promoter. For example, in an embodiment of the invention, a bacterial host cell, such as an E. coli, includes a polynucleotide encoding the T7 RNA polymerase gene operably linked to a lac promoter and expression of the polymerase and the chain is induced by incubation of the host cell with IPTG (isopropyl-beta-D-thiogalactopyranoside). See US4952496 and US5693489 or Studier & Moffatt, Use of bacteriophage T7 RNA polymerase to direct selective high-level expression of cloned genes, J. Mol. Biol. 1986 May 5;189(1): 113-30.

[0063] There are several methods by which to produce recombinant antibodies which are known in the art. One example of a method for recombinant production of antibodies is disclosed in US4816567.

[0064] Transformation can be by any known method for introducing polynucleotides (e.g., DNA or RNA, including mRNA) into a host cell. Methods for introduction of heterologous polynucleotides into mammalian cells are well known in the art and include dextran-mediated transfection, calcium phosphate precipitation, polybrene-mediated transfection, protoplast fusion, electroporation, encapsulation of the polynucleotide(s) in liposomes, lipid nanoparticle technology, biolistic injection and direct microinjection of the DNA into nuclei. In addition, nucleic acid molecules may be introduced into mammalian cells by viral vectors such as lentivirus or adeno-associated virus. Methods of transforming cells are well known in the art. See, for example, U.S. Pat. Nos. 4,399,216; 4,912,040; 4,740,461 and 4,959,455. In some embodiments, an antibody or antigen-binding fragment thereof of the present disclosure can be introduced to a subject in nucleic acid form (e.g, DNA or RNA, including mRNA), such that the subject's own cells produce the antibody. The present disclosure further provides modifications to nucleotide sequences encoding the anti-CoV-S antibodies described herein that result in increased antibody expression, increased antibody stability, increased nucleic acid (e.g., mRNA) stability, or improved affinity or specificity of the antibodies for the CoV spike protein.

[0065] Thus, the present invention includes recombinant methods for making an anti-CoV-S antigen-binding protein, such as an antibody or antigen-binding fragment thereof of the present invention, or an immunoglobulin chain thereof, comprising (i) introducing one or more polynucleotides (e.g., including the nucleotide sequence of any one or more of the sequences of Table 2) encoding light and / or heavy immunoglobulin chains, or CDRs, of the antigen-binding protein, e.g., of Table 1, for example, wherein the polynucleotide is in a vector; and / or integrated into a host cell chromosome and / or is operably linked to a promoter; (ii) culturing the host cell (e.g., CHO or Pichia or Pichia pastoris) under condition favorable to expression of the polynucleotide and, (iii) optionally, isolating the antigen-binding protein, (e.g., antibody or fragment) or chain from the host cell and / or medium in which the host cell is grown. For example, a polynucleotide can be integrated into a host cell chromosome through targeted insertion with a vector such as adeno-associated virus (AAV), e.g., after cleavage of the chromosome using a gene editing system (e.g., CRISPR (for example, CRISPR-Cas9), TALEN, megaTAL, zinc finger, or Argonaute). Targeted insertions can take place, for example, at host cell loci such as an albumin or immunoglopbulin genomic locus. Alternatively, insertion can be at a random locus, e.g., using a vector such as lentivirus. When making an antigen-binding protein (e.g., antibody or antigen-binding fragment) comprising more than one immunoglobulin chain, e.g., an antibody that comprises two heavy immunoglobulin chains and two light immunoglobulin chains, co-expression of the chains in a single host cell leads to association of the chains, e.g., in the cell or on the cell surface or outside the cell if such chains are secreted, so as to form the antigen-binding protein (e.g., antibody or antigen-binding fragment). The methods include those wherein only a heavy immunoglobulin chain or only a light immunoglobulin chain (e.g., any of those discussed herein including mature fragments and / or variable domains thereof) is expressed. Such chains are useful, for example, as intermediates in the expression of an antibody or antigen-binding fragment that includes such a chain. For example, the present invention also includes anti-CoV-S antigen-binding proteins, such as antibodies and antigen-binding fragments thereof, comprising a heavy chain immunoglobulin (or variable domain thereof or comprising the CDRs thereof) encoded by a polynucleotide comprising a nucleotide sequence set forth in Table 2 and a light chain immunoglobulin (or variable domain thereof or comprising the CDRs thereof) encoded by a nucleotide sequence set forth in Table 2 which are the product of such production methods, and, optionally, the purification methods set forth herein. For example, in some embodiments, the product of the method is an anti-CoV-S antigen-binding protein which is an antibody or fragment comprising an HCVR comprising an amino acid sequence set forth in Table 1 and an LCVR comprising an amino acid sequence set forth in Table 1, wherein the HCVR and LCVR sequences are selected from a single antibody listed in Table 1. In some embodiments, the product of the method is an anti-CoV-S antigen-binding protein which is an antibody or fragment comprising HCDR1, HCDR2, and HCDR3 comprising amino acid sequences set forth in Table 1 and LCDR1, LCDR2, and LCDR3 comprising amino acid sequences set forth in Table 1, wherein the six CDR sequences are selected from a single antibody listed in Table 1. In some embodiments, the product of the method is an anti-CoV-S antigen-binding protein which is an antibody or fragment comprising a heavy chain comprising an HC amino acid sequence set forth in Table 1 and a light chain comprising an LC amino acid sequence set forth in Table 1.

[0066] Eukaryotic and prokaryotic host cells, including mammalian cells, may be used as hosts for expression of an anti-CoV-S antigen-binding protein. Such host cells are well known in the art and many are available from the American Type Culture Collection (ATCC). These host cells include, inter alia, Chinese hamster ovary (CHO) cells, NS0, SP2 cells, HeLa cells, baby hamster kidney (BHK) cells, monkey kidney cells (COS), human hepatocellular carcinoma cells (e.g., Hep G2), A549 cells, 3T3 cells, HEK-293 cells and a number of other cell lines. Mammalian host cells include human, mouse, rat, dog, monkey, pig, goat, bovine, horse and hamster cells. Other cell lines that may be used are insect cell lines (e.g., Spodopterafrugiperda or Trichoplusia ni), amphibian cells, bacterial cells, plant cells and fungal cells. Fungal cells include yeast and filamentous fungus cells including, for example, Pichiapastoris, Pichia finlandica, Pichia trehalophila, Pichia koclamae, Pichia membranaefaciens, Pichia minuta (Ogataea minuta, Pichia lindneri), Pichia opuntiae, Pichia thermotolerans, Pichia salictaria, Pichia guercuum, Pichia pijperi, Pichia stiptis, Pichia methanolica, Pichia sp., Saccharomyces cerevisiae, Saccharomyces sp., Hansenula polymorpha, Kluyveromyces sp., Kluyveromyces lactis, Candida albicans, Aspergillus nidulans, Aspergillus niger, Aspergillus oryzae, Trichoderma reesei, Chrysosporium lucknowense, Fusarium sp., Fusarium gramineum, Fusarium venenatum, Physcomitrella patens and Neurospora crassa. The present invention includes an isolated host cell (e.g., a CHO cell) comprising an antigen-binding protein, such as those of Table 1; or a polynucleotide encoding such a polypeptide thereof.

[0067] The term "specifically binds" refers to those antigen-binding proteins (e.g., mAbs) having a binding affinity to an antigen, such as a CoV-S protein (e.g., SARS-CoV-2-S), expressed as K D , of at least about 10 -8< M, as measured by real-time, label free bio-layer interferometry assay, for example, at 25°C or 37°C, e.g., an Octet® HTX biosensor, or by surface plasmon resonance, e.g., BIACORE™, or by solution-affinity ELISA. The present invention includes antigen-binding proteins that specifically bind to a CoV-S protein.

[0068] The terms "antigen-binding portion" or "antigen-binding fragment" of an antibody or antigen-binding protein, and the like, as used herein, include any naturally occurring, enzymatically obtainable, synthetic, or genetically engineered polypeptide or glycoprotein that specifically binds an antigen to form a complex. Non-limiting examples of antigen-binding fragments include: (i) Fab fragments; (ii) F(ab') 2 fragments; (iii) Fd fragments; (iv) Fv fragments; (v) single-chain Fv (scFv) molecules; (vi) dAb fragments; and (vii) minimal recognition units consisting of the amino acid residues that mimic the hypervariable region of an antibody (e.g., an isolated complementarity determining region (CDR) such as a CDR3 peptide), or a constrained FR3-CDR3-FR4 peptide. Other engineered molecules, such as domain-specific antibodies, single domain antibodies, domain-deleted antibodies, chimeric antibodies, CDR-grafted antibodies, diabodies, triabodies, tetrabodies, minibodies, nanobodies (e.g., as defined in WO08 / 020079 or WO09 / 138519) (e.g., monovalent nanobodies, bivalent nanobodies, etc.), small modular immunopharmaceuticals (SMIPs), and shark variable IgNAR domains, are also encompassed within the expression "antigen-binding fragment," as used herein. In an embodiment of the invention, the antigen-binding fragment comprises three or more CDRs of an antibody of Table 1 (e.g., CDR-H1, CDR-H2 and CDR-H3; or CDR-L1, CDR-L2 and CDR-L3).

[0069] An antigen-binding fragment of an antibody will, in an embodiment of the invention, comprise at least one variable domain. The variable domain may be of any size or amino acid composition and will generally comprise at least one CDR, which is adjacent to or in frame with one or more framework sequences. In antigen-binding fragments having a V H domain associated with a V L domain, the V H and V L domains may be situated relative to one another in any suitable arrangement. For example, the variable region may be dimeric and contain V H - V H , V H - V L or V L - V L dimers. Alternatively, the antigen-binding fragment of an antibody may contain a monomeric V H or V L domain.

[0070] In certain embodiments, an antigen-binding fragment of an antibody may contain at least one variable domain covalently linked to at least one constant domain. Non-limiting, exemplary configurations of variable and constant domains that may be found within an antigen-binding fragment of an antibody of the present invention include: (i) V H -C H 1; (ii) V H -C H 2; (iii) V H -C H 3; (iv) V H -C H 1-C H 2; (v) V H -C H 1-C H 2-C H 3; (vi) V H -C H 2-C H 3; (vii) V H -C L ; (viii) V L -C H 1; (ix) V L -C H 2; (x) V L -C H 3; (xi) V L -C H 1-C H 2; (xii) V L -C H 1-C H 2-C H 3; (xiii) V L -C H 2-C H 3; and (xiv) V L -C L . In any configuration of variable and constant domains, including any of the exemplary configurations listed above, the variable and constant domains may be either directly linked to one another or may be linked by a full or partial hinge or linker region. A hinge region may consist of at least 2 (e.g., 5, 10, 15, 20, 40, 60 or more) amino acids, which result in a flexible or semi-flexible linkage between adjacent variable and / or constant domains in a single polypeptide molecule. Moreover, an antigen-binding fragment of an antibody of the present invention may comprise a homo-dimer or hetero-dimer (or other multimer) of any of the variable and constant domain configurations listed above in non-covalent association with one another and / or with one or more monomeric V H or V L domain (e.g., by disulfide bond(s)).

[0071] Antigen-binding proteins (e.g., antibodies and antigen-binding fragments) may be mono-specific or multi-specific (e.g., bi-specific). Multispecific antigen-binding proteins are discussed further herein.

[0072] In specific embodiments, antibody or antibody fragments of the invention may be conjugated to a moiety such a ligand or a therapeutic moiety ("immunoconjugate"), such as an anti-viral drug, a second anti-influenza antibody, or any other therapeutic moiety useful for treating a viral infection, e.g., influenza viral infection. See below.

[0073] The present invention also provides a complex comprising an anti-CoV-S antigen-binding protein, e.g., antibody or antigen-binding fragment, discussed herein complexed with CoV-S polypeptide or an antigenic fragment thereof and / or with a secondary antibody or antigen-binding fragment thereof (e.g., detectably labeled secondary antibody) that binds specifically to the anti-CoV-S antibody or fragment. In an embodiment of the invention, the antibody or fragment is in vitro (e.g., is immobilized to a solid substrate) or is in the body of a subject. In an embodiment of the invention, the CoV-S is in vitro (e.g., is immobilized to a solid substrate) or is on the surface of a virus or is in the body of a subject. Immobilized anti-CoV-S antibodies and antigen-binding fragments thereof which are covalently linked to an insoluble matrix material (e.g., glass or polysaccharide such as agarose or sepharose, e.g., a bead or other particle thereof) are also part of the present invention; optionally, wherein the immobilized antibody is complexed with CoV-S or antigenic fragment thereof or a secondary antibody or fragment thereof.

[0074] "Isolated" antigen-binding proteins, antibodies or antigen-binding fragments thereof, polypeptides, polynucleotides and vectors, are at least partially free of other biological molecules from the cells or cell culture from which they are produced. Such biological molecules include nucleic acids, proteins, other antibodies or antigen-binding fragments, lipids, carbohydrates, or other material such as cellular debris and growth medium. An isolated antibody or antigen-binding fragment may further be at least partially free of expression system components such as biological molecules from a host cell or of the growth medium thereof. Generally, the term "isolated" is not intended to refer to a complete absence of such biological molecules or to an absence of water, buffers, or salts or to components of a pharmaceutical formulation that includes the antibodies or fragments.

[0075] The term "epitope" refers to an antigenic determinant (e.g., a CoV-S polypeptide) that interacts with a specific antigen-binding site of an antigen-binding protein, e.g., a 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 be linear or conformational, that is, composed of non-linear amino acids. In certain 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 certain embodiments, may have specific three-dimensional structural characteristics, and / or specific charge characteristics.

[0076] Methods for determining the epitope of an antigen-binding protein, e.g., antibody or fragment or polypeptide, include alanine scanning mutational analysis, peptide blot analysis (Reineke (2004) Methods Mol. Biol. 248: 443-63), peptide cleavage analysis, crystallographic studies and NMR analysis. In addition, methods such as epitope excision, epitope extraction and chemical modification of antigens can be employed (Tomer (2000) Prot. Sci. 9: 487-496). Another method that can be used to identify the amino acids within a polypeptide with which an antigen-binding protein (e.g., antibody or fragment or polypeptide) (e.g., coversin) interacts is hydrogen / deuterium exchange detected by mass spectrometry. In general terms, the hydrogen / deuterium exchange method involves deuterium-labeling the protein of interest, followed by binding the antigen-binding protein, e.g., antibody or fragment or polypeptide, to the deuterium-labeled protein. Next, the CoV-S protein / antigen-binding protein complex is transferred to water and exchangeable protons within amino acids that are protected by the antibody complex undergo deuterium-to-hydrogen back-exchange at a slower rate than exchangeable protons within amino acids that are not part of the interface. As a result, amino acids that form part of the protein / antigen-binding protein interface may retain deuterium and therefore exhibit relatively higher mass compared to amino acids not included in the interface. After dissociation of the antigen-binding protein (e.g., antibody or fragment or polypeptide), the target protein is subjected to protease cleavage and mass spectrometry analysis, thereby revealing the deuterium-labeled residues which correspond to the specific amino acids with which the antigen-binding protein interacts. See, e.g., Ehring (1999) Analytical Biochemistry 267: 252-259; Engen and Smith (2001) Anal. Chem. 73: 256A-265A.

[0077] The term "competes" as used herein, refers to an antigen-binding protein (e.g., antibody or antigen-binding fragment thereof) that binds to an antigen (e.g., CoV-S) and inhibits or blocks the binding of another antigen-binding protein (e.g., antibody or antigen-binding fragment thereof) to the antigen. The term also includes competition between two antigen-binding proteins e.g., antibodies, in both orientations, i.e., a first antibody that binds and blocks binding of second antibody and vice versa. In certain embodiments, the first antigen-binding protein (e.g., antibody) and second antigen-binding protein (e.g., antibody) may bind to the same epitope. Alternatively, the first and second antigen-binding proteins (e.g., antibodies) may bind to different, but, for example, overlapping epitopes, wherein binding of one inhibits or blocks the binding of the second antibody, e.g., via steric hindrance. Competition between antigen-binding proteins (e.g., antibodies) may be measured by methods known in the art, for example, by a real-time, label-free bio-layer interferometry assay. Epitope mapping (e.g., via alanine scanning or hydrogen-deuterium exchange (HDX)) can be used to determine whether two or more antibodies are non-competing (e.g., on a spike protein receptor binding domain (RBD) monomer), competing for the same epitope, or competing but with diverse micro-epitopes (e.g., identified through HDX). In an embodiment of the invention, competition between a first and second anti-CoV-S antigen-binding protein (e.g., antibody) is determined by measuring the ability of an immobilized first anti-CoV-S antigen-binding protein (e.g., antibody) (not initially complexed with CoV-S protein) to bind to soluble CoV-S protein complexed with a second anti-CoV-S antigen-binding protein (e.g., antibody). A reduction in the ability of the first anti-CoV-S antigen-binding protein (e.g., antibody) to bind to the complexed CoV-S protein, relative to uncomplexed CoV-S protein, indicates that the first and second anti-CoV-S antigen-binding proteins (e.g., antibodies) compete. The degree of competition can be expressed as a percentage of the reduction in binding. Such competition can be measured using a real time, label-free bio-layer interferometry assay, e.g., on an Octet RED384 biosensor (Pall ForteBio Corp.), ELISA (enzyme-linked immunosorbent assays) or SPR (surface plasmon resonance).

[0078] Binding competition between anti-CoV-S antigen-binding proteins (e.g., monoclonal antibodies (mAbs)) can be determined using a real time, label-free bio-layer interferometry assay on an Octet RED384 biosensor (Pall ForteBio Corp.). For example, to determine competition between two anti-CoV-S monoclonal antibodies, the anti-CoV-S mAb can be first captured onto anti-hFc antibody coated Octet biosensor tips (Pall ForteBio Corp., # 18-5060) by submerging the tips into a solution of anti-CoV-S mAb (subsequently referred to as "mAb1"). As a positive-control for blocking, the antibody captured biosensor tips can then be saturated with a known blocking isotype control mAb (subsequently referred to as "blocking mAb") by dipping into a solution of blocking mAb. To determine if mAb2 competes with mAb1, the biosensor tips can then be subsequently dipped into a co-complexed solution of CoV-S polypeptide and a second anti-CoV-S mAb (subsequently referred to as "mAb2"), that had been pre-incubated for a period of time and binding of mAb1 to the CoV-S polypeptide can be determined. The biosensor tips can be washed in buffer in between every step of the experiment. The real-time binding response can be monitored during the course of the experiment and the binding response at the end of every step can be recorded.

[0079] For example, in an embodiment of the invention, the competition assay is conducted at 25 °C and pH about 7, e.g., 7.4, e.g., in the presence of buffer, salt, surfactant and a non-specific protein (e.g., bovine serum albumin).

[0080] Typically, an antibody or antigen-binding fragment of the invention which is modified in some way retains the ability to specifically bind to CoV-S, e.g., retains at least 10% of its CoV-S binding activity (when compared to the parental antibody) when that activity is expressed on a molar basis. Preferably, an antibody or antigen-binding fragment of the invention retains at least 20%, 50%, 70%, 80%, 90%, 95% or 100% or more of the CoV-S binding affinity as the parental antibody. It is also intended that an antibody or antigen-binding fragment of the invention can include conservative or non-conservative amino acid substitutions (referred to as "conservative variants" or "function conserved variants" of the antibody) that do not substantially alter its biologic activity.

[0081] A "variant" of a polypeptide, such as an immunoglobulin chain (e.g., mAb8021 V H , V L , HC, or LC, mAb8028 V H , V L , HC, or LC, or mAb8029 V H , V L , HC, or LC), refers to a polypeptide comprising an amino acid sequence that is at least about 70-99.9% (e.g., 70, 72, 74, 75, 76, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 99.5, 99.9%) identical or similar to a referenced amino acid sequence that is set forth herein (e.g., SEQ ID NO: 2, 10, 18, 20, 22, 30, 38, 40, 42, 50, 58, or 60); when the comparison is performed by a BLAST algorithm wherein the parameters of the algorithm are selected to give the largest match between the respective sequences over the entire length of the respective reference sequences (e.g., expect threshold: 10; word size: 3; max matches in a query range: 0; BLOSUM 62 matrix; gap costs: existence 11, extension 1; conditional compositional score matrix adjustment).

[0082] A "variant" of a polynucleotide refers to a polynucleotide comprising a nucleotide sequence that is at least about 70-99.9% (e.g., at least about 70, 72, 74, 75, 76, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 99.5, or 99.9%) identical to a referenced nucleotide sequence that is set forth herein (e.g., SEQ ID NO: 1, 9, 17, 19, 21, 29, 37, 39, 41, 49, 57, or 59); when the comparison is performed by a BLAST algorithm wherein the parameters of the algorithm are selected to give the largest match between the respective sequences over the entire length of the respective reference sequences (e.g., expect threshold: 10; word size: 28; max matches in a query range: 0; match / mismatch scores: 1, -2; gap costs: linear).

[0083] Anti-CoV-S antigen-binding proteins, e.g., antibodies and antigen-binding fragments thereof of the present invention, in an embodiment of the invention, include a heavy chain immunoglobulin variable region having at least 70% (e.g., 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or greater) amino acid sequence identity to the HCVR amino acid sequences set forth in Table 1; and / or a light chain immunoglobulin variable region having at least 70% (e.g., 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or greater) amino acid sequence identity to the LCVR amino acid sequences set forth in Table 1. In some instances the antibody retains the exact CDR sequences of Table 1. In other words, the changes lie outside of the CDR regions. Variant antibodies typically retain the binding specificity along with certain other properties of the starting candidate antibody.

[0084] In addition, a variant anti-CoV-S antigen-binding protein may include a polypeptide comprising an amino acid sequence that is set forth herein except for one or more (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) mutations such as, for example, missense mutations (e.g., conservative substitutions), non-sense mutations, deletions, or insertions. For example, the present invention includes antigen-binding proteins which include an immunoglobulin light chain variant comprising an LCVR amino acid sequence set forth in Table 1 but having one or more of such mutations and / or an immunoglobulin heavy chain variant comprising an HCVR amino acid sequence set forth in Table 1 but having one or more of such mutations. In an embodiment of the invention, a variant anti-CoV-S antigen-binding protein includes an immunoglobulin light chain variant comprising CDR-L1, CDR-L2 and CDR-L3 wherein one or more (e.g., 1 or 2 or 3) of such CDRs has one or more of such mutations (e.g., conservative substitutions) and / or an immunoglobulin heavy chain variant comprising CDR-H1, CDR-H2 and CDR-H3 wherein one or more (e.g., 1 or 2 or 3) of such CDRs has one or more of such mutations (e.g., conservative substitutions). Substitutions can be in a CDR, framework, or constant region. In some instances, antibodies of the invention retain the exact CDR sequences of Table 1 but may have the above described mutations in the remainder of the HCVR / LCVR, or heavy / light chain. Variant antibodies can retain the functions of the starting candidate antibody (see below).

[0085] The invention further provides variant anti-CoV-S antigen-binding proteins, e.g., antibodies or antigen-binding fragments thereof, comprising one or more variant CDRs (e.g., any one or more of CDR-L1, CDR-L2, CDR-L3, CDR-H1, CDR-H2 and / or CDR-H3) that are set forth herein with at least 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 99.9% sequence identity or similarity to, e.g., the heavy chain and light chain CDRs of Table 1. Once again, such variants can retain the function of the starting candidate antibody (see below).

[0086] Embodiments of the present invention also include variant antigen-binding proteins, e.g., anti-CoV-S antibodies and antigen-binding fragments thereof, that comprise immunoglobulin V H s and V L s; or HCs and LCs, which comprise an amino acid sequence having 70% or more (e.g., 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or greater) overall amino acid sequence identity or similarity to the amino acid sequences of the corresponding V H s, V L s, HCs or LCs specifically set forth herein, but wherein the CDR-L1, CDR-L2, CDR-L3, CDR-H1, CDR-H2 and CDR-H3 of such immunoglobulins are not variants and comprise CDR amino acid sequence set forth in Table 1. Thus, in such embodiments, the CDRs within variant antigen-binding proteins are not, themselves, variants.

[0087] Conservatively modified variant anti-CoV-S antibodies and antigen-binding fragments thereof are also part of the present invention. A "conservatively modified variant" or a "conservative substitution" refers to a variant wherein there is one or more substitutions of amino acids in a polypeptide with other amino acids having similar characteristics (e.g. charge, side-chain size, hydrophobicity / hydrophilicity, backbone conformation and rigidity, etc.). Such changes can frequently be made without significantly disrupting the biological activity of the antibody or fragment. Those of skill in this art recognize that, in general, single amino acid substitutions in non-essential regions of a polypeptide do not substantially alter biological activity (see, e.g., Watson et al. (1987) Molecular Biology of the Gene, The Benjamin / Cummings Pub. Co., p. 224 (4th Ed.)). In addition, substitutions of structurally or functionally similar amino acids are less likely to significantly disrupt biological activity.

[0088] 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.

[0089] Function-conservative variants of the anti-CoV-S antibodies and antigen-binding fragments thereof are also part of the present invention. Any of the variants of the anti-CoV-S antibodies and antigen-binding fragments thereof (as discussed herein) may be "function-conservative variants". Such function-conservative variants may, in some cases, also be characterized as conservatively modified variants. "Function-conservative variants," as used herein, refers to variants of the anti-CoV-S antibodies or antigen-binding fragments thereof in which one or more amino acid residues have been changed without significantly altering one or more functional properties of the antibody or fragment. In an embodiment of the invention, a function-conservative variant anti-CoV-S antibody or antigen-binding fragment thereof of the present invention comprises a variant amino acid sequence and exhibits one or more of the following functional properties: Inhibits growth of coronavirus (e.g., SARS-CoV-2, SARS-CoV, and / or MERS-CoV) in ACE2- and / or TMPRSS2-expressing cells (e.g., Calu-3 cells); Does not significantly bind to MDCK / Tet-on cells which do not express ACE2 and / or TMPRSS2; Limits spread of coronavirus infection (e.g., by SARS-CoV-2, SARS-CoV, and / or MERS-CoV) of cells, e.g., Calu-3, in vitro; and / or Protects a mouse engineered to express the human TMPRSS2 and / or ACE2 protein from death caused by coronavirus infection (e.g., SARS-CoV-2, SARS-CoV, or MERS-CoV), for example, wherein the mice are infected with an otherwise lethal dose of the virus, optionally when combined with a second therapeutic agent. Protects a mouse engineered to express the human TMPRSS2 and / or ACE2 protein from weight loss caused by coronavirus infection (e.g., SARS-CoV-2, SARS-CoV, or MERS-CoV), for example, wherein the mice are infected with a dose of the virus that would otherwise cause weight loss, optionally when combined with a second therapeutic agent. In some instances, an antibody may have the CDR sequences of the starting candidate antibody and be a functionally conserved antibody. For example, an antibody of the invention may comprise the six CDR sequences of the mAb10933 antibody and (i) exhibit a neutralization potency (IC 50 ) against wild-type VSV-SARS-CoV-2-S psuedoparticles in Vero cells of less than 50 pM (e.g., as in Example 12), (ii) exhibit a relative neutralization of VSV-SARS-CoV-2 variants encoding S protein of at least 95% compared to wild-type (e.g., of the variants identified in Table 27 of Example 12), (iii) exhibit a neutralization potency (IC 50 ) against VSV-SARS-CoV-2 variants encoding spike protein of less than 150 pM (e.g., the variants identified in Table 28 of Example 12), (iv) exhibit a blocking potency (IC 50 ) of less than 7.1 pM (e.g., less than 7.0 pM, less than 6.5 pM, less than 6.0 pM, less than 5.5 pM, less than 5.0 pM, less than 4.5 pM, or less than 4.0 pM) in an ELISA-based assay measuring the antibody's ability to block binding of SARS-CoV-2 S protein RBD to human ACE2 (e.g., as in Example 14), and / or (v) exhibit neutralization of live SARS-CoV-2 virus. Similarly, an antibody may comprise the six CDR sequences of the mAb 10987 antibody and (i) exhibit a neutralization potency (IC 50 ) against wild-type VSV-SARS-CoV-2-S psuedoparticles in Vero cells of less than 50 pM (e.g., as in Example 12), (ii) exhibit a relative neutralization of VSV-SARS-CoV-2 variants encoding spike protein of at least 95% compared to wild-type (e.g., of the variants identified in Table 27 of Example 12), (iii) exhibit a neutralization potency (IC 50 ) against VSV-SARS-CoV-2 variants encoding S protein of less than 55 pM (e.g., the variants identified in Table 28 of Example 12), (iv) exhibit a blocking potency (IC 50 ) of less than 185 pM (e.g., less than 180 pM, less than 170 pM, less than 160 pM, less than 150 pM, less than 140 pM, less than 130 pM, less than 120 pM, less than 110 pM, less than 100 pM, or less than 90 pM) in an ELISA-based assay measuring the antibody's ability to block binding of SARS-CoV-2 S protein RBD to human ACE2 (e.g., as in Example 14), and / or (v) exhibit neutralization of live virus.These properties also apply to variants of the 10933 and 10987 antibodies.

[0090] A "neutralizing" or "antagonist" anti-CoV-S antigen-binding protein, e.g., antibody or antigen-binding fragment, refers to a molecule that inhibits an activity of CoV-S to any detectable degree, e.g., inhibits the ability of CoV-S to bind to a receptor such as ACE2, to be cleaved by a protease such as TMPRSS2, or to mediate viral entry into a host cell or viral reproduction in a host cell.

[0091] Table 1 refers to antigen-binding proteins, such as antibodies and antigen-binding fragments thereof, that comprise the heavy chain or V H (or a variant thereof) and light chain or V L (or a variant thereof) as set forth below; or that comprise a V H that comprises the CDRs thereof (CDR-H1 (or a variant thereof), CDR-H2 (or a variant thereof) and CDR-H3 (or a variant thereof)) and a V L that comprises the CDRs thereof (CDR-L1 (or a variant thereof), CDR-L2 (or a variant thereof) and CDR-L3 (or a variant thereof)), e.g., wherein the immunoglobulin chains, variable regions and / or CDRs comprise the specific amino acid sequences described below.

[0092] The antibodies described herein also include embodiments wherein the V H is fused to a wild-type IgG4 (e.g., wherein residue 108 is S) or to IgG4 variants (e.g., wherein residue 108 is P).

[0093] Antibodies and antigen-binding fragments of the present invention comprise immunoglobulin chains including the amino acid sequences set forth herein as well as cellular and in vitro post-translational modifications to the antibody. For example, the present invention includes antibodies and antigen-binding fragments thereof that specifically bind to CoV-S comprising heavy and / or light chain amino acid sequences set forth herein (e.g., CDR-H1, CDR-H2, CDR-H3, CDR-L1, CDR-L2 and / or CDR-L3) as well as antibodies and fragments wherein one or more amino acid residues is glycosylated, one or more Asn residues is deamidated, one or more residues (e.g., Met, Trp and / or His) is oxidized, the N-terminal Gln is pyroglutamate (pyroE) and / or the C-terminal Lysine is missing.

[0094] The amino acid and nucleotide sequences of exemplary anti-SARS-CoV-2-Spike protein (SARS-CoV-2-S) antibodies are shown in the Table of Exemparly Sequences, below. Table of Exemplary Sequences Antibody Designation Component Part Sequence SEQ ID NO mAb10933 Amino Acids HCVR202HCDR1GFTFSDYY204HCDR2ITYSGSTI206HCDR3ARDRGTTMVPFDY208LCVR210LCDR1QDITNY212LCDR2AAS55LCDR3QQYDNLPLT214HC216LC218Nucleic Acids HCVR201HCDR1GGATTCACCTTCAGTGACTACTAC203HCDR2ATTACTTATAGTGGTAGTACCATA205HCDR3207LCVR209LCDR1CAGGACATTACCAACTAT211LCDR2GCTGCATCC54LCDR3CAACAGTATGATAATCTCCCTCTCACT213HC215LC217mAb10934 Amino Acids HCVR220HCDR1GITFSNAW222HCDR2IKSKTDGGTT224HCDR3TTARWDWYFDL226LCVR228LCDR1QDIWNY230LCDR2DAS194LCDR3QQHDDLPPT232HC234LC236Nucleic Acids HCVR219HCDR1GGAATCACTTTCAGTAACGCCTGG221HCDR2ATTAAAAGCAAAACTGATGGTGGGACAACA223HCDR3ACCACAGCGAGGTGGGACTGGTACTTCGATCTC225LCVR227LCDR1CAGGACATTTGGAATTAT229LCDR2GATGCATCC193LCDR3CAACAGCATGATGATCTCCCTCCGACC231HC233LC235mAb10987 Amino Acids HCVR640HCDR1GFTFSNYA642HCDR2ISYDGSNK499HCDR3ASGSDYGDYLLVY644LCVR646LCDR1SSDVGGYNY648LCDR2DVS650LCDR3NSLTSISTWV652HC654LC656Nucleic Acids HCVR639HCDR1GGATTCACCTTCAGTAACTATGCT641HCDR2ATATCATATGATGGAAGTAATAAA498HCDR3643LCVR645LCDR1AGCAGTGACGTTGGTGGTTATAACTAT647LCDR2GATGTCAGT649LCDR3AACTCTTTGACAAGCATCAGCACTTGGGTG651HC653LC655mAb10989 Amino Acids HCVR678HCDR1GYIFTGYY680HCDR2INPNSGGA682HCDR3ARGSRYDWNQNNWFDP684LCVR686LCDR1SSDVGTYNY688LCDR2DVS650LCDR3SSFTTSSTVV690HC692LC694Nucleic Acids HCVR677HCDR1GGATACATCTTCACCGGCTACTAT679HCDR2ATCAACCCTAACAGTGGTGGCGCA681HCDR3683LCVR685LCDR1AGCAGTGACGTTGGTACTTATAACTAT687LCDR2GATGTCAGT649LCDR3AGCTCATTTACAACCAGCAGCACTGTGGTT689HC691LC693 Administration of Antibodies

[0095] The present invention provides methods for administering an anti-CoV-S antigen-binding protein of the present invention, e.g., those of Table 1, comprising introducing the antigen-binding protein into the body of a subject (e.g., a human). For example, the method comprises piercing the body of the subject with a needle of a syringe and injecting the antigen-binding protein into the body of the subject, e.g., into the vein, artery, tumor, muscular tissue or subcutis of the subject.

[0096] The present invention provides a vessel (e.g., a plastic or glass vial, e.g., with a cap or a chromatography column, hollow bore needle or a syringe cylinder) comprising an anti-CoV-S antigen-binding protein of the present invention, e.g., those of Table 1.

[0097] The present invention also provides an injection device comprising one or more antigen-binding proteins (e.g., antibody or antigen-binding fragment) that bind specifically to CoV-S, e.g., those of Table 1, or a pharmaceutical composition thereof. The injection device may be packaged into a kit. An injection device is a device that introduces a substance into the body of a subject via a parenteral route, e.g., intramuscular, subcutaneous or intravenous. For example, an injection device may be a syringe (e.g., pre-filled with the pharmaceutical composition, such as an auto-injector) which, for example, includes a cylinder or barrel for holding fluid to be injected (e.g., comprising the antibody or fragment or a pharmaceutical composition thereof), a needle for piecing skin and / or blood vessels for injection of the fluid; and a plunger for pushing the fluid out of the cylinder and through the needle bore. In an embodiment of the invention, an injection device that comprises an antigen-binding protein, e.g., an antibody or antigen-binding fragment thereof, from a combination of the present invention, or a pharmaceutical composition thereof is an intravenous (IV) injection device. Such a device can include the antigen-binding protein or a pharmaceutical composition thereof in a cannula or trocar / needle which may be attached to a tube which may be attached to a bag or reservoir for holding fluid (e.g., saline) introduced into the body of the subject through the cannula or trocar / needle. The antibody or fragment or a pharmaceutical composition thereof may, in an embodiment of the invention, be introduced into the device once the trocar and cannula are inserted into the vein of a subject and the trocar is removed from the inserted cannula. The IV device may, for example, be inserted into a peripheral vein (e.g., in the hand or arm); the superior vena cava or inferior vena cava, or within the right atrium of the heart (e.g., a central IV); or into a subclavian, internal jugular, or a femoral vein and, for example, advanced toward the heart until it reaches the superior vena cava or right atrium (e.g., a central venous line). In an embodiment of the invention, an injection device is an autoinjector; a jet injector or an external infusion pump. A jet injector uses a high-pressure narrow jet of liquid which penetrate the epidermis to introduce the antibody or fragment or a pharmaceutical composition thereof to a subject's body. External infusion pumps are medical devices that deliver the antibody or fragment or a pharmaceutical composition thereof into a subject's body in controlled amounts. External infusion pumps may be powered electrically or mechanically. Different pumps operate in different ways, for example, a syringe pump holds fluid in the reservoir of a syringe, and a moveable piston controls fluid delivery, an elastomeric pump holds fluid in a stretchable balloon reservoir, and pressure from the elastic walls of the balloon drives fluid delivery. In a peristaltic pump, a set of rollers pinches down on a length of flexible tubing, pushing fluid forward. In a multi-channel pump, fluids can be delivered from multiple reservoirs at multiple rates.Preparation of Human Antibodies

[0098] Methods for generating human antibodies in transgenic mice are known in the art. Any such known methods can be used in the context of the present invention to make human antibodies that specifically bind to CoV-S. An immunogen comprising any one of the following can be used to generate antibodies to CoV-S. In certain embodiments of the invention, the antibodies of the invention are obtained from mice immunized with a full length, native CoV-S, or with a live attenuated or inactivated virus, or with DNA encoding the protein or fragment thereof. Alternatively, the CoV-S protein or a fragment thereof may be produced using standard biochemical techniques and modified and used as immunogen. In one embodiment of the invention, the immunogen is a recombinantly produced CoV-S protein or fragment thereof. In certain embodiments of the invention, the immunogen may be a CoV-S polypeptide vaccine. In certain embodiments, one or more booster injections may be administered. In certain embodiments, the immunogen may be a recombinant CoV-S polypeptide expressed in E. coli or in any other eukaryotic or mammalian cells such as Chinese hamster ovary (CHO) cells.

[0099] Using VELOCIMMUNE® technology (see, for example, US 6,596,541, Regeneron Pharmaceuticals, VELOCIMMUNE®) or any other known method for generating monoclonal antibodies, high affinity chimeric antibodies to CoV-S can be initially isolated having a human variable region and a mouse constant region. The VELOCIMMUNE® technology involves generation of a transgenic mouse having a genome comprising human heavy and light chain variable regions operably linked to endogenous mouse constant region loci such that the mouse produces an antibody comprising a human variable region and a mouse constant region in response to antigenic stimulation. The DNA encoding the variable regions of the heavy and light chains of the antibody are isolated and operably linked to DNA encoding the human heavy and light chain constant regions. The DNA is then expressed in a cell capable of expressing the fully human antibody.

[0100] Generally, a VELOCIMMUNE® mouse is challenged with the antigen of interest, and lymphatic cells (such as B-cells) are recovered from the mice that express antibodies. The lymphatic cells may be fused with a myeloma cell line to prepare immortal hybridoma cell lines, and such hybridoma cell lines are screened and selected to identify hybridoma cell lines that produce antibodies specific to the antigen of interest. DNA encoding the variable regions of the heavy chain and light chain may be isolated and linked to desirable isotypic constant regions of the heavy chain and light chain. Such an antibody protein may be produced in a cell, such as a CHO cell. Alternatively, DNA encoding the antigen-specific chimeric antibodies or the variable domains of the light and heavy chains may be isolated directly from antigen-specific lymphocytes.

[0101] Initially, high affinity chimeric antibodies are isolated having a human variable region and a mouse constant region. As in the experimental section below, the antibodies are characterized and selected for desirable characteristics, including affinity, selectivity, epitope, etc. The mouse constant regions are replaced with a desired human constant region to generate the fully human antibody of the invention, for example wild-type or modified IgG1 or IgG4. While the constant region selected may vary according to specific use, high affinity antigen-binding and target specificity characteristics reside in the variable region.Anti-Coronavirus Spike Protein Antibodies Comprising Fc Variants

[0102] According to certain embodiments of the present invention, anti-CoV-S antigen-binding proteins, e.g., antibodies or antigen-binding fragments, are provided comprising an Fc domain comprising one or more mutations, which, for example, enhance or diminish antibody binding to the FcRn receptor, e.g., at acidic pH as compared to neutral pH. For example, the present invention includes anti-CoV-S antibodies comprising a mutation in the C H 2 or a C H 3 region of the Fc domain, wherein the mutation(s) increases the affinity of the Fc domain to FcRn in an acidic environment (e.g., in an endosome where pH ranges from about 5.5 to about 6.0). Such mutations may result in an increase in serum half-life of the antibody when administered to an animal. Non-limiting examples of such Fc modifications include, e.g., a modification at position 250 (e.g., E or Q); 250 and 428 (e.g., L or F); 252 (e.g., L / Y / F / W or T), 254 (e.g., S or T), and 256 (e.g., S / R / Q / E / D or T); or a modification at position 428 and / or 433 (e.g., H / L / R / S / P / Q or K) and / or 434 (e.g., A, W, H, F or Y [N434A, N434W, N434H, N434F or N434Y]); or a modification at position 250 and / or 428; or a modification at position 307 or 308 (e.g., 308F, V308F), and 434. In one embodiment, the modification comprises a 428L (e.g., M428L) and 434S (e.g., N434S) modification; a 428L, 2591 (e.g., V259I), and 308F (e.g., V308F) modification; a 433K (e.g., H433K) and a 434 (e.g., 434Y) modification; a 252, 254, and 256 (e.g., 252Y, 254T, and 256E) modification; a 250Q and 428L modification (e.g., T250Q and M428L); and a 307 and / or 308 modification (e.g., 308F or 308P). In yet another embodiment, the modification comprises a 265A (e.g., D265A) and / or a 297A (e.g., N297A) modification.

[0103] For example, the present invention includes anti-CoV-S antigen-binding proteins, e.g., antibodies or antigen-binding fragments, comprising an Fc domain comprising one or more pairs or groups of mutations selected from the group consisting of: 250Q and 248L (e.g., T250Q and M248L); 252Y, 254T and 256E (e.g., M252Y, S254T and T256E); 428L and 434S (e.g., M428L and N434S); 2571 and 3111 (e.g., P257I and Q311I); 2571 and 434H (e.g., P257I and N434H); 376V and 434H (e.g., D376V and N434H); 307A, 380A and 434A (e.g., T307A, E380A and N434A); and 433K and 434F (e.g., H433K and N434F).

[0104] Anti-CoV-S antigen-binding proteins, e.g., antibodies and antigen-binding fragments thereof, that comprise a V H and / or V L as set forth herein comprising any possible combinations of the foregoing Fc domain mutations, are contemplated within the scope of the present invention.

[0105] The present invention also includes anti-CoV-S antigen-binding proteins, antibodies or antigen-binding fragments, comprising a V H set forth herein and a chimeric heavy chain constant (C H ) region, wherein the chimeric C H region comprises segments derived from the C H regions of more than one immunoglobulin isotype. For example, the antibodies of the invention may comprise a chimeric C H region comprising part or all of a C H 2 domain derived from a human IgG1, human IgG2 or human IgG4 molecule, combined with part or all of a C H 3 domain derived from a human IgG1, human IgG2 or human IgG4 molecule. According to certain embodiments, the antibodies of the invention comprise a chimeric C H region having a chimeric hinge region. For example, a chimeric hinge may comprise an "upper hinge" amino acid sequence (amino acid residues from positions 216 to 227 according to EU numbering) derived from a human IgG1, a human IgG2 or a human IgG4 hinge region, combined with a "lower hinge" sequence (amino acid residues from positions 228 to 236 according to EU numbering) derived from a human IgG1, a human IgG2 or a human IgG4 hinge region. According to certain embodiments, the chimeric hinge region comprises amino acid residues derived from a human IgG1 or a human IgG4 upper hinge and amino acid residues derived from a human IgG2 lower hinge. An antibody comprising a chimeric C H region as described herein may, in certain embodiments, exhibit modified Fc effector functions without adversely affecting the therapeutic or pharmacokinetic properties of the antibody. (See, e.g., WO2014 / 022540).Immunoconjugates

[0106] The invention encompasses an anti-CoV-S antigen-binding proteins, e.g., antibodies or antigen-binding fragments, conjugated to another moiety, e.g., a therapeutic moiety (an "immunoconjugate"), such as a toxoid or an anti-viral drug to treat influenza virus infection. In an embodiment of the invention, an anti-CoV-S antibody or fragment is conjugated to any of the further therapeutic agents set forth herein. As used herein, the term "immunoconjugate" refers to an antigen-binding protein, e.g., an antibody or antigen-binding fragment, which is chemically or biologically linked to a radioactive agent, a cytokine, an interferon, a target or reporter moiety, an enzyme, a peptide or protein or a therapeutic agent. The antigen-binding protein may be linked to the radioactive agent, cytokine, interferon, target or reporter moiety, enzyme, peptide or therapeutic agent at any location along the molecule so long as it is able to bind its target (CoV-S). Examples of immunoconjugates include antibody-drug conjugates and antibody-toxin fusion proteins. In one embodiment of the invention, the agent may be a second, different antibody that binds specifically to CoV-S. The type of therapeutic moiety that may be conjugated to the anti-CoV-S antigen-binding protein (e.g., antibody or fragment) will take into account the condition to be treated and the desired therapeutic effect to be achieved. See, e.g., Arnon et al., "Monoclonal Antibodies For Immunotargeting Of Drugs In Cancer Therapy", Monoclonal Antibodies And Cancer Therapy, Reisfeld et al. (eds.), pp. 243-56 (Alan R. Liss, Inc. 1985); Hellstrom et al., "Antibodies For Drug Delivery", Controlled Drug Delivery (2nd Ed.), Robinson et al. (eds.), pp. 623-53 (Marcel Dekker, Inc. 1987); Thorpe, "Antibody Carriers Of Cytotoxic Agents In Cancer Therapy: A Review", Monoclonal Antibodies 1984: Biological And Clinical Applications, Pinchera et al. (eds.), pp. 475-506 (1985); "Analysis, Results, And Future Prospective Of The Therapeutic Use Of Radiolabeled Antibody In Cancer Therapy", Monoclonal Antibodies For Cancer Detection And Therapy, Baldwin et al. (eds.), pp. 303-16 (Academic Press 1985), and Thorpe et al., "The Preparation And Cytotoxic Properties Of Antibody-Toxin Conjugates", Immunol. Rev., 62: 119-58 (1982).Multi-specific Antibodies

[0107] The present invention includes anti-CoV-S antigen-binding proteins, e.g., antibodies and antigen-binding fragments thereof, as well as methods of use thereof and methods of making such antigen-binding proteins. The term "anti-CoV-S" antigen-binding proteins, e.g., antibodies or antigen-binding fragments, includes multispecific (e.g., bispecific or biparatopic) molecules that include at least one first antigen-binding domain that specifically binds to CoV-S (e.g., an antigen-binding domain from an antibody of Table 1) and at least one second antigen-binding domain that binds to a different antigen or to an epitope in CoV-S which is different from that of the first antigen-binding domain. In some embodiments, the first antigen-binding domain and the second antigen-binding domain are both selected from the antigen-binding domains of Table 1. In an embodiment of the invention, the first and second epitopes overlap. In another embodiment of the invention, the first and second epitopes do not overlap. For example, in an embodiment of the invention, a multispecific antibody is a bispecific IgG antibody (e.g., IgG1 or IgG4) that includes a first antigen-binding domain that binds specifically to CoV-S including the heavy and light immunoglobulin chain of an antibody of Table 1, and a second antigen-binding domain that binds specifically to a different epitope of CoV-S. In some embodiments, a bispecific IgG antibody (e.g., IgG1 or IgG4) includes a first antigen-binding domain that binds specifically to CoV-S and a second binding domain that binds to a host cell protein, e.g., ACE2 or TMPRSS2.

[0108] The antibodies of Table 1 include multispecific molecules, e.g., antibodies or antigen-binding fragments, that include the CDR-Hs and CDR-Ls, V H and V L , or HC and LC of those antibodies, respectively (including variants thereof as set forth herein).

[0109] In an embodiment of the invention, an antigen-binding domain that binds specifically to CoV-S, which may be included in a multispecific molecule, comprises: (1) (i) a heavy chain variable domain sequence that comprises CDR-H1, CDR-H2, and CDR-H3 amino acid sequences set forth in Table 1, and (ii) a light chain variable domain sequence that comprises CDR-L1, CDR-L2, and CDR-L3 amino acid sequences set forth in Table 1; or, (2) (i) a heavy chain variable domain sequence comprising an amino acid sequence set forth in Table 1, and (ii) a light chain variable domain sequence comprising an amino acid sequence set forth in Table 1; or, (3) (i) a heavy chain immunoglobulin sequence comprising an amino acid sequence set forth in Table 1, and (ii) a light chain immunoglobulin sequence comprising an amino acid sequence set forth in Table 1.

[0110] In an embodiment of the invention, the multispecific antibody or fragment includes more than two different binding specificities (e.g., a trispecific molecule), for example, one or more additional antigen-binding domains which are the same or different from the first and / or second antigen-binding domain.

[0111] In one embodiment of the invention, a bispecific antigen-binding fragment comprises a first scFv (e.g., comprising V H and V L sequences of Table 1) having binding specificity for a first epitope (e.g., CoV-S) and a second scFv having binding specificity for a second, different epitope. For example, in an embodiment of the invention, the first and second scFv are tethered with a linker, e.g., a peptide linker (e.g., a GS linker such as (GGGGS) n (SEQ ID NO: 834) wherein n is, for example, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10). Other bispecific antigen-binding fragments include an F(ab) 2 of a bispecific IgG antibody which comprises the heavy and light chain CDRs of Table 1 and of another antibody that binds to a different epitope.Therapeutic Methods

[0112] The present invention provides methods for treating or preventing viral infection (e.g., coronavirus infection) by administering a therapeutically effective amount of anti-CoV-S antigen-binding protein, e.g., antibody or antigen-binding fragment, (e.g., of Table 1) to a subject (e.g., a human) in need of such treatment or prevention.

[0113] Coronavirus infection may be treated or prevented, in a subject, by administering an anti-CoV-S antigen-binding protein of the present invention to a subject.

[0114] An effective or therapeutically effective dose of anti-CoV-S antigen-binding protein, e.g., antibody or antigen-binding fragment (e.g., of Table 1), for treating or preventing a viral infection refers to the amount of the antibody or fragment sufficient to alleviate one or more signs and / or symptoms of the infection in the treated subject, whether by inducing the regression or elimination of such signs and / or symptoms or by inhibiting the progression of such signs and / or symptoms. The dose amount may vary depending upon the age and the size of a subject to be administered, target disease, conditions, route of administration, and the like. In an embodiment of the invention, an effective or therapeutically effective dose of antibody or antigen-binding fragment thereof of the present invention, for treating or preventing viral infection, e.g., in an adult human subject, is about 0.01 to about 200 mg / kg, e.g., up to about 150 mg / kg. In an embodiment of the invention, the dosage is up to about 10.8 or 11 grams (e.g., about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 or 11 grams). Depending on the severity of the infection, the frequency and the duration of the treatment can be adjusted. In certain embodiments, the antigen-binding protein of the present invention can be administered at an initial dose, followed by one or more secondary doses. In certain embodiments, the initial dose may be followed by administration of a second or a plurality of subsequent doses of 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.

[0115] As used herein, the term "subject" refers to a mammal (e.g., rat, mouse, cat, dog, cow, pig, sheep, horse, goat, rabbit), preferably a human, for example, in need of prevention and / or treatment of a disease or disorder such as viral infection or cancer. The subject may have a viral infection, e.g., an influenza infection, or be predisposed to developing an infection. Subjects predisposed to developing an infection, or subjects who may be at elevated risk for contracting an infection (e.g., of coronavirus or influenza virus), include subjects with compromised immune systems because of autoimmune disease, subjects receiving immunosuppressive therapy (for example, following organ transplant), subjects afflicted with human immunodeficiency syndrome (HIV) or acquired immune deficiency syndrome (AIDS), subjects with forms of anemia that deplete or destroy white blood cells, subjects receiving radiation or chemotherapy, or subjects afflicted with an inflammatory disorder. Additionally, subjects of very young (e.g., 5 years of age or younger) or old age (e.g., 65 years of age or older) are at increased risk. Moreover, a subject may be at risk of contracting a viral infection due to proximity to an outbreak of the disease, e.g. subject resides in a densely-populated city or in close proximity to subjects having confirmed or suspected infections of a virus, or choice of employment, e.g. hospital worker, pharmaceutical researcher, traveler to infected area, or frequent flier.

[0116] "Treat" or "treating" means to administer an anti-CoV-S antigen-binding protein, e.g., antibody or antigen-binding fragment of the present invention (e.g., of Table 1), to a subject having one or more signs or symptoms of a disease or infection, e.g., viral infection, for which the antigen-binding protein is effective when administered to the subject at an effective or therapeutically effective amount or dose (as discussed herein).

[0117] The present invention also encompasses prophylactically administering an anti-CoV-S antigen-binding protein, e.g., antibody or antigen-binding fragment thereof of the present invention (e.g., of Table 1), to a subject who is at risk of viral infection so as to prevent such infection. Passive antibody-based immunoprophylaxis has proven an effective strategy for preventing subject from viral infection. See e.g., Berry et al., Passive broad-spectrum influenza immunoprophylaxis. Influenza Res Treat. 2014; 2014:267594. Epub 2014 Sep 22; and Jianqiang et al., Passive immune neutralization strategies for prevention and control of influenza A infections, Immunotherapy. 2012 February; 4(2): 175-186; Prabhu et al., Antivir Ther. 2009;14(7):911-21, Prophylactic and therapeutic efficacy of a chimeric monoclonal antibody specific for H5 hemagglutinin against lethal H5N1 influenza. "Prevent" or "preventing" means to administer an anti-CoV-S antigen-binding protein, e.g., antibody or antigen-binding fragment of the present invention (e.g., of Table 1), to a subject to inhibit the manifestation of a disease or infection (e.g., viral infection) in the body of a subject, for which the antigen-binding protein is effective when administered to the subject at an effective or therapeutically effective amount or dose (as discussed herein).

[0118] In an embodiment of the invention, a sign or symptom of a viral infection in a subject is survival or proliferation of virus in the body of the subject, e.g., as determined by viral titer assay (e.g., coronavirus propagation in embryonated chicken eggs or coronavirus spike protein assay). Other signs and symptoms of viral infection are discussed herein.

[0119] As noted above, in some embodiments the subject may be a non-human animal, and the antigen-binding proteins (e.g., antibodies and antigen-binding fragments) discussed herein may be used in a veterinary context to treat and / or prevent disease in the non-human animals (e.g., cats, dogs, pigs, cows, horses, goats, rabbits, sheep, and the like).

[0120] The present invention provides a method for treating or preventing viral infection (e.g., coronavirus infection) or for inducing the regression or elimination or inhibiting the progression of at least one sign or symptom of viral infection such as: fever or feeling feverish / chills; cough; sore throat; runny or stuffy nose; sneezing; muscle or body aches; headaches; fatigue (tiredness); vomiting; diarrhea; respiratory tract infection; chest discomfort; shortness of breath; bronchitis; and / or pneumonia, which sign or symptom is secondary to viral infection, in a subject in need thereof (e.g., a human), by administering a therapeutically effective amount of anti-CoV-S antigen-binding protein (e.g., of Table 1) to the subject, for example, by injection of the protein into the body of the subject.Combinations and Pharmaceutical Compositions

[0121] To prepare pharmaceutical compositions of the anti-CoV-S antigen-binding proteins, e.g., antibodies and antigen-binding fragments thereof (e.g., of Table 1), antigen-binding protein is admixed with a pharmaceutically acceptable carrier or excipient. See, e.g., Remington's Pharmaceutical Sciences and U.S. Pharmacopeia: National Formulary, Mack Publishing Company, Easton, Pa. (1984); Hardman, et al. (2001) Goodman and Gilman's The Pharmacological Basis of Therapeutics, McGraw-Hill, New York, N.Y.; Gennaro (2000) Remington: The Science and Practice of Pharmacy, Lippincott, Williams, and Wilkins, New York, N.Y.; Avis, et al. (eds.) (1993) Pharmaceutical Dosage Forms: Parenteral Medications, Marcel Dekker, NY; Lieberman, et al. (eds.) (1990) Pharmaceutical Dosage Forms: Tablets, Marcel Dekker, NY; Lieberman, et al. (eds.) (1990) Pharmaceutical Dosage Forms: Disperse Systems, Marcel Dekker, NY; Weiner and Kotkoskie (2000) Excipient Toxicity and Safety, Marcel Dekker, Inc., New York, N.Y. In an embodiment of the invention, the pharmaceutical composition is sterile. Such compositions are part of the present invention.

[0122] The scope of the present invention includes desiccated, e.g., freeze-dried, compositions comprising an anti-CoV-S antigen-binding proteins, e.g., antibody or antigen-binding fragment thereof (e.g., of Table 1), or a pharmaceutical composition thereof that includes a pharmaceutically acceptable carrier but substantially lacks water.

[0123] In a further embodiment of the invention, a further therapeutic agent that is administered to a subject in association with an anti-CoV-S antigen-binding protein, e.g., antibody or antigen-binding fragment thereof (e.g., of Table 1), disclosed herein is administered to the subject in accordance with the Physicians' Desk Reference 2003 (Thomson Healthcare; 57th edition (Nov. 1, 2002)).

[0124] The mode of administration can vary. Routes of administration include oral, rectal, transmucosal, intestinal, parenteral; intramuscular, subcutaneous, intradermal, intramedullary, intrathecal, direct intraventricular, intravenous, intraperitoneal, intranasal, intraocular, inhalation, insufflation, topical, cutaneous, transdermal or intra-arterial.

[0125] The present invention provides methods for administering an anti-CoV-S antigen-binding protein, e.g., antibody or antigen-binding fragment thereof (e.g., of Table 1), comprising introducing the protein into the body of a subject. For example, the method comprises piercing the body of the subject with a needle of a syringe and injecting the antigen-binding protein into the body of the subject, e.g., into the vein, artery, tumor, muscular tissue or subcutis of the subj ect.

[0126] The present invention provides a vessel (e.g., a plastic or glass vial, e.g., with a cap or a chromatography column, hollow bore needle or a syringe cylinder) comprising any of the anti-CoV-S antigen-binding proteins, e.g., antibodies or antigen-binding fragments thereof (e.g., of Table 1), polypeptides (e.g., an HC, LC, V H or V L of Table 1) or polynucleotides (e.g., of Table 2) or vectors set forth herein or a pharmaceutical composition thereof comprising a pharmaceutically acceptable carrier.

[0127] In an embodiment of the present disclosure, an anti-CoV-S antigen-binding protein, e.g., antibody or antigen-binding fragment thereof of the present invention (e.g., of Table 1), is administered in association with one or more further therapeutic agents. A further therapeutic agent includes, but is not limited to: an anti-inflammatory agent, an antimalarial agent, a second antibody or antigen-binding fragment thereof that specifically binds TMPRSS2, and a second antibody or antigen-binding fragment thereof that specifically binds to CoV-S. In some embodiments, an antimalarial agent is chloroquine or hydroxychloroquine. In some embodiments, an anti-inflammatory agent is an antibody such as sarilumab, tocilizumab, or gimsilumab. In some embodiments, the further therapeutic agent is a second antibody or antigen-binding fragment disclosed herein, e.g., of Table 1. In certain embodiments, one, two, three, four, or more antibodies, or antigen-binding fragments thereof, of Table 1 can be administered in combination (e.g., concurrently or sequentially). Particular combinations of antibodies of Table 1 are listed in the Table of Exemplary Antibody Combinations, below (each number representing a specific combination, e.g., mAb10989 and mAb10987 is Combination 1, mAb10989 and mAb10934 is Combination 2, and so on). In some embodiments, a combination of antibodies can be selected from among those binding to different epitope clusters. For example, certain antibodies described herein belong to epitope clusters as follows: Cluster 1, mAb10987, mAb10922, mAb10936, and mAb10934; Cluster 2, mAb10989, mAb10977, and mAb10933; Cluster 3, mAb 10920; Cluster 4, mAb 10954, mAb 10986, and mAb 10964; and Cluster 5, mAb10984. Thus, a combination of two antibodies can be selected from, for example, Cluster 1 and Cluster 2, Cluster 1 and Cluster 3, Cluster 1 and Cluster 4, Cluster 1 and Cluster 5, Cluster 2 and Cluster 3, Cluster 2 and Cluster 4, Cluster 2 and Cluster 5, Cluster 3 and Cluster 4, Cluster 3 and Cluster 5, and Cluster 4 and Cluster 5. In some embodiments, an antibody that specifically binds TMPRSS2 is H1H7017N, as described in International Patent Pub. No. WO / 2019 / 147831. Table of Exemplary Antibody Combinations mAb 10989mAb 10987mAb 10934mAb 10933mAb 10920mAb 10922mAb 10936mAb 10954mAb 10964mAb 10977mAb 10984mAb 10986mAb 10989X1234567891011mAb 1098712X13141516171819202122mAb 109342324X252627282930313233mAb 10933343536X3738394041424344mAb 1092045464748X49505152535455mAb 109225657585960X616263646566mAb 10936676869707172X7374757677mAb 1095478798081828384X85868788mAb 109648990919293949596X979899mAb 10977100101102103104105106107108X109110mAb 10984111112113114115116117118119120X121mAb 10986122123124125126127128129130131132X

[0128] In some embodiments, anti-CoV-S antigen-binding proteins (e.g., anti-SARS-CoV-2-S antibodies or antigen-binding fragments thereof) from different human donors may be combined. The present invention includes a composition comprising two (or more) anti-SARS-CoV-2-S antibodies or antigen-binding fragments comprising variable domains from human subjects, wherein the two (or more) antibodies or antigen-binding fragments are derived from different subjects (e.g., two different human subjects). Antibody variable regions derived from human B cells are discussed, e.g., in Examples 1 and 2 (Table 3), which describes that variable domains cloned from such B cells are combined with a constant region not from those B cells to produce hybrid antibodies. The source (Donor) of such antibody variable regions is shown in the Table of Exemplary Human-Derived Antibody Variable Regions, below. In some embodiments, a composition may comprise a combination of an antibody or antigen-binding fragment thereof with variable domains derived from donor 1 and an antibody or antigen-binding fragment thereof with variable domains derived from donor 2. In some embodiments, a composition may comprise a combination of an antibody or antigen-binding fragment thereof with variable domains derived from donor 1 and an antibody or antigen-binding fragment thereof with variable domains derived from donor 3. In some embodiments, a composition may comprise a combination of an antibody or antigen-binding fragment thereof with variable domains derived from donor 2 and an antibody or antigen-binding fragment thereof with variable domains derived from donor 3. In some embodiments, a composition may comprise a combination of mAb10987 (e.g., an antibody comprising the CDRs, the variable regions, or the heavy and light chain sequences shown in Table 1) from Donor 1, and mAb10989 (e.g., an antibody comprising the CDRs, the variable regions, or the heavy and light chain sequences shown in Table 1) from Donor 3. Table of Exemplary Human-Derived Antibody Variable Regions mAb Donor mAb10954Donor 3mAb10955Donor 3mAb10956Donor 3mAb10957Donor 3mAb 10964Donor 1mAb 10965Donor 2mAb10966Donor 3mAb10967Donor 3mAb10970Donor 1mAb10971Donor 1mAb10977Donor 1mAb10984Donor 1mAb10985Donor 1mAb10986Donor 1mAb10987Donor 1mAb10988Donor 3mAb 10989Donor 3mAb 10969Donor 1

[0129] In some embodiments, the further therapeutic agent is an anti-viral drug and / or a vaccine. As used herein, the term "anti-viral drug" refers to any anti-infective drug or therapy used to treat, prevent, or ameliorate a viral infection in a subject. The term "anti-viral drug" includes, but is not limited to a cationic steroid antimicrobial, leupeptin, aprotinin, ribavirin, or interferon-alpha2b. Methods for treating or preventing virus (e.g., coronavirus) infection in a subject in need of said treatment or prevention by administering an antibody or antigen-binding fragment of Table 1 in association with a further therapeutic agent are part of the present invention.

[0130] For example, in an embodiment of the invention, the further therapeutic agent is a vaccine, e.g., a coronavirus vaccine. In an embodiment of the invention, a vaccine is an inactivated / killed virus vaccine, a live attenuated virus vaccine or a virus subunit vaccine.

[0131] For example, in an embodiment of the invention, the further therapeutic agent is: (camostat mesylate); (nafamostat mesylate); (bromhexine hydrochloride (BHH)); (4-(2-aminomethyl)benzenesulfonyl fluoride hydrochloride (AEBSF)); or (polyamide). See Shen et al. Biochimie 142: 1-10 (2017).

[0132] In an embodiment of the invention, the anti-viral drug is an antibody or antigen-binding fragment that binds specifically to coronavirus, e.g., SARS-CoV-2, SARS-CoV, or MERS-CoV. Exemplary anti-CoV-S antibodies include, but are not limited to: H4sH15188P; H1H15188P; H1H15211P; H1H15177P; H4sH15211P; H1H15260P2; H1H15259P2; H1H15203P; H4sH15260P2; H4sH15231P2; H1H15237P2; H1H15208P; H1H15228P2; H1H15233P2; H1H15264P2; H1H15231P2; H1H15253P2; H1H15215P; and H1H15249P2, as set forth in International patent application publication no. WO / 2015 / 179535, or an antigen-binding fragment thereof, e.g., wherein the antibody or fragment comprises a light chain immunoglobulin that includes CDR-L1, CDR-L2 and CDR-L3 (e.g., the V L or light chain thereof); and a heavy chain that includes CDR-H1, CDR-H2 and CDR-H3 (e.g., the V H or heavy chain thereof) of any of the foregoing anti-CoV-S antibodies.

[0133] In a certain embodiment of the invention, the further therapeutic agent is not aprotinin, leupeptin, a cationic steroid antimicrobial, an influenza vaccine (e.g., killed, live, attenuated whole virus or subunit vaccine), or an antibody against influenza virus (e.g., an anti-hemagglutinin antibody).

[0134] The term "in association with" indicates that the components, an anti-CoV-S antigen-binding protein, e.g., antibody or antigen-binding fragment thereof of the present invention, along with another agent, can be formulated into a single composition, e.g., for simultaneous delivery, or formulated separately into two or more compositions (e.g., a kit). Each component can be administered to a subject at a different time than when the other component is administered; for example, each administration may be given non-simultaneously (e.g., separately or sequentially) at intervals over a given period of time. Moreover, the separate components may be administered to a subject by the same or by a different route (e.g., wherein an anti-CoV-S antibody or antigen-binding fragment thereof.Kits

[0135] Further provided are kits comprising one or more components that include, but are not limited to, an anti-CoV-S antigen-binding protein, e.g., an antibody or antigen-binding fragment as discussed herein (e.g., of Table 1), in association with one or more additional components including, but not limited to, a further therapeutic agent, as discussed herein. The antigen-binding protein and / or the further therapeutic agent can be formulated as a single composition or separately in two or more compositions, e.g., with a pharmaceutically acceptable carrier, in a pharmaceutical composition.

[0136] In one embodiment of the invention, the kit includes an anti-CoV-S antigen-binding protein, e.g., an antibody or antigen-binding fragment thereof of the invention (e.g., of Table 1), or a pharmaceutical composition thereof in one container (e.g., in a sterile glass or plastic vial) and a further therapeutic agent in another container (e.g., in a sterile glass or plastic vial).

[0137] In another embodiment, the kit comprises a combination of the invention, including an anti-CoV-S antigen-binding protein, e.g., antibody or antigen-binding fragment thereof of the invention (e.g., of Table 1), or pharmaceutical composition thereof in combination with one or more further therapeutic agents formulated together, optionally, in a pharmaceutical composition, in a single, common container.

[0138] If the kit includes a pharmaceutical composition for parenteral administration to a subject, the kit can include a device (e.g., an injection device) for performing such administration. For example, the kit can include one or more hypodermic needles or other injection devices as discussed above containing the anti-CoV-S antigen-binding protein, e.g., antibody or antigen-binding fragment thereof of the present invention (e.g., of Table 1).

[0139] The kit can include a package insert including information concerning the pharmaceutical compositions and dosage forms in the kit. Generally, such information aids patients and physicians in using the enclosed pharmaceutical compositions and dosage forms effectively and safely. For example, the following information regarding a combination of the invention may be supplied in the insert: pharmacokinetics, pharmacodynamics, clinical studies, efficacy parameters, indications and usage, contraindications, warnings, precautions, adverse reactions, overdosage, proper dosage and administration, how supplied, proper storage conditions, references, manufacturer / distributor information and patent information.Diagnostic Uses of the Antibodies

[0140] The anti-CoV-S antigen-binding proteins, e.g., antibodies or antigen-binding fragments thereof of the present invention (e.g., of Table 1), may be used to detect and / or measure CoV-S in a sample. Exemplary assays for CoV-S may include, e.g., contacting a sample with an anti-CoV-S antigen-binding protein of the invention, wherein the anti-CoV-S antigen-binding protein is labeled with a detectable label or reporter molecule or used as a capture ligand to selectively isolate CoV-S from samples. The presence of an anti-CoV-S antigen-binding protein complexed with CoV-S indicates the presence of CoV-S in the sample. Alternatively, an unlabeled anti-CoV-S antibody can be used in combination with a secondary antibody which is itself detectably labeled. The detectable label or reporter molecule can be a radioisotope, such as 3< H, 14< C, 32< P, 35< S, or 125< I; a fluorescent or chemiluminescent moiety such as fluorescein isothiocyanate, or rhodamine; or an enzyme such as alkaline phosphatase, β-galactosidase, horseradish peroxidase, or luciferase. Specific exemplary assays that can be used to detect or measure CoV-S in a sample include neutralization assays, enzyme-linked immunosorbent assay (ELISA), radioimmunoassay (RIA), and fluorescence-activated cell sorting (FACS). Thus, the present invention includes a method for detecting the presence of spike protein polypeptide in a sample comprising contacting the sample with an anti-CoV-S antigen-binding protein and detecting the presence of a CoV-S / anti-CoV-S antigen-binding protein wherein the presence of the complex indicates the presence of CoV-S.

[0141] An anti-CoV-S antigen-binding protein of the invention (e.g., of Table 1) may be used in a Western blot or immune-protein blot procedure for detecting the presence of CoV-S or a fragment thereof in a sample. Such a procedure forms part of the present invention and includes the steps of e.g.: (1) providing a membrane or other solid substrate comprising a sample to be tested for the presence of CoV-S, e.g., optionally including the step of transferring proteins from a sample to be tested for the presence of CoV-S (e.g., from a PAGE or SDS-PAGE electrophoretic separation of the proteins in the sample) onto a membrane or other solid substrate using a method known in the art (e.g., semi-dry blotting or tank blotting); and contacting the membrane or other solid substrate to be tested for the presence of CoV-S or a fragment thereof with an anti-CoV-S antigen-binding protein of the invention. Such a membrane may take the form, for example, of a nitrocellulose or vinyl-based (e.g., polyvinylidene fluoride (PVDF)) membrane to which the proteins to be tested for the presence of CoV-S in a non-denaturing PAGE (polyacrylamide gel electrophoresis) gel or SDS-PAGE (sodium dodecyl sulfate polyacrylamide gel electrophoresis) gel have been transferred (e.g., following electrophoretic separation in the gel). Before contacting the membrane with the anti-CoV-S antigen-binding protein, the membrane is optionally blocked, e.g., with non-fat dry milk or the like so as to bind non-specific protein binding sites on the membrane. (2) washing the membrane one or more times to remove unbound anti-CoV-S antigen-binding protein and other unbound substances; and (3) detecting the bound anti-CoV-S antigen-binding protein.

[0142] Detection of the bound antigen-binding protein indicates that the CoV-S protein is present on the membrane or substrate and in the sample. Detection of the bound antigen-binding protein may be by binding the antigen-binding protein with a secondary antibody (an anti-immunoglobulin antibody) which is detectably labeled and, then, detecting the presence of the secondary antibody label.

[0143] The anti-CoV-S antigen-binding proteins (e.g., antibodies and antigen-binding fragments (e.g., of Table 1)) disclosed herein may also be used for immunohistochemistry. Such a method forms part of the present invention and comprises, e.g., (1) contacting tissue to be tested for the presence of CoV-S protein with an anti-CoV-S antigen-binding protein of the invention; and (2) detecting the antigen-binding protein on or in the tissue.

[0144] If the antigen-binding protein itself is detectably labeled, it can be detected directly. Alternatively, the antigen-binding protein may be bound by a detectably labeled secondary antibody wherein the label is then detected.EXAMPLES

[0145] The following examples 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 methods and compositions of the invention and are not intended to limit the scope of what the inventors regard as their invention. Efforts have been made to ensure accuracy with respect to numbers used (e.g., amounts, temperature, etc.) but some experimental errors and deviations should be accounted for. Unless indicated otherwise, parts are parts by weight, molecular weight is average molecular weight, temperature is in degrees Centigrade, room temperature is about 25°C, and pressure is at or near atmospheric.Example 1: Generation of human antibodies to SARS-CoV-2 spike protein (SARS-CoV-2-S)

[0146] Human antibodies to SARS-CoV-2-Spike protein (SARS-CoV-2-S) were generated in a VELOCIMMUNE® mouse comprising DNA encoding human immunoglobulin heavy and kappa light chain variable regions or human immunoglobulin heavy and lambda light chain variable regions. Each mouse was immunized with a vector expressing the SARS-CoV-2-S receptor binding domain (RBD) (amino acids 1-1273 of NCBI accession number (MN908947.3), SEQ ID NO: 832), followed by a booster with a SARS-CoV-2-S vector or a SARS-CoV-2-S protein. The antibody immune response was monitored by a SARS-CoV-2-S-specific immunoassay. When a desired immune response was achieved, lymphocytes were harvested and fused with mouse myeloma cells to preserve their viability and form hybridoma cell lines. The hybridoma cell lines were screened and selected to identify cell lines that produce SARS-CoV-2-S-specific antibodies. Anti-SARS-CoV-2-S antibodies were also isolated directly from antigen-positive mouse B cells without fusion to myeloma cells, as described in U.S. Patent 7582298, herein specifically incorporated by reference in its entirety. Using this method, fully human anti-SARS-CoV-2-S antibodies (i.e., antibodies possessing human variable domains and human constant domains) were obtained.

[0147] Antibody variable regions were also isolated from human blood samples. Whole blood was received from patients 3-4 weeks after a laboratory-confirmed PCR positive test for SARS-CoV- 2 and symptomatic COVID-19 disease. Red blood cells were lysed using an ammonium chloride based lysis buffer (Life Technologies) and B cells were enriched by negative selection. Single B cells that bound the SARS-CoV-2 spike protein were isolated by fluorescent-activated cell sorting (FACS). Isolated B cells were single-well plated and mixed with antibody light and heavy variable region-specific PCR primers. cDNAs for each single B cell were synthesized via a reverse transcriptase (RT) reaction. Each resulting RT product was then split and transferred into two corresponding wells for subsequent antibody heavy and light chain PCRs. One set of the resulting RT products was first amplified by PCR using a 5' degenerate primer specific for antibody heavy variable region leader sequence or a 5' degenerate primer specific for antibody light chain variable region leader sequence and a 3' primer specific for antibody constant region, to form an amplicon. The amplicons were then amplified again by PCR using a 5' degenerate primer specific for antibody heavy variable region framework 1 or a 5' degenerate primer specific for antibody light chain variable region framework 1 and a 3' primer specific for antibody constant region, to generate amplicons for cloning. The antibody heavy chain and light chain derived PCR products were cloned into expression vectors containing heavy constant region and light constant region, respectively, thereby producing expression vectors for hybrid antibodies. The expression vectors expressing full-length heavy and light chain pairs were transfected into CHO cells to produce antibody proteins for testing.

[0148] The biological properties of exemplary antibodies generated in accordance with the methods of this Example are described in detail in the Examples set forth below.Example 2: Heavy and light chain variable region amino acid and nucleotide sequences

[0149] Table 1 sets forth the amino acid sequence identifiers of the heavy and light chain variable regions and CDRs, as well as the heavy chain and light chain sequences, of exemplary anti-SARS-CoV-2-S antibodies. The corresponding nucleic acid sequence identifiers are set forth in Table 2. Table 1: Amino Acid Sequence Identifiers SEQ ID NOs Antibody Designation HCVR HCDR1 HCDR2 HCDR3 LCVR LCDR1 LCDR2 LCDR3 HC LC mAb109132468101214161820mAb1091522242628303234363840mAb109162468101214164220mAb1091744462649515355575961mAb1091822242628303234366340mAb 1092065676971737555777981mAb1092183852687899155939597mAb 1092299101103105107109111113115117mAb 1092311912112312512712955131133135mAb 10924137139141143145147149151153155mAb 10925656769717375557715781mAb 10926838526878991559315997mAb1092799101103105107109111113161117mAb 1092811912112312512712955131163135mAb 10929137139141143145147149151165155mAb1093016716917117317512955177179181mAb1093116716917117317512955177183181mAb109321851872618919175194196198200mAb1093320220420620821021255214216218mAb10934220222224226228230194232234236mAb109352382426240242244194246248250mAb1093625225425625826012955262264266mAb1093726827027227427612955278280282mAb 10940284169286288290292294296298300mAb109383022426304306308194310312314mAb1093931618731932132332555327329331mAb109413338526336338340294296342344mAb109421851872618919175194196346200mAb1094320220420620821021255214348218mAb 10944220222224226228230194232350236mAb109452382426240242244194246352250mAb 1094625225425625826012955262354266mAb1094726827027227427612955278356282mAb 109483022426304306308194310358314mAb 1094931618731932132332555327360331mAb109513338526336338340294296362344mAb10950284169286288290292294296364300mAb109543668526370372244194375377379mAb1095538138326385387389194310392394mAb1095639618726399401389194403405407mAb10957409411264144165355418420422mAb109583668526370372244194375424379mAb1095938138326385387389194310426394mAb 1096039618726399401389194403428407mAb10961409411264144165355418430422mAb 1096443243443643844044255445447449mAb 109654514532645545745934462464466mAb 1096646818726470472389194474476478mAb 1096748024483485487389194489491493mAb 10969495497499501503389194214506508mAb109705102426512514516194518520522mAb109715244112652853053255534536538mAb 1097343243443643844044255445540449mAb109744514532645545745934462542466mAb1097546818726470472389194474544478mAb1097648024483485487389194489546493mAb 10977548550552554556558294560562564mAb 10978495497499501503389194214566508mAb 109795102426512514516194518568522mAb 109805244112652853053255534570538mAb10981548550552554556558294560572564mAb 10982574187576578580582584586588590mAb 10983574187576578580582584586592590mAb 10984594596265986001214602604606mAb10985608169610612614616584618620622mAb 1098662462626628630582632634636638mAb10987640642499644646648650652654656mAb10988658660662664666668670672674676mAb10989678680682684686688650690692694mAb 10990594596265986001214602696606mAb10991608169610612614616584618698622mAb 1099262462626628630582632634700638mAb 10993640642499644646648650652702656mAb 10994658660662664666668670672704676mAb 10995678680682684686688650690706694mAb 10996708242671171312955715717719mAb 10997708242671171312955715721719mAb 109987231872672572712955729731733mAb 109997231872672572712955729735733mAb11000737242673974174355745747749mAb11001737242673974174355745751749mAb11002753242675571312955715757719mAb11003753242675571312955715759719mAb10914444626495153555776261mAb110047647664997687709155772774776mAb110057647664997687709155772778776mAb11006780782267847865355788790792mAb11007780782267847865355788794792mAb1100879624267988005355802804806mAb1100979624267988005355802808806mAb11010810812814816818129820822824826mAb11011810812814816818129820822828826 Table 2: Nucleic Acid Sequence Identifiers SEQ ID NOs Antibody Designation HCVR HCDR 1 HCDR 2 HCDR 3 LCVR LCDR 1 LCDR 2 LCDR 3 HC LC mAb10913135791113151719mAb1091521232527293133353739mAb10916135791113154119mAb1091743454748505254565860mAb1091821232527293133356239mAb 1092064666870727454767880mAb1092182844786889054929496mAb 1092298100102104106108110112114116mAb 1092311812012212412612854130132134mAb 10924136138140142144146148150152154mAb 10925646668707274547615680mAb 10926828447868890549215896mAb1092798100102104106108110112160116mAb 1092811812012212412612854130162134mAb 10929136138140142144146148150164154mAb1093016616817017217412854176178180mAb1093116616817017217412854176182180mAb1093218418647188190192193195197199mAb1093320120320520720921154213215217mAb10934219221223225227229193231233235mAb109352372347239241243193245247249mAb1093625125325525725912854261263265mAb1093726726927127327512854277279281mAb 10940283168285287289291293295297299mAb109383012347303305307193309311313mAb1093931531731832032232454326328330mAb1094133233447335337339293295341343mAb1094218418647188190192193195345199mAb1094320120320520720921154213347217mAb 10944219221223225227229193231349235mAb109452372347239241243193245351249mAb 1094625125325525725912854261353265mAb1094726726927127327512854277355281mAb 109483012347303305307193309357313mAb 1094931531731832032232454326359330mAb1095133233447335337339293295361343mAb10950283168285287289291293295363299mAb10954365367368369371373193374376378mAb1095538038247384386388193390391393mAb1095639539747398400388193402404406mAb109574084104124134155254417419421mAb10958365367368369371373193374423378mAb1095938038247384386388193390425393mAb 1096039539747398400388193402427406mAb109614084104124134155254417429421mAb 10964431433435437439441443444446448mAb 1096545045247454456458460461463465mAb 10966467397412469471388193473475477mAb10967479481482484486388193488490492mAb 10969494496498500502388193504505507mAb10970509481412511513515193517519521mAb1097152352552652752953154533535537mAb 10973431433435437439441443444539448mAb1097445045247454456458460461541465mAb10975467397412469471388193473543477mAb10976479481482484486388193488545492mAb 10977547549551553555557293559561563mAb 10978494496498500502388193504565507mAb 10979509481412511513515193517567521mAb 1098052352552652752953154533569537mAb10981547549551553555557293559571563mAb 10982573186575577579581583585587589mAb 10983573186575577579581583585591589mAb 10984593595475975991113601603605mAb10985607168609611613615583617619621mAb 1098662362547627629581631633635637mAb10987639641498643645647649651653655mAb10988657659661663665667669671673675mAb10989677679681683685687649689691693mAb 10990593595475975991113601695605mAb10991607168609611613615583617697621mAb 1099262362547627629581631633699637mAb 10993639641498643645647649651701655mAb 10994657659661663665667669671703675mAb 10995677679681683685687649689705693mAb 109967077094771071212854714716718mAb 109977077094771071212854714720718mAb 109987221864772472612854728730732mAb 109997221864772472612854728734732mAb11000736234773874074254744746748mAb11001736234773874074254744750748mAb11002752234775471212854714756718mAb11003752234775471212854714758718mAb109147604547485052545676160mAb110047637654987677699054771773775mAb110057637654987677699054771777775mAb11006779781477837855254787789791mAb11007779781477837855254787793791mAb11008795709477977995254801803805mAb11009795709477977995254801807805mAb11010809811813815817128819821823825mAb11011809811813815817128819821827825

[0150] Antibodies disclosed herein have fully human variable regions but can have mouse constant regions (e.g., a mouse IgG1 Fc or a mouse IgG2 Fc (a or b isotype)) or human constant regions (e.g., a human IgG1 Fc or a human IgG4 Fc). As will be appreciated by a person of ordinary skill in the art, an antibody having a particular Fc isotype can be converted to an antibody with a different Fc isotype (e.g., an antibody with a mouse IgG1 Fc can be converted to an antibody with a human IgG4, etc.), but in any event, the variable domains (including the CDRs) - which are indicated by the numerical identifiers shown in Tables 1 and 2 will remain the same, and the binding properties to antigen are expected to be identical or substantially similar regardless of the nature of the constant domain.

[0151] The variable regions of antibodies derived from VELOCIMMUNE® mice and from human samples were sequenced by Next Generation Sequencing and the repertoire for heavy and light chain pairs was identified (Figure 10A and Figure 10B). The predominant lineage of VI antibodies utilized VH3-53 paired with VK1-9, VK1-33, or VK1-39 while human-derived antibodies utilized VH3-66 paired with VK1-33 or VH2-70 paired with VK1-39. Further analysis of overlaid sequences showed strong overlap in the repertoire of isolated kappa chains between VI and human-derived antibodies. Although the repertoire of Lambda chains did not overlap well, that may be due to only two lambda mice being included in this trial. The average CDR length for heavy chain was similar between VI and human derived antibodies with an average length of 13 and 14.5 amino acids, respectively. Average kappa CDR length was the same for VI and human derived antibodies at 9 amino acids and was close for lambda chains with an average length of 11.1 and 10.6 amino acids, respectively. Availability of humanized mouse and human-derived antibodies allowed for more diversity of V genes and enabled the later identification of noncompeting antibodies.

[0152] As described above, the antibodies were obtained from hybridomas generated from VELOCIMMUNE® mice, by direct isolation from antigen-positive VELOCIMMUNE® mouse B cells, or derived from variable regions cloned from antigen-positive human B cells. A summary of these sources is shown in Table 3. Table 3: Antibody / Variable Region sources Antibody Source mAb10913mouse B cellsmAb10915mouse B cellsmAb10916mouse B cellsmAb10917mouse B cellsmAb10918mouse B cellsmAb 10920mouse B cellsmAb 10921mouse B cellsmAb 10922mouse B cellsmAb 10923mouse B cellsmAb 10924mouse B cellsmAb 10925mouse B cellsmAb 10926mouse B cellsmAb 10927mouse B cellsmAb 10928mouse B cellsmAb 10929mouse B cellsmAb10930mouse B cellsmAb10931mouse B cellsmAb 10932mouse B cellsmAb 10933mouse B cellsmAb 10934mouse B cellsmAb10935mouse B cellsmAb10936mouse B cellsmAb 10937mouse B cellsmAb 10940mouse B cellsmAb10938mouse B cellsmAb10939mouse B cellsmAb10941mouse B cellsmAb10942mouse B cellsmAb10943mouse B cellsmAb 10944mouse B cellsmAb 10945mouse B cellsmAb 10946mouse B cellsmAb10947mouse B cellsmAb 10948mouse B cellsmAb 10949mouse B cellsmAb10951mouse B cellsmAb10950mouse B cellsmAb10954human B cellsmAb10955human B cellsmAb10956human B cellsmAb10957human B cellsmAb10958human B cellsmAb10959human B cellsmAb 10960human B cellsmAb10961human B cellsmAb 10964human B cellsmAb 10965human B cellsmAb 10966human B cellsmAb10967human B cellsmAb 10969human B cellsmAb10970human B cellsmAb10971human B cellsmAb 10973human B cellsmAb10974human B cellsmAb10975human B cellsmAb10976human B cellsmAb 10977human B cellsmAb 10978human B cellsmAb 10979human B cellsmAb 10980human B cellsmAb10981human B cellsmAb 10982mouse B cellsmAb 10983mouse B cellsmAb 10984human B cellsmAb10985human B cellsmAb 10986human B cellsmAb10987human B cellsmAb10988human B cellsmAb10989human B cellsmAb 10990human B cellsmAb10991human B cellsmAb 10992human B cellsmAb 10993human B cellsmAb 10994human B cellsmAb10995human B cellsmAb 10996hybridomamAb 10997hybridomamAb 10998hybridomamAb 10999hybridomamAb11000hybridomamAb11001hybridomamAb11002hybridomamAb11003hybridomamAb10914mouse B cellsmAb11004hybridomamAb11005hybridomamAb11006hybridomamAb11007hybridomamAb11008hybridomamAb11009hybridomamAb11010hybridomamAb11011hybridoma Example 3: Characterization of hybridoma supernatants by binding ELISA

[0153] An ELISA binding assay was performed to identify antibody supernatants that bound to the SARS-CoV-2-Spike protein receptor binding domain (RBD). A protein composed of the RBD of SARS-CoV-2 (amino acids 319-541) expressed with a 6X histidine tag and two myc epitope tags at the C-terminus (SARS-CoV-2-S-RBD-mmH; see also NCBI Accession Number MN908947.3) was coated at 1 µg / ml on a 96-well plate in PBS buffer overnight at 4°C. Nonspecific binding sites were subsequently blocked using a 0.5% (w / v) solution of BSA in PBS. Antibody supernatants or media alone were diluted 1:40 or 1 :50 in the PSA+0.5% BSA blocking buffer and transferred to the washed microtiter plates. After one hour of incubation at room temperature, the wells were washed, and plate-bound supernatant was detected with either goat-anti-human IgG antibody conjugated with horseradish peroxidase (HRP) (Jackson Immunoresearch), or anti-mouse IgG antibody conjugated with horseradish peroxidase (HRP) (Jackson Immunoresearch). The plates were then developed using TMB substrate solution (BD Biosciences) according to manufacturer's recommendation and absorbance at 450nm was measured on a Victor X5 plate reader.

[0154] The ability of anti-SARS-CoV-2-S antibodies to bind the receptor binding domain of SARS-CoV-2-S (SARS-CoV-2-S-RBD) was assessed, as described above, using a binding ELISA with the SARS-CoV-2-S-RBD-mmH protein coated on a microplate. Single point antibody supernatant binding to SARS-COV-2-S-RBD-mmH coated on 96-well microtiter plates was detected with an HRP conjugated anti-hFc or anti-mFc antibody.

[0155] The binding results of three trials are summarized in Table 4. The SARS-CoV-2 binding signals (absorbance 450nm) are indicated, with the media only background provided as a negative reference per experiment. A sample marked IC (Inconclusive) had an experimental anomaly to the plate and is therefore reported without a value. As shown in comparison to the media only control, the supernatants tested showed substantial binding to the SARS-CoV-2-S-RBD. Table 4: Supernatant binding to SARS-CoV-2 spike protein receptor binding domain Supernatant Supernatant Dilution Detection Antibody Binding Signal (absorbance at 450nm) mAb109131:50a-hFc2.752mAb109141:50a-hFc2.857mAb109151:50a-hFc2.76mAb109321:50a-hFc2.718mAb109331:50a-hFc2.762mAb109341:50a-hFc2.688mAb109351:50a-hFc2.676mAb109361:50a-hFc2.644mAb109371:50a-hFc2.664mAb 109201:50a-hFc2.683mAb109211:50a-hFc2.633mAb 109221:50a-hFc2.595mAb 109231:50a-hFc2.353mAb 109241:50a-hFc2.269mAb109301:50a-hFc2.451mAb109381:50a-hFc2.536mAb109391:50a-hFc2.516mAb 109401:50a-hFc2.77mAb109411:50a-hFcICmAb 109821:50a-hFc2.537mAb 109841:50a-hFc0.716mAb109851:50a-hFc2.35mAb 109861:50a-hFc2.331mAb109871:50a-hFc2.438mAb109881:50a-hFc3.062mAb109891:50a-hFc3.116mAb 109691:50a-hFc2.629mAb109701:50a-hFc2.807mAb109711:50a-hFc3.052mAb 109641:50a-hFc3.086mAb 109651:50a-hFc2.918mAb 109661:50a-hFc0.421mAb109671:50a-hFc1.732mAb109541:50a-hFc1.963mAb109551:50a-hFc2.469mAb109561:50a-hFc2.6mAb109571:50a-hFc2.49mAb 109771:50a-hFc2.925mAb110101:40a-mFc2.896mAb110041:40a-mFc2.908mAb110001:40a-mFc2.725mAb110061:40a-mFc2.619mAb110081:40a-mFc2.907mAb 109981:40a-mFc2.835mAb 109961:40a-mFc2.826mAb110021:40a-mFc2.581Media only1:50a-hFc0.069Media only1:40a-mFc0.058Media only1:50a-hFc0.055 Example 4: Antibody binding to SARS-CoV-2-S-expressing virus-like particle

[0156] To investigate the ability of a panel of anti-SARS-CoV-2-S monoclonal antibodies to bind the SARS-CoV-2 spike glycoprotein, an in vitro binding assay utilizing SARS-CoV-2 spike protein-expressing viral-like particles (VLPs) in an electrochemiluminescence based detection platform (MSD) was developed.

[0157] To transiently express the SARS-CoV-2 spike protein (NCBI Accession number MN908947.3, amino acids 16-1211; SEQ ID NO: 833), Vesicular stomatitis virus (VSV) lacking glycoprotein G (VSV delta G) was pseudotyped with SARS-CoV-2 spike protein (VSV-SARS-CoV-2-S) and generated in HEK293T cells. As a negative binding control, VSV delta G was pseudotyped with VSV G protein (VSV-G).

[0158] Experiments were carried out according to following procedure. The two types of VLPs described above were diluted in PBS, seeded into 96-well carbon electrode plates (MULTI-ARRAY high bind plate, MSD), and incubated overnight at 4 °C to allow the VLPs to adhere. Nonspecific binding sites were blocked by 2% BSA (w / v) in PBS for 1 hour at room temperature. Supernatants containing antibodies produced from SARS CoV-2-immunized mice or infected human sera, along with media-only controls which were diluted 1:10 or 1:20 in 1x PBS + 0.5% BSA buffer, were added to the plate-bound particles. The plates were then incubated for 1 hour at room temperature with shaking, after which the plates were washed with 1x PBS to remove the unbound antibodies using an AquaMax2000 plate washer (MDS Analytical Technologies). The plate-bound antibodies were detected with a SULFO-TAGTM-conjugated anti-human IgG antibody (Jackson Immunoresearch) or a SULFO-TAGTM-conjugated anti-mouse IgG antibody (Jackson Immunoresearch) for 1 hour at room temperature. After washes, the plates were developed with the Read Buffer (MSD) according to manufacturer's recommended procedure and the luminescent signals were recorded with a SECTOR Imager 600 (Meso Scale Development) instrument. Direct binding signals (in RLU) were captured, and a ratio of SARS-CoV-2-S-expressing VLPs to the irrelevant VLP was calculated.

[0159] The ability of the anti-SARS-CoV-2-S monoclonal antibodies to bind to SARS-CoV-2-S-expressing VLPs compared with binding to irrelevant VSV-expressing VLPs was assessed using an immunobinding assay, as described above. Single-point binding to the immobilized VLPs on 96-well High Bind plates (MSD) was performed with an antibody supernatant dilution of 1:10 or 1:20, bound for 1 hour, and detected using SULFO-TAGTM-conjugated anti-human IgG or anti-mouse IgG antibody. The binding signals from electrochemiluminescence were recorded on a Sector Imager 600 (MSD). RLU values were determined for the antibody binding to VLPs. Ratios were calculated comparing the SARS-CoV-2-S-expressing VLP binding signals to control VLPs.

[0160] The binding results from three experiments are summarized in Table 5. A signal observed from SARS-COV-2-S-expressing VLPs indicates binding, while comparison with negative VLPs provides a relative background. Media alone samples provide baseline signals of secondary antibody binding to samples with no supernatant. The 46 antibodies bound specifically at >4-fold higher than the media-only samples (20-35 RLU) on the SARS-CoV-2-S-expressing VLPs, with a range of binding signals from 85-13,600 RLU. The ratios of SARS-CoV-2-S-expressing VSV: VSV-VLPs (negative control) ranged from 1.1- 22.7, with many having high background on VSV-VLPs. The ratio of mAb11002 of 0.9 is likely due to a low concentration of monoclonal antibody in the supernatant sample. Table 5: SARS-CoV-2-S VLP binding Supernatant Supernatant Dilution Secondary Detection Antibody vsv-VLP Binding Signal (RLU) VSV-SARS-CoV-2-S VLP Binding Signal (RLU) Ratio of Binding Signals: VSV-SARS-CoV-2-S / VSV-VLP mAb109131:10a-hFc215532441.5mAb109141:10a-hFc388551811.3mAb109151:10a-hFc98090229.2mAb109321:10a-hFc9891045110.6mAb109331:10a-hFc5079661.9mAb109341:10a-hFc387650411.3mAb109351:10a-hFc208738671.9mAb109361:10a-hFc232580763.5mAb109371:10a-hFc140419201.4mAb 109201:10a-hFc8366100411.2mAb109211:10a-hFc119454364.6mAb 109221:10a-hFc147322291.5mAb 109231:10a-hFc122418591.5mAb 109241:10a-hFc4879692mAb109301:10a-hFc176932071.8mAb109381:10a-hFc123266235.4mAb109391:10a-hFc177750742.9mAb 109401:10a-hFc60620723.4mAb109411:10a-hFc67345886.8mAb 109821:10a-hFc117820161.7mAb 109841:10a-hFc248689893.6mAb109851:10a-hFc204932791.6mAb 109861:10a-hFc2044108315.3mAb109871:10a-hFc183924501.3mAb109881:10a-hFc183223051.3mAb109891:10a-hFc67219993mAb 109691:10a-hFc309633131.1mAb109701:10a-hFc136457124.2mAb109711:10a-hFc113572666.4mAb 109641:10a-hFc143986016mAb 109651:10a-hFc74313701.8mAb 109661:10a-hFc142865744.6mAb109671:10a-hFc144695106.6mAb109541:10a-hFc64163089.8mAb109551:10a-hFc93217881.9mAb109561:10a-hFc103015811.5mAb109571:10a-hFc60455449.2mAb 109771:10a-hFc4141136003.3mAb110101:20a-mFc963633.8mAb110041:20a-mFc1104063.7mAb110001:20a-mFc3335921.8mAb110061:20a-mFc165374722.7mAb110081:20a-mFc1033243.1mAb 109981:20a-mFc742182.9mAb 109961:20a-mFc51851.7mAb110021:20a-mFc1561460.9Media only1:10a-hFc30351.2Media only1:20a-mFc35200.6Media only1:10a-hFc39290.7 Example 5: Antibody neutralization of VSV-SARS-CoV-2-S pseudovirus infectivity

[0161] To investigate the ability of a panel of anti-SARS-CoV-2-S monoclonal antibodies to neutralize SARS-CoV-2, an in vitro neutralization assay utilizing VSV-SARS-CoV-2-S pseudovirus was developed.

[0162] As described above, VSV pseudotype viruses were generated by transiently transfecting 293T cells with a plasmid encoding for SARS-CoV-2 spike protein. Cells were seeded in 15 cm plates at 1.2x10 7< cells per plate in DMEM complete media one day prior to transfection with 15 µg / plate spike protein DNA using 125 µL Lipofectamine LTX, 30 µL PLUS reagent, and up to 3 mL Opti-Mem. 24 hours post transfection, the cells were washed with 10 mL PBS, then infected with an MOI of 0.1 VSV ΔG:mNeon< virus in 10 mL Opti-Mem. Virus was incubated on cells for 1 hour, with gentle rocking every 10 minutes. Cells were washed 3 times with 10 mL PBS, then overlaid with 20 mL Infection media before incubation at 37 C, 5% CO 2 for 24 hours. Supernatant was collected into 250 mL centrifuge tubes on ice, then centrifuged at 3000 rpm for 5 minutes to pellet any cellular debris, aliquoted on ice, then frozen to -80 °C. Infectivity was tested on Vero cells prior to use in neutralization assays. This material will be referred to as VSV-SARS-CoV-2-S.Neutralization assay with VSV-SARS-CoV-2-S

[0163] On day 1, Vero cells were seeded at 80% confluency in T225 flasks. To seed cells, media was removed from the cells, the cells were washed with 20mL PBS (Gibco: 20012-043), and 5mL TrypLE was added and incubated for ∼5 minutes at 37 °C until the cells dislodged. 5 mL of complete DMEM was added to inactivate the trypsin, and pipetted up and down to distribute the cells. To count the resuspended cells, 20,000 Vero cells were plated in 100 µL prewarmed Complete DMEM per well in a 96 Well Black Polystyrene Microplate (Corning: 3904).

[0164] On day 2, VSV-SARS-CoV-2-S was thawed on ice and diluted 1:1 with infection media.

[0165] In a V-bottom 96 well plate, a dilution of each supernatant was generated in 60ul infection media. For media (negative) controls, 60 µl of diluted conditioned media was added to the wells. 60 µL of diluted VSV-SARS-CoV-2-S were added to every well except the media control wells. To those wells, 60 µL of infection media was added. Pseudoviruses were then incubated with supernatant dilutions for 30 minutes at room temperature. Media was removed from the Vero cell plates, 100 µL of supernatant / pseudovirus mixtures were transferred to the cells, and the plate was incubated at 37 °C, 5% CO 2 for 24 hours. The final supernatant dilutions of 1:4 and 1:20, and for some samples 1:100, were used to assess neutralization of VSV-SARS-CoV-2-S pseudoviruses.

[0166] On day 3, after the 24 hr incubation, supernatant was removed from the cell wells and replaced with 100 µL of PBS. The plates were then read on a SpectraMax i3 with MiniMax imaging cytometer.

[0167] The ability of the anti-SARS-CoV-2-S antibodies to neutralize VSV-based SARS-CoV-2-S-expressing pseudotyped virus was assessed using a neutralization fluorescence focus assay. The binding results of three assays are summarized below. The neutralization potency of antibody at each dilution is represented as a percentage compared to mock supernatant control. All antibodies demonstrated neutralization capacity, and particularly for the set of antibodies that were evaluated 1:100, those showing higher neutralization may represent more potent neutralization capacity. Table 6: Neutralization of VLPs Supernatant Neutralization (1:4 dilution) Neutralization (1:20 dilution) Neutralization (1:100 dilution) mAb1091399.595.569.1mAb1091494.274.843.6mAb1091596.774.229.6mAb1093299.894.668mAb1093399.898.988.4mAb1093499.999.898.4mAb1093599.698.588.8mAb1093699.799.192.9mAb1093797.587.756.3mAb 1092099.595.569.1mAb1092198.291.446.1mAb 1092299.899.188.4mAb 1092399.592.967.7mAb 1092498.185.455.2mAb1093099.191.159mAb1093898.18354.2mAb1093998.690.564mAb 109409789.966.4mAb1094198.992.973.8mAb 1098297.483.844.5mAb 1098499.895.183.4mAb1098599.788.463.5mAb 1098699.79886mAb1098799.397.794.6mAb1098897.687.662.2mAb1098910099.898.2mAb 1096997.29163.7mAb1097099.696.782.4mAb1097199.59773.9mAb 1096499.799.794.1mAb 1096598.587.668.6mAb 1096699.595.576.2mAb1096798.991.469.2mAb1095499.89670.7mAb1095598.888.662.7mAb1095697.184.161.6mAb1095797.676.448mAb 1097795.57947.7mAb110108554NTmAb110047740NTmAb110009882NTmAb110069154NTmAb110089677NTmAb 109988859NTmAb 109968558NTmAb1100235-1NT*NT: not tested Example 6: Characterization of antibodies in an antibody-dependent cell-mediated toxicity surrogate assay

[0168] The ability of antibodies targeting the spike protein of SARS-CoV-2 to interact with FcγR3a, an Fc-receptor prominently expressed on natural killer (NK) cells that induces antibody dependent cell-mediated cytotoxicity (ADCC), was measured in a surrogate bioassay using reporter cells and target cells bound to antibodies. This assay used Jurkat T cells that were engineered to express the reporter gene luciferase under the control of the transcription factor NFAT (NFAT-Luc) along with the high affinity human FcγR3a 176< Val allotype receptor (Jurkat / NFAT-Luc / hFcγR3a 176< Val). Target cells were engineered Jurkat T cells expressing human CD20 (used as a positive control with a CD20-targeting human IgG1 antibody) and the full-length SARS-CoV-2 spike protein controlled by a doxycycline-inducible promoter. Reporter cells were incubated with target cells and engagement of FcγR3a via the Fc domain of human IgG1 antibodies bound to target cells led to the activation of the transcription factor NFAT in the reporter cells and drove the expression of luciferase which was then measured via a luminescence readout.

[0169] Jurkat T cells were engineered to constitutively express full length human CD20 (amino acids M1-P297 of NCBI accession number NP_690605.1), Tet3G transactivator protein (cloned using a Takara pEF1α-Tet3G Vector, Catalog # 631167), as well as a doxycycline-inducible full-length SARS-CoV-2 spike protein (amino acids M1-T1273 of NCBI accession number YP_009724390.1). Engineered Jurkat / Tet3G / hCD20 / SARS-CoV2 spike protein-expressing cells were sorted for high expression of the spike protein and subsequently maintained in RPMI + 10% Tet-free FBS + P / S / G + 500 µg / ml G418 + 1 µg / ml puromycin + 250 µg / ml hygromycin growth medium.

[0170] Jurkat T cells were engineered to stably express a Nuclear Factor of Activated T-cells (NFAT) luciferase reporter construct along with the high affinity human FcγR3a 176< Val allotype receptor (amino acids M1-K254 of NCBI accession number P08637 VAR_003960). Engineered reporter cells were maintained in RPMI1640 + 10% FBS + P / S / G + 0.5 (µ / ml puromycin + 500 (µg / ml G418 growth media.

[0171] 36 hours prior to the start of the surrogate ADCC assay, 5 x 10 5< target cells / ml were induced in RPMI + 10% Tet-free FBS + P / S / G cell culture media containing 1 µg / ml doxycycline (Sigma). A day before the experiment, reporter cells were split to a density of 7.5 x 10 5< cells / ml in RPMI 1640 + 10% FBS + P / S / G + 0.5 µg / ml puromycin + 500 µg / ml G418 growth media.

[0172] Briefly, on the day of the experiment, the target and reporter cells were transferred into assay media (RPMI + 10% Tet-free FBS + P / S / G) and added at a 3:2 ratio (3 x 10 4< / well target cells and 2 x 10 4< / well reporter cells) to 384-well white microtiter plates, followed by the addition of anti-SARS-CoV-2-S antibody supernatant of varying concentrations. A positive control (CD20 antibody with human IgG1) sample and a negative control sample containing no antibody was included on each plate to normalize detected ADCC activities of anti-SARS-CoV-2-S antibody supernatants. Plates were incubated at 37 °C / 5% CO 2 for 5 h followed by the addition of an equal volume of ONE-Glo™ (Promega) reagent to lyse cells and detect luciferase activity. The emitted light was captured in Relative Light Units (RLU) on a multi-label plate reader Envision (PerkinElmer), and data was analyzed and normalized using the following equation: ADCC activity % = 100 × Mean RLU test samples − Mean RLU background signal Mean RLU positive control − Mean RLU background signal

[0173] The ability of anti-SARS-COV-2-S antibodies to activate FcyR3a receptors was evaluated in a surrogate ADCC assay using Jurkat / NFAT-Luc / FcγR3a 176< Val) as reporter cells and Jurkat / hCD20 / SARS-CoV2 Spike as target cells. Each antibody tested contained an IgG1 domain.

[0174] Table 7 summarizes the results, showing the raw luciferase activity and the calculated % of positive control are indicated. A range of % ADCC activity was observed indicating FcγR3a activation by the antibody supernatants. All samples demonstrated some measure of surrogate ADCC activity, and 10 of the antibody supernatants demonstrated surrogate ADCC activity better than observed in positive controls. Table 7: ADCC surrogate activity of anti-SARS-CoV-2-S antibody supernatants. mAb ADCC Mean RLU ADCC (Activity (%) mAb 1091311,480111.9mAb1091421,960265.8mAb1091514,280153mAb1093213,020108.8mAb109339,74068.5mAb1093411,68092mAb1093511,54090.4mAb1093615,160133.8mAb1093712,340100.1mAb 1092015,480137.8mAb1092110,08067.7mAb 109229,14056.3mAb 1092313,340107.1mAb 109247,22033mAb109308,90053.4mAb1093812,960102.5mAb109399,44059.7mAb 1094012,520106.2mAb1094110,34077.2mAb 109827,90059.4mAb 1098467806.8mAb1098558402.8mAb 1098662004.4mAb109871202029.4mAb1098872008.7mAb109891020021.5mAb 109691050023.1mAb10970764010.6mAb10971748010mAb 1096463805.1mAb 1096567806.9mAb 10966708010.4mAb1096767408.6mAb1095469409.8mAb1095567408.7mAb1095667608.8mAb10957712010.8mAb 109771298033.8 Example 7: Anti-SARS-CoV-2-S antibody binding specificity assay

[0175] A Luminex binding assay was performed to determine the binding of anti-SARS-COV-2-S antibodies to a panel of antigens. For this assay, antigens were amine-coupled or captured by streptavidin to Luminex microspheres as follows: approximately 10 million MagPlex microspheres (Luminex Corp., MagPlex Microspheres, Cat. No. MC10000 and MC12000), were resuspended by vortexing in 500 µL 0.1M NaPO 4 , pH 6.2 (activation buffer) and then centrifuged to remove the supernatant. Microspheres were protected from light, as they are light sensitive. The microspheres were resuspended in 160 µL of activation buffer and the carboxylate groups (-COOH) were activated by addition of 20 µL of 50 mg / mL of N-hydroxysuccinimide (NHS, Thermo Scientific, Cat. No. 24525) followed by addition of 20 µL of 50 mg / mL 1-ethyl-3-[3-dimethylaminopropyl]carbodiimide (EDC, ThermoScientific, Cat. No. 22980) at 25 °C. After 10 minutes, the pH of the reaction was reduced to 5.0 with the addition of 600 µL 50 mM MES, pH 5 (coupling buffer), and the microspheres were vortexed and centrifuged to remove supernatant. The activated microspheres were immediately mixed with 500 µL of 25 µg / mL of the protein antigen or Streptavidin in coupling buffer and incubated for two hours at 25 °C. The coupling reaction was quenched by addition of 50 µL of 1M Tris-HCl, pH 8.0 and the microspheres were vortexed, centrifuged, and washed three times with 800 µL of PBS 0.005% (Tween20 0.05%), to remove uncoupled proteins and other reaction components. Microspheres were resuspended in 1 mL of PBS 2% BSA 0.05% Na Azide at 10 million microspheres / mL. For Streptavidin capture of antigens, 500 µL of 12.5 µg / mL of biotinylated protein in PBS was added to Streptavidin-coupled microspheres and incubated for one hour at 25 °C. Microspheres were vortexed, centrifuged, and washed three times with 800 µL of PBS, and then blocked using 500 µL 30mM Biotin (Millipore-Sigma, Cat. No. B4501) in 0.15M Tris pH 8.0. Microspheres were incubated for 30 minutes then vortexed, centrifuged, and washed three times with 800 µL of PBS. Microspheres were resuspended in 1 mL of PBS 2% BSA 0.05% Na Azide at 10 million microspheres / mL.

[0176] Microspheres for the different proteins and biotinylated proteins were mixed at 2700 beads / ml, and 75 µL of microspheres were plated per well on a 96 well ProcartaPlex flat bottom plate (ThermoFisher, Cat. No: EPX-44444-000) and mixed with 25 µL of individual anti-SARS-CoV-2 supernatant containing antibody. Samples and microspheres were incubated for two hours at 25°C and then washed twice with 200 µL of DPBS with 0.05% Tween 20. To detect bound antibody levels to individual microspheres, 100 µL of 2.5 µg / mL R-Phycoerythrin conjugated goat F(ab')2 anti-human kappa (Southern Biotech, Cat# 2063-09) in blocking buffer (for antibodies with murine Fc regions) or 100 µL of 1.25 µg / mL R-Phycoerythrin AffiniPure F(ab') 2 Fragment Goat Anti-Mouse IgG, F(ab') 2 Fragment Specific (Jackson Immunoresearch, Cat. No: 115-116-072) in blocking buffer (for antibodies with human Fc regions), was added and incubated for 30 minutes at 25 °C. After 30 minutes, the samples were washed twice with 200 µl of washing buffer and resuspended in 150 µL of wash buffer. The plates were read in a Luminex FlexMap 3D® (Luminex Corp.) and Luminex xPonent® software version 4.3 (Luminex Corp.). The SARS-CoV-2 proteins used in the assay are as follows: RBD_(R319-F541).mmh: SEQ ID NO: 829 RBD_(R319-F541).mFc: SEQ ID NO: 830 RBD_(R319-F541).hFc): SEQ ID NO: 831

[0177] The results of the Luminex binding are shown in Table 8 and Table 9 as median fluorescence intensity (MFI) signal intensities. The results show that the 46 anti-SARS-CoV-2-S antibody supernatants bound specifically to SARS-CoV-2-S RBD proteins. These results also show that five of these antibodies cross-react with SARS Coronavirus spike RBD proteins with binding signal greater than 1000 MFI. Example 8: Anti-SARS-CoV-2-S antibody diversity assay

[0178] A binding assay was performed to determine the binding profile of anti-SARS-COV-2-S antibodies. For this assay, antigens were amine coupled as described for the Luminex binding assay above. Briefly, approximately 9 million MagPlex microspheres for 16 different bead regions (Luminex Corp., MagPLex Microspheres, Cat. No. MagPLex MC10000 and MC12000), were resuspended by vortexing in 500 µL 0.1M NaPO 4 , pH 6.2 and then centrifuged to remove the supernatant. The microspheres were resuspended in 160 µL of activation buffer and the carboxylate groups (-COOH) were activated by addition of 20 µL of 50 mg / mL of N-hydroxysuccinimide (NHS, Thermo Scientific, Cat# 24525) followed by addition of 20 µL of 50 mg / mL of 1-ethyl-3-[3-dimethylaminopropyl]carbodiimide (EDC, ThermoScientific, Cat# 22980) at 25°C. After 10 minutes, the pH of the reaction was reduced to 5.0 with the addition of 600 µL of 50 mM MES, pH 5 (coupling buffer), and the microspheres were vortexed and centrifuged to remove supernatant. The activated microspheres were immediately mixed with 500 µL of 20 µg / mL of SARS-CoV-2 Spike Protein (RBD)(R319-F541)-mmH in coupling buffer and incubated for two hours at 25°C. The coupling reaction was quenched by addition of 50 µL of 1M Tris-HCl, pH 8.0 and the microspheres were vortexed, centrifuged, and washed three times with 1000 µL of PBS. Microspheres were resuspended in 250 µL of PBS at 9 million microspheres / mL.

[0179] 15 out of the 16 microsphere regions with amine-coupled protein were modified for the binning assay as follows: microspheres were washed twice with PBS 5% DMSO, and 500 µl of a chemical or enzyme were dissolved per manufacturing recommendations and added at 10 nM to the amine-coupled microspheres described above. This was subsequently vortexed and incubated for 2 hours at room temperature with rotation. Wash microspheres 3 times with PBS 2% BSA. Microspheres were resuspended in 1 mL of PBS at 9 million microspheres / mL.

[0180] Protein-modified and protein-unmodified (intact) microspheres were mixed at 2700 beads / ml, and 75 µL of microspheres were plated per well on a 96 well ProcartaPlex 96 well flat bottom plate (ThermoFisher, Cat. No: EPX-44444-000) and mixed with 25 µL of individual anti-SARS-CoV-2-S supernatant-containing antibody. Samples and microspheres were incubated for two hours at 25 °C and then washed twice with 200 µL of DPBS with 0.05% Tween 20. To detect bound antibody levels to individual microspheres, 100 µL of 2.5 µg / mL R-Phycoerythrin conjugated goat F(ab')2 anti-human kappa (Southern Biotech, Cat# 2063-09) in blocking buffer (for antibodies with hFc), or 100 µL of 1.25 µg / mL R-Phycoerythrin AffiniPure F(ab') 2 Fragment Goat Anti-Mouse IgG, F(ab') 2 Fragment Specific (Jackson Immunoresearch, Cat. No: 115-116-072) in blocking buffer (for antibodies with mFc), or 100 µL of 1.25 µg / mL R-Phycoerythrin Anti-His (Biolegend, Cat. No: 362603) in blocking buffer (for ACE-2 control, R&D, Cat. No. 933-ZN), was added and incubated for 30 minutes at 25 °C. After 30 minutes, the samples were washed twice with 200 µl of washing buffer and resuspended in 150 µL of wash buffer. The plates were read in FlexMap 3D® (Luminex Corp.) and Luminex xPonent® software version 4.3 (Luminex Corp.).

[0181] The results of the Luminex binning results are shown in Table 10 as median fluorescence intensity (MFI) signal intensities. To determine clusters, data was normalized to the intact protein (unmodified microspheres) and clustered. The 46 anti-SARS-CoV-2 antibodies were classified in 9 clusters with 2 or more antibodies, and 11 antibodies were classified as single nodes. Clusters were assigned by based on these results of the hierarchical clustering and dendrogram. These results show that the 46 anti-SARS-CoV-2-S antibody supernatants had diverse binding characteristics and profiles, suggesting that the collection of antibodies bound to different epitopes on the SARS-CoV-2 spike protein. Example 9: Biacore binding kinetics of anti-SARS-CoV-2-S monoclonal antibodies

[0182] Equilibrium dissociation constants (K D ) for different SARS-CoV-2-S antibodies from primary supernatants from CHOt cells or from hybridomas were determined using a real-time surface plasmon resonance-based Biacore T200 / Biacore 8K biosensor. All binding studies were performed in 10mM HEPES, 150mM NaCl, 3mM EDTA, and 0.05% v / v Surfactant Tween-20, pH 7.4 (HBS-ET) running buffer at 25 °C. The Biacore CM5 sensor chip surface was first derivatized by amine coupling with either mouse anti-human Fc specific mAb or rabbit anti-mouse Fcγ monoclonal antibody (GE, Catalog # BR-1008-38) to capture anti-SARS-CoV-2 antibodies. Binding studies were performed on a human SARS-CoV-2 RBD extracellular domain expressed with a C-terminal myc-myc-hexahistidine tag (SARS-COV-2 RBD-MMH), SARS-CoV-2 RBD extracellular domain expressed with a C-terminal mouse IgG2a (SARS-COV-2 RBD-mFc), or SARS-CoV-2 RBD extracellular domain expressed with a C-terminal human IgG1 (SARS-COV-2 RBD-hFc). Single concentrations of SARS-COV-2 RBD-MMH, (100nM); SARS-COV-2 RBD-mFc (50nM), or SARS-COV-2 RBD-hFc (50nM), prepared in HBS-ET running buffer, were injected for 1.5 minutes at a flow rate of 30µL / min while the dissociation of antibody-bound different SARS-CoV-2 RBD reagents was monitored for 2 minutes in HBS-ET running buffer. At the end of each cycle, the SARS-CoV-2 RBD antibody capture surface was regenerated using either a 10 sec injection of 20mM phosphoric acid for the mouse anti-human Fc specific monoclonal antibody surface or a 40 sec injection of 10mM Glycine, HCl, pH1.5 for the rabbit anti-mouse Fcγ specific polyclonal antibody. The association rate (k a ) and dissociation rate (k d ) were determined by fitting the real-time binding sensorgrams to a 1:1 binding model with mass transport limitation using BiaEvaluation software v3.1 or Biacore Insight Evaluation software v2.0. or curve-fitting software. Binding dissociation equilibrium constant (K D ) and dissociative half-life (t½) were calculated from the kinetic rates as: K D M = kd ka , and t½ min = ln 2 60 ∗ kd

[0183] Binding kinetics parameters for different SARS-CoV-2 monoclonal antibodies binding to different anti-SARS-COV-2 RBD reagents of the invention at 25°C are shown in Tables 11 and 12. Table 11: Binding kinetics of SARS-COV-2 RBD-MMH binding to anti-SARS-CoV-2 monoclonal antibodies at 25 °C Supernatant mAb Capture Level (RU) 50nM Ag Bound (RU) k a k d KD t½ (1 / Ms) (1 / s) (M) (min) mAb1091320103814.91E+052.28E-024.64E-080.5mAb1091431691743.49E+051.36E-023.89E-080.8mAb109158241098.85E+043.18E-043.59E-0936.3mAb1093222613268.50E+041.26E-041.48E-0992mAb1093314144281.05E+064.08E-033.88E-092.8mAb1093429189811.01E+064.35E-034.32E-092.7mAb1093532936942.11E+053.99E-031.89E-082.9mAb1093624917173.03E+058.81E-042.91E-0913.1mAb1093718465043.81E+055.73E-031.50E-082mAb 1092012952346.22E+052.20E-023.54E-080.5mAb1092110241419.52E+044.99E-045.24E-0923.1mAb 1092223957863.91E+052.00E-035.11E-095.8mAb 1092312783222.94E+056.04E-032.06E-081.9mAb 109247661661.97E+053.65E-031.85E-083.2mAb1093031373288.90E+041.85E-032.08E-086.2mAb1093821671806.60E+043.48E-045.28E-0933.2mAb1093915052411.69E+053.38E-032.00E-083.4mAb 1094021496983.34E+052.38E-037.15E-094.9mAb1094118112889.85E+045.17E-045.25E-0922.3mAb 1098210961881.32E+052.71E-032.06E-084.3mAb 1098416543871.55E+053.70E-042.39E-0931.2mAb1098519747499.41E+051.45E-031.54E-098mAb 1098615605243.21E+052.56E-047.97E-1045.2mAb1098712423564.50E+051.04E-022.32E-081.1mAb1098812272911.27E+063.52E-022.77E-080.3mAb109896922571.60E+063.14E-031.96E-093.7mAb 1096922004271.80E+054.71E-032.61E-082.5mAb1097018654381.37E+057.99E-045.82E-0914.4mAb1097114823581.68E+054.49E-042.67E-0925.8mAb 1096412084601.06E+067.56E-047.14E-1015.3mAb 1096510461681.19E+052.73E-032.28E-084.2mAb 1096614223431.57E+054.40E-042.81E-0926.3mAb1096714211751.12E+051.08E-049.66E-10106.9mAb1095411503382.34E+054.05E-041.73E-0928.5mAb1095510321991.38E+052.69E-031.95E-084.3mAb1095613031842.02E+055.31E-032.62E-082.2mAb109577361631.34E+053.15E-042.35E-0936.7mAb 10977221572.33E+057.17E-043.08E-0916.1mAb1101010271083.35E+051.48E-034.42E-097.8mAb1100411111611.88E+053.12E-031.66E-083.7mAb11000381161.40E+052.41E-021.72E-070.5mAb110061118498.97E+043.67E-044.10E-0931.5mAb11008887566.73E+044.00E-035.94E-082.9mAb 109981155691.95E+052.28E-021.17E-070.5mAb 10996616281.53E+051.10E-027.18E-081.1mAb11002107083.21E+052.54E-027.93E-080.5 Table 12: Binding kinetics of SARS-COV-2 RBD-mFc or SARS-COV-2 RBD-hFc binding to anti-SARS-CoV-2-S monoclonal antibodies at 25 °C Supernatant mAb Capture Level (RU) 50nM Ag Bound (RU) k a k d KD t½ (1 / Ms) (1 / s) (M) (min) mAb109139615756.23E+051.52E-042.44E-1076.1mAb1091414673131.83E+051.00E-05*5.47E-111155*mAb109153921412.81E+051.00E-05*3.56E-111155*mAb1093210603722.42E+051.00E-05*4.13E-111155*mAb109336814651.23E+062.12E-041.73E-1054.4mAb1093414019491.41E+061.17E-048.32E-1198.3mAb1093516678303.83E+051.00E-05*2.61E-111155*mAb1093611716996.52E+051.00E-05*1.53E-111155*mAb109379045756.39E+057.28E-051.14E-10158.7mAb 109206173577.02E+052.92E-044.16E-1039.5mAb109214891702.66E+051.00E-05*3.75E-111155*mAb 1092212868287.19E+052.42E-053.36E-11478.2mAb 109236133626.51E+052.83E-054.35E-11407.7mAb 109244652233.67E+058.13E-052.22E-10142.1mAb1093021564492.32E+051.00E-05*4.31E-111155*mAb1093813633333.11E+051.00E-05*3.22E-111155*mAb109399043242.99E+051.15E-053.87E-111004.3mAb 1094015088935.61E+052.86E-055.09E-11403.8mAb1094111323712.60E+051.00E-05*2.15E-111155*mAb 109825292363.10E+051.69E-055.44E-11683.6mAb 1098412135734.02E+051.00E-05*2.49E-111155*mAb10985146310401.09E+061.27E-051.17E-11910.9mAb 1098611687526.33E+051.00E-05*1.58E-111155*mAb 109879026328.20E+051.70E-042.08E-1067.8mAb109888926281.24E+063.46E-042.79E-1033.4mAb109895053782.07E+069.30E-054.50E-11124.2mAb 1096916587383.05E+051.51E-054.96E-11764mAb1097013706613.48E+051.00E-05*2.88E-111155*mAb1097110815563.95E+051.00E-05*2.53E-111155*mAb 109648756511.43E+061.00E-05*7.00E-121155*mAb 109657623222.97E+051.00E-05*3.36E-111155*mAb 109669214304.02E+051.00E-05*2.49E-111155*mAb109679453553.99E+051.00E-05*2.51E-111155*mAb109547344145.77E+051.00E-05*1.73E-111155*mAb109556342923.96E+052.34E-055.92E-11493.6mAb109568423393.74E+051.48E-043.95E-1078mAb109574492093.58E+051.00E-05*2.79E-111155*mAb 109771611025.56E+051.04E-041.87E-10110.9mAb1101010141634.24E+051.00E-05*2.36E-111155*mAb1100411012413.46E+056.63E-051.91E-10174.2mAb11000380614.38E+051.83E-034.17E-096.3mAb110061112751.88E+051.00E-05*5.32E-111155*mAb110088721101.61E+051.15E-047.15E-10100.4mAb1099811402273.30E+055.21E-041.58E-0922.2mAb10996629832.88E+059.32E-043.24E-0912.4mAb110021068602.69E+054.49E-031.67E-082.6 *: Estimated value based on the limit of measurement of the dissociative rate constant and dissociative half-life under the experimental conditions. Example 10: Characterization of anti-SARS-CoV-2-S monoclonal antibodies by blocking ELISA

[0184] An ELISA-based blocking assay was developed to determine the ability of anti-SARS-CoV2-S antibodies to block the binding of the SARS-CoV-2 spike protein receptor binding domain (RBD) to human angiotensin converting enzyme 2 (hACE2).

[0185] The SARS-CoV-2 protein used in the experiments was comprised of the receptor binding domain (RBD) portion of the SARS-CoV-2 spike protein (amino acids Arg319 to Phe541) expressed with the Fc portion of the human IgG1 at the c-terminus (SARS-CoV-2 RBD-hFc; see NCBI accession number MN908947.3) The human ACE2 protein used in the experiments was purchased from R&D systems and is comprised of amino acids glutamine 18 to serine 740 with a c-terminal 10X-Histidine tag (hACE2-His; NCBI accession number Q9BYF1).

[0186] Experiments were carried out using the following procedure. A monoclonal anti-Penta-His antibody (Qiagen) was coated at 1 µg / ml in PBS on a 96-well microtiter plate overnight at 4 °C. The hACE2-His receptor was added at 0.2 µg / ml in PBS and bound for 2 hours at room temperature. Nonspecific binding sites were subsequently blocked using a 0.5% (w / v) solution of BSA in PBS. In other microtiter plates, a constant amount of 10 pM or 15 pM (as indicated in Table 13) of SARS-CoV-2 RBD-hFc protein was bound with antibodies diluted 1:10 or 1:20 in PBS + 0.5% BSA. These antibody-protein complexes, after a one-hour incubation, were transferred to the microtiter plate coated with hACE2-His. After 1.5 hours of incubation at RT, the wells were washed, and plate-bound SARS-CoV-2 RBD-hFc protein was detected with goat-anti-human IgG antibody conjugated with horseradish peroxidase (HRP) (Jackson). The plates were then developed using TMB substrate solution (BD Biosciences, catalog #555214) according to manufacturer's recommendation and absorbance at 450nm was measured on a Victor X5 plate reader.

[0187] Data analysis was performed by calculating the % reduction of signal of the fixed SARS-CoV-2-S RBD-hFc concentration in the presence of the antibody vs in the absence of the antibody. In the calculation, binding signal of the sample of the constant SARS-CoV-2-S RBD-hFc without the presence of the antibody for each plate was referenced as 100% binding or 0% blocking; and the baseline signal of the sample of media only without the presence of SARS-CoV-2 RBD-hFc was referenced as 0% binding or 100% blocking.

[0188] The ability of anti-SARS-CoV-2-S antibodies to block SARS-CoV-2-S RBD from binding to human ACE2 was assessed using a blocking ELISA format. Single point test antibody supernatant blocking of either 10 pM or 15 pM SARS-CoV-2-S RBD-hFc binding to hACE2-His, which was presented on anti-His antibody coated on 96-well microtiter plates, was detected with an HRP conjugated anti-hFc antibody.

[0189] The blocking results of three assays are summarized in Table 13. The SARS-CoV-2-S binding signal (450 nm) and theG calculated % blocking are indicated. A range of blocking is observed for the test samples. For samples where an NA is indicated in columns 6 and 7, a plate-corrected value is included in columns 4 and 5, as data was consistent with a single plate switch occurring for those samples. 43 of 46 antibody supernatants blocked greater than 50% of the SARS-CoV-2-S RBD-hFc binding to plate-coated human ACE2, with 16 of them blocking >90% of the signal. Table 13: Blocking ELISA Results Supernatant SARS-CoV-2 RBD Fixed Concentration Supernatant dilution Plate corrected SRAS-CoV-2 RBD-hFc Binding to His presented ACE2 (Abs 450nm) Plate corrected SARS-CoV-2 RBD-hFc Binding to His presented ACE2 % Blocking SARS-CoV-2 RBD-hFc Binding to His presented ACE2 (Abs 450nm) SARS-CoV-2 RBD-hFc Binding to His presented ACE2 % Blocking mAb1091315pM1:100.20680.50.20680.5mAb1091415pM1:100.32659.10.32659.1mAb1091515pM1:100.17189.70.17189.7mAb1093215pM1:100.25457.30.25457.3mAb1093315pM1:100.15896.30.15896.3mAb1093415pM1:100.209780.20978mAb1093515pM1:100.23869.40.23869.4mAb1093615pM1:100.23470.60.23470.6mAb1093715pM1:100.17688.10.17688.1mAb 1092015pM1:100.601-56.50.601-56.5mAb1092115pM1:100.19282.70.19282.7mAb 1092215pM1:100.18186.40.18186.4mAb 1092315pM1:100.23743.60.23743.6mAb 1092415pM1:100.17578.20.17578.2mAb1093015pM1:100.24142.50.24142.5mAb1093815pM1:100.16987.50.16987.5mAb1093915pM1:100.20465.60.20465.6mAb 1094015pM1:100.15295.20.15295.2mAb1094115pM1:100.17497.20.17497.2mAb 1098215pM1:100.19583.50.19583.5mAb 1098415pM1:100.16696.3NANAmAb1098515pM1:100.16297NANAmAb 1098615pM1:100.15897.8NANAmAb1098715pM1:100.24381.8NANAmAb1098815pM1:100.244840.24484mAb1098915pM1:100.155101.80.155101.8mAb 1096915pM1:100.22187.80.22187.8mAb1097015pM1:100.16497.70.16497.7mAb1097115pM1:100.1796.70.1796.7mAb 1096415pM1:100.16996.90.16996.9mAb 1096515pM1:100.15898.80.15898.8mAb 1096615pM1:100.15794.20.15794.2mAb1096715pM1:100.14597.90.14597.9mAb1095415pM1:100.14797.30.14797.3mAb1095515pM1:100.16292.70.16292.7mAb1095615pM1:100.18984.50.18984.5mAb1095715pM1:100.15495.10.15495.1mAb 1097715pM1:100.31571.50.31571.5mAb1101010pM1:200.18682.10.18682.1mAb1100410pM1:200.211700.21170mAb1100010pM1:200.17372.70.17372.7mAb1100610pM1:200.236580.23658mAb1100810pM1:200.21369.10.21369.1mAb1099810pM1:200.18561.60.18561.6mAb1099610pM1:200.295-18.10.295-18.1mAb1100210pM1:200.17779.20.17779.2 Example 11: Epitope mapping of anti-SARS-CoV-2-S monoclonal antibodies to spike glycoprotein by Hydrogen-Deuterium Exchange Mass Spectrometry

[0190] Hydrogen-Deuterium Exchange Mass Spectrometry (HDX-MS) was performed to determine the amino acid residues of the SARS-CoV-2 Spike Protein Receptor Binding Domain (RBD (amino acids R319-F541)) that interact with mAb10989, mAb10987, mAb10934, mAb10933, mAb10920, mAb10922, mAb10936, mAb10954, mAb10964, mAb10977, mAb10984, and mAb10986. A general description of the HDX-MS method is set forth in e.g., Ehring (1999) Analytical Biochemistry 267(2):252-259; and Engen and Smith (2001) Anal. Chem. 73:256A-265A.

[0191] The HDX-MS experiments were performed on an integrated HDX-MS platform, consisting of a Leaptec HDX PAL system for the deuterium labeling and quenching, a Waters Acquity I-Class (Binary Solvent Manager) for the sample digestion and loading, a Waters Acquity I-Class (Binary Solvent Manager) for the analytical gradient, and a Thermo Q Exactive HF mass spectrometer for peptide mass measurement.

[0192] The labeling solution was prepared as PBS buffer in D 2 O at pD 7.0 (10 mM phosphate buffer, 140 mM NaCl, and 3 mM KCl, equivalent to pH 7.4 at 25°C). For deuterium labeling, 10 µL of the RBD protein or RBD protein premixed with each one of the 12 antibodies listed above were incubated at 20 °C with 90 µL of D 2 O labeling solution for various timepoints, in duplicate. For mAb10989, mAb10987, mAb10934, and mAb10933, the time points were 0 min (non-deuterated control), 5 min, and 10 min. For mAb10920, mAb10922, mAb10936, mAb 10954, mAb 10964, mAb 10977, mAb 10984, and mAb 10986, the time points were 0 min (non-deuterated control) and 10 min. The deuteration reaction was quenched by adding 90 µL of pre-chilled quench buffer (0.5 M TCEP-HCl, 4 M urea and 0.5% formic acid) to each sample for a 90 second incubation at 20 °C. The quenched samples were then injected into the Leaptec HDX PAL system for online pepsin / protease XIII digestion. The digested peptides were trapped by a C18 column (2.1 mm x 5 mm, Waters) and separated by another C18 column (2.1 mm x 50 mm, Waters) at -5 °C with a 20 minute gradient (for mAb 10989, mAb 10987, mAb 10934, and mAb10933) or a 10 minute gradient (for mAb10920, mAb10922, mAb10936, mAb10954, mAb 10956, mAb 10964, mAb 10977, and mAb 10984) from 0% to 90% of movbile phase B solution (mobile phase A solution: 0.5% formic acid and 4.5% acetonitrile in water, mobile phase B solution: 0.5% formic acid in acetonitrile). The eluted peptides were analyzed by a Thermo Q Exactive HF mass spectrometry in LC-MS / MS or LC-MS mode.

[0193] The LC-MS / MS data from the undeuterated RBD protein sample were searched against a database including amino acid sequences of the RBD protein, pepsin, protease XIII, and their reversed sequences using Byonic search engine (Protein Metrics). The search parameters were set as default using non-specific enzymatic digestion and human glycosylation as common variable modification. The list of identified peptides was then imported into HDExaminer software (version 3.1) to calculate the deuterium uptake (D-uptake) and differences in deuterium uptake percentage (Δ%D) for all deuterated samples. Difference in deuterium uptake percentage (Δ%D) was calculated as follows. Difference in deuterium uptake ΔD = D − uptake RBD − mAb − D − uptake RBD alone Difference in deuterium uptake percentage Δ % D = ΔD Theoretical maximum D uptake of the peptide × 100

[0194] A total of 190 peptides from the RBD were identified from both RBD alone and RBD in complex with mAb10989 samples, representing 86.06% sequence coverage of the RBD. Any peptide that exhibited a reduction in deuterium uptake of 5% or greater (i.e., a Δ%D value of less than -5%, such as -6%, -10%, and so on) upon mAb binding was defined as significantly protected. Peptides corresponding to amino acids 467-513 (DISTEIYQAGSTPCNGVEGFNCYFPLQSYGFQPTNGVGYQPYRVVVL) (SEQ ID NO: 835) of the RBD were significantly protected by mAb10989.

[0195] A total of 187 peptides from the RBD were identified from both RBD alone and RBD in complex with mAb10987 samples, representing 86.06% sequence coverage of RBD. Any peptide that exhibited a reduction in deuterium uptake of 5% or greater (i.e., a Δ%D value of less than -5%, such as -6%, -10%, and so on) upon mAb binding was defined as significantly protected. Peptides corresponding to amino acids 432-452 (CVIAWNSNNLDSKVGGNYNYL) (SEQ ID NO: 836) of the RBD were significantly protected by mAb10987.

[0196] A total of 188 peptides from the RBD were identified from both RBD alone and RBD in complex with mAb10934 samples, representing 86.06% sequence coverage of the RBD. Any peptide that exhibited a reduction in deuterium uptake of 5% or greater (i.e., a Δ%D value of less than -5%, such as -6%, -10%, and so on) upon mAb binding was defined as significantly protected. Peptides corresponding to amino acids 432-452 (CVIAWNSNNLDSKVGGNYNYL) (SEQ ID NO: 836), 467-474 (DISTEIYQ) (SEQ ID NO: 837), and 480-513 (CNGVEGFNCYFPLQSYGFQPTNGVGYQPYRVVVL) (SEQ ID NO: 838) of the RBD were significantly protected by mAb 10934.

[0197] A total of 188 peptides from the RBD were identified from both RBD alone and RBD in complex with mAb10933 samples, representing 86.06% sequence coverage of the RBD. Any peptide that exhibited a reduction in deuterium uptake of 5% or greater (i.e., a Δ%D value of less than -5%, such as -6%, -10%, and so on) upon mAb binding was defined as significantly protected. Peptides corresponding to amino acids 467-510 (DISTEIYQAGSTPCNGVEGFNCYFPLQSYGFQPTNGVGYQPYRV) (SEQ ID NO: 839) of the RBD were significantly protected by mAb 10933.

[0198] A total of 75 peptides from the RBD were identified from both RBD alone and RBD in complex with mAb 10920 samples, representing 83.27% sequence coverage of the RBD. Any peptide that exhibited a reduction in deuterium uptake of 5% or greater (i.e., a Δ%D value of less than -5%, such as -6%, -10%, and so on) upon mAb binding was defined as significantly protected. Peptides corresponding to amino acids 471-486 (EIYQAGSTPCNGVEGF) (SEQ ID NO: 840), and 491-515 (PLQSYGFQPTNGVGYQPYRVVVLSF) (SEQ ID NO: 841) of the RBD were significantly protected by mAb 10920.

[0199] A total of 86 peptides from the RBD were identified from both RBD alone and RBD in complex with mAb 10922 samples, representing 87.25% sequence coverage of the RBD. Any peptide that exhibited a reduction in deuterium uptake of 5% or greater (i.e., a Δ%D value of less than -5%, such as -6%, -10%, and so on) upon mAb binding was defined as significantly protected. Peptides corresponding to amino acids 432-452 (CVIAWNSNNLDSKVGGNYNYL) (SEQ ID NO: 836) of the RBD were significantly protected by mAb10922.

[0200] A total of 81 peptides from the RBD were identified from both RBD alone and RBD in complex with mAb10936 samples, representing 82.07% sequence coverage of the RBD. Any peptide that exhibited a reduction in deuterium uptake of 5% or greater (i.e., a Δ%D value of less than -5%, such as -6%, -10%, and so on) upon mAb binding was defined as significantly protected. Peptides corresponding to amino acids 351-360 (YAWNRKRISN) (SEQ ID NO: 842), 432-452 (CVIAWNSNNLDSKVGGNYNYL) (SEQ ID NO: 836), 467-486 (DISTEIYQAGSTPCNGVEGF) (SEQ ID NO: 843), and 491-513 (PLQSYGFQPTNGVGYQPYRVVVL) (SEQ ID NO: 844) of the RBD were significantly protected by mAb10936.

[0201] A total of 84 peptides from the RBD were identified from both RBD alone and RBD in complex with mAb 10954 samples, representing 87.25% sequence coverage of the RBD. Any peptide that exhibited a reduction in deuterium uptake of 5% or greater (i.e., a Δ%D value of less than -5%, such as -6%, -10%, and so on) upon mAb binding was defined as significantly protected. Peptides corresponding to amino acids 400-422 (FVIRGDEVRQIAPGQTGKIADYN) (SEQ ID NO: 845), 453-486 (YRLFRKSNLKPFERDISTEIYQAGSTPCNGVEGF) (SEQ ID NO: 846), and 490-515 (FPLQSYGFQPTNGVGYQPYRVVVLSF) (SEQ ID NO: 847) of the RBD were significantly protected by mAb 10954.

[0202] A total of 109 peptides from the RBD were identified from both RBD alone and RBD in complex with mAb10964 samples, representing 83.67% sequence coverage of RBD. Any peptide that exhibited a reduction in deuterium uptake of 5% or greater (i.e., a Δ%D value of less than -5%, such as -6%, -10%, and so on) upon mAb binding was defined as significantly protected. Peptides corresponding to amino acids 401-424 (VIRGDEVRQIAPGQTGKIADYNYK) (SEQ ID NO: 848), and 471-513 (EIYQAGSTPCNGVEGFNCYFPLQSYGFQPTNGVGYQPYRVVVL) (SEQ ID NO: 849) of the RBD were significantly protected by mAb 10964.

[0203] A total of 78 peptides from the RBD were identified from both RBD alone and RBD in complex with mAb 10977 samples, representing 87.25% sequence coverage of the RBD. Any peptide that exhibited a reduction in deuterium uptake of 5% or greater (i.e., a Δ%D value of less than -5%, such as -6%, -10%, and so on) upon mAb binding was defined as significantly protected. Peptides corresponding to amino acids 351-364 (YAWNRKRISNCVAD) (SEQ ID NO: 850), and 471-486 (EIYQAGSTPCNGVEGF) (SEQ ID NO: 840) of the RBD were significantly protected by mAb 10977.

[0204] A total of 88 peptides from the RBD were identified from both RBD alone and RBD in complex with mAb 10984 samples, representing 87.25% sequence coverage of RBD. Any peptide that exhibited a reduction in deuterium uptake of 5% or greater (i.e., a Δ%D value of less than -5%, such as -6%, -10%, and so on) upon mAb binding was defined as significantly protected. Peptides corresponding to amino acids 400-422 (FVIRGDEVRQIAPGQTGKIADYN) (SEQ ID NO: 845), and 453-486 (YRLFRKSNLKPFERDISTEIYQAGSTPCNGVEGF) (SEQ ID NO: 846) of the RBD were significantly protected by mAb10984.

[0205] A total of 84 peptides from the RBD were identified from both RBD alone and RBD in complex with mAb 10986 samples, representing 87.25% sequence coverage of the RBD. Any peptide that exhibited a reduction in deuterium uptake of 5% or greater (i.e., a Δ%D value of less than -5%, such as -6%, -10%, and so on) upon mAb binding was defined as significantly protected. Peptides corresponding to amino acids 400-422 (FVIRGDEVRQIAPGQTGKIADYN) (SEQ ID NO: 845), 453-486 (YRLFRKSNLKPFERDISTEIYQAGSTPCNGVEGF) (SEQ ID NO: 846), and 490-515 (FPLQSYGFQPTNGVGYQPYRVVVLSF) (SEQ ID NO: 847) of the RBD were significantly protected by mAb 10986.

[0206] In sum, the majority of the neutralizing antibodies tested contact the RBD in a manner that overlaps the RBD residues that comprise the ACE2 interface; furthermore, the antibodies can be grouped based on their pattern of contacting the RBD surface, as shown in Figure 15. The above data are also summarized in Tables 14-25. Table 14: Spike protein receptor binding domain (RBD) peptides with significant protection upon formation of RBD-mAb compared to RBD alone RBD Residues5 min incubation10 min incubationRBD-mAb10989RBDRBD-mAb10989RBDD-uptakeD-uptakeΔDD-uptakeD-uptakeΔDΔ%D467-4742.673.16-0.492.533.17-0.64-10.5 470-4730.480.98-0.500.470.98-0.51-28.0 470-4740.991.46-0.470.991.44-0.45-16.9 471-4740.510.89-0.380.510.89-0.38-20.9 475-4862.202.93-0.732.112.94-0.83-9.7 475-4873.314.50-1.193.614.48-0.87-11.4 475-4892.774.48-1.712.784.53-1.75-16.0 475-4902.634.96-2.332.674.97-2.30-19.8 480-4891.823.67-1.851.773.69-1.92-26.2 483-4860.310.78-0.470.300.78-0.48-26.5 487-4890.050.40-0.350.020.39-0.37-40.4 487-4900.110.90-0.790.110.84-0.73-42.3 487-4910.101.05-0.950.101.03-0.93-52.0 487-4950.621.59-0.970.671.57-0.90-17.4 487-5095.636.99-1.365.687.02-1.34-8.3 487-5106.087.37-1.296.087.44-1.36-7.7 487-5125.726.48-0.765.606.77-1.17-5.1 487-5135.156.16-1.015.076.14-1.07-5.3 488-4900.030.22-0.190.000.23-0.23-23.2 488-4910.040.37-0.330.040.36-0.32-36.3 Table 15: Spike protein RBD peptides with significant protection upon formation of RBD-mAb10987 complex comparing to RBD alone RBD5 min incubation10 min incubationRBD-mAb 10987RBDRBD-mAb 10987RBDResiduesD-uptakeD-uptakeΔDD-uptakeD-uptakeΔDΔ%D432-4411.622.17-0.551.642.18-0.54-7.6 432-4495.606.59-0.995.546.59-1.05-7.1 432-4526.207.49-1.296.207.46-1.26-7.5 433-4411.502.00-0.501.492.01-0.52-8.1 440-4523.954.81-0.864.034.80-0.77-8.3 442-4492.492.98-0.492.602.99-0.39-8.2 Table 16: RBD peptides with significant protection upon formation of RBD-mAb10934 complex comparing to RBD alone RBD5 min incubation10 min incubationRBD-niAb10934RBDRBD-mAb10934RBDResiduesD-uptakeD-uptakeΔDD-uptakeD-uptakeΔDΔ%D432-4525.707.49-1.795.627.46-1.84-10.6 433-4411.602.00-0.401.632.01-0.38-6.2 434-4412.242.42-0.182.132.52-0.39-5.3 440-4523.124.81-1.693.104.80-1.70-17.1 442-4492.372.98-0.612.372.99-0.62-11.4 442-4522.674.21-1.542.664.23-1.57-19.1 443-4522.533.78-1.252.523.78-1.26-17.5 444-4511.792.73-0.941.802.73-0.93-17.2 444-4521.823.09-1.271.753.09-1.34-20.7 445-4521.242.42-1.181.242.43-1.19-22.0 467-4742.643.16-0.522.583.17-0.59-10.2 470-4730.510.98-0.470.550.98-0.43-25.0 470-4741.031.46-0.431.011.44-0.43-16.0 471-4740.560.89-0.330.550.89-0.34-18.6 480-4893.193.67-0.483.193.69-0.50-6.8 487-4890.040.40-0.360.060.39-0.33-38.6 487-4900.540.90-0.360.530.84-0.31-18.8 487-4910.631.05-0.420.701.03-0.33-20.5 487-4950.731.59-0.860.711.57-0.86-16.0 487-5095.556.99-1.445.577.02-1.45-8.9 487-5105.897.37-1.486.007.44-1.44-8.5 487-5134.376.16-1.794.796.14-1.35-7.9 488-5094.505.49-0.994.605.52-0.92-6.2 488-5105.846.58-0.745.656.67-1.02-5.4 490-5095.166.01-0.855.306.12-0.82-5.8 490-5125.156.37-1.225.306.28-0.98-6.4 490-5134.906.10-1.205.056.05-1.00-6.1 503-5091.191.39-0.201.211.41-0.20-5.5 Table 17: RBD peptides with significant protection upon formation of RBD-mAb10933 complex comparing to RBD alone RBD5 min incubation10 min incubationRBD-mAb10933RBDRBD-mAb10933RBDResiduesD-uptakeD-uptakeΔDD-uptakeD-uptakeΔDΔ%D467-4742.523.16-0.642.553.17-0.62-11.7 470-4741.031.46-0.431.031.44-0.41-15.6 471-4740.540.89-0.350.540.89-0.35-19.5 475-4873.624.50-0.883.634.48-0.85-9.6 475-4893.214.48-1.273.264.53-1.27-11.8 480-4861.792.06-0.271.872.07-0.20-5.1 480-4892.133.67-1.542.183.69-1.51-21.2 483-4860.610.78-0.170.620.78-0.16-9.3 487-4890.020.40-0.380.020.39-0.37-41.6 487-4900.420.90-0.480.400.84-0.44-25.6 487-4910.461.05-0.590.461.03-0.57-32.0 487-4950.741.59-0.850.821.57-0.75-14.8 487-5096.016.99-0.986.147.02-0.88-5.7 487-5106.297.37-1.086.147.44-1.30-7.0 488-4900.190.22-0.030.130.23-0.10-7.4488-4910.260.37-0.110.250.36-0.11-12.3 Table 18: RBD peptides with significant protection upon formation of RBD-mAb10920 complex comparing to RBD alone RBD10 min incubationRBD-mAb 10920RBDResiduesD-uptakeD-uptakeΔDΔ%D471-4864.635.40-0.77-6.6 475-4862.743.27-0.53-6.5 491-5135.456.57-1.12-6.6 495-5104.515.43-0.92-8.5 495-5134.415.13-0.72-5.4 496-5153.584.35-0.77-5.4 Table 19: RBD peptides with significant protection upon formation of RBD-mAb10922 complex comparing to RBD alone RBD10 min incubationRBD-mAb 10922RBDResiduesD-uptakeD-uptakeΔDΔ%D432-4411.862.23-0.37-5.3 442-4523.524.57-1.05-13.0 Table 20: RBD peptides with significant protection upon formation of RBD-mAb10936 complex comparing to RBD alone RBD10 min incubationRBD-mAb10936RBDResiduesD-uptakeD-uptakeΔDΔ%D351-3602.683.10-0.42-5.9 432-4411.852.23-0.38-5.3 442-4522.554.57-2.02-25.0 443-4522.984.01-1.03-14.2 467-4700.690.84-0.15-8.1 471-4864.735.40-0.67-5.8 491-5135.486.57-1.09-6.4 495-5104.385.43-1.05-9.8 Table 21: RBD peptides with significant protection upon formation of RBD-mAb10954 complex comparing to RBD alone RBD10 min incubationRBD-mAb10954RBDResiduesD-uptakeD-uptakeΔDΔ%D400-4203.674.56-0.89-5.5 401-4203.394.22-0.83-5.5 401-4213.444.28-0.84-5.2 406-4203.324.10-0.78-7.2 406-4213.234.11-0.88-7.6 406-4223.414.16-0.75-5.9 407-4202.863.62-0.76-7.7 407-4222.973.74-0.77-6.6 453-4661.532.23-0.70-7.1 453-4703.634.53-0.90-6.7 453-4714.425.22-0.80-5.6 471-4864.345.40-1.06-9.1 472-4864.475.29-0.82-7.6 490-5125.646.65-1.01-5.9 490-5135.616.57-0.96-5.3 491-5135.266.57-1.31-7.7 493-5124.865.69-0.83-5.7 493-5134.745.72-0.98-6.4 495-5104.775.43-0.66-6.2 495-5134.105.13-1.03-7.6 496-5123.604.60-1.00-8.6 496-5153.434.35-0.92-6.4 Table 22: RBD peptides with significant protection upon formation of RBD-mAb10964 complex comparing to RBD alone RBD10 min incubationRBD-mAb 10964RBDResiduesD-uptakeD-uptakeΔDΔ%D401-4213.874.84-0.97-6.0 406-4193.343.91-0.57-5.8 406-4203.474.15-0.68-6.3 406-4213.534.22-0.69-5.9 406-4223.664.37-0.71-5.6 406-4243.314.24-0.93-6.5 410-4223.043.56-0.52-5.8 471-4864.655.41-0.76-6.4 475-4893.344.56-1.22-11.3 480-4892.323.19-0.87-12.1 487-5096.387.58-1.20-7.4 495-5134.505.20-0.70-5.2 496-5124.174.80-0.63-5.4 496-5133.904.85-0.95-7.5 Table23: RBD peptides with significant protection upon formation of RBD-mAb10977 complex comparing to RBD alone RBD10 min incubationRBD-mAb 10977RBDResiduesD-uptakeD-uptakeΔDΔ%D351-3644.825.38-0.56-5.2 471-4863.815.40-1.59-13.6 472-4864.205.29-1.09-10.1 Table24: RBD peptides with significant protection upon formation of RBD-mAb10984 complex comparing to RBD alone RBD10 min incubationRBD-mAb 10984RBDResiduesD-uptakeD-uptakeΔDΔ%D400-4203.734.56-0.83-5.2 401-4213.474.28-0.81-5.1 406-4203.354.10-0.75-7.0 406-4213.314.11-0.80-6.9 406-4223.474.16-0.69-5.5 407-4202.883.62-0.74-7.5 407-4222.943.74-0.80-6.8 453-4661.512.23-0.72-7.3 453-4703.704.53-0.83-6.2 453-4714.495.22-0.73-5.1 471-4864.455.40-0.95-8.1 472-4864.635.29-0.66-6.1 Table 25: RBD peptides with significant protection upon formation of RBD-mAb10986 complex comparing to RBD alone RBD10 min incubationRBD-mAb 10986RBDResiduesD-uptakeD-uptakeΔDΔ%D400-4203.584.56-0.98-6.1 400-4213.604.61-1.01-5.9 401-4203.304.22-0.92-6.1 401-4213.294.28-0.99-6.1 401-4223.444.43-0.99-5.8 406-4203.284.10-0.82-7.6 406-4213.244.11-0.87-7.5 406-4223.354.16-0.81-6.4 407-4202.813.62-0.81-8.2 407-4222.913.74-0.83-7.1 453-4661.532.23-0.70-7.1 453-4703.554.53-0.98-7.3 453-4714.415.22-0.81-5.6 471-4864.135.40-1.27-10.9 490-5105.136.44-1.31-8.6 490-5125.336.65-1.32-7.7 490-5135.256.57-1.32-7.3 491-5134.296.57-2.28-13.3 493-5124.465.69-1.23-8.5 493-5134.625.72-1.10-7.2 495-5133.895.13-1.24-9.3 496-5133.364.53-1.17-9.3 496-5153.054.35-1.30-9.1 Example 12: Neutralization of SARS-CoV-2 wild-type and variant spike proteins

[0207] To test whether anti-SARS-CoV-2 spike protein antibodies can neutralize SARS-CoV-2 variants, these antibodies were screened against a panel of VSV pseudotype viruses expressing wild-type and variant spike proteins. VSV pseudotype viruses were generated by transiently transfecting 293T cells with a plasmid encoding the SARS-CoV-2 spike protein or the same plasmid containing nucleotide variations that encode for known variants of the SARS-CoV-2 spike protein amino acid sequence. All plasmids were confirmed by Sanger sequencing. Cells were seeded in 15 cm plates at 1.2x10 7< cells per plate in DMEM Complete Media (1000 mL DMEM, Gibco; 100 mL FBS, Gibco; 10 mL PSG, Gibco) one day prior to transfection with 15 µg / plate Spike DNA using 125 µL Lipofectamine LTX, 30 µL PLUS reagent, and up to 3 mL Opti-Mem. 24 hours post transfection, the cells were washed with 10 mL PBS, then infected with an MOI of 0.1 VSV ΔG:mNeon< virus in 10 mL of Opti-Mem. Virus was incubated on cells for 1 hour, with gentle rocking every 10 minutes. Cells were washed 3 times with 10 mL PBS, then overlaid with 20 mL Infection media (1000 mL DMEM, Gibco; 10 mL Sodium Pyruvate, Gibco; 7 mL BSA, Sigma; 5 mL Gentamicin, Gibco) before incubation at 37 °C, 5% CO 2 for 24 hours. Pseudovirus supernatant was collected into 250 mL centrifuge tubes on ice, then centrifuged at 3000 rpm for 5 minutes to pellet any cellular debris, aliquoted on ice, then frozen to -80 °C. Infectivity was tested on Vero cells prior to use in neutralization assays. This material will be referred to as VSV ΔG:mNeon< / Spike pseudovirus, or VSV ΔG:mNeon< / Spike_(variant amino acid mutation) (for example, VSV ΔG:mNeon< / Spike_H49Y).

[0208] On Day 1, Vero cells were seeded to 80% confluency in T225 flasks, the cells were washed with PBS (Gibco: 20012-043), TrypLE was added to detach cells from the flask, and Complete DMEM was added to inactivate trypsin. 20,000 Vero cells were plated in in 100 µL of prewarmed Complete DMEM per well in 96 Well Black Polystyrene Microplate (Corning: 3904). On Day 2, VSV ΔG:mNeon< / Spike pseudovirus was thawed on ice and diluted with Infection media. Antibodies were diluted in a U-bottom 96 well plate, generating a dilution of each antibody in 210 µl Infection media at 2X assay concentration. 120 µL of diluted antibodies were transferred to a fresh U-bottom plate, and media and an IgG1 control antibody were added to each plate. 120 µl of diluted pseudovirus was added to every well except the media control wells. To those wells, 120 µL of Infection media was added. Pseudovirus with antibodies were incubated for 30 minutes at room temperature, then media was removed from Vero cells. 100 µL of antibody / pseudovirus mixture were added to the cells, and then incubated at 37 °C, 5% CO 2 for 24 hours. On day 3, supernatant was removed from cell wells and replaced with 100 µL of PBS. Plates were read on a SpectraMax i3 with MiniMax imaging cytometer.

[0209] In addition to testing neutralization capacity with non-replicating VSV-SARS-CoV-2-S virus, antibodies also were tested with SARS-CoV-2 virus. Monoclonal antibodies and antibody combinations were serially diluted in DMEM (Quality Biological), supplemented with 10% (v / v) heat inactivated fetal bovine serum (Sigma), 1% (v / v) penicillin / streptomycin (Gemini Bio-products) and 1% (v / v) L-glutamine (2 mM final concentration, Gibco) (VeroE6 media) to a final volume of 250 µL. Next, 250 µL of VeroE6 media containing SARS-CoV-2 (WA-1) (1000 PFU / mL) was added to each serum dilution and to 250 µL media as an untreated control. The virus-antibody mixtures were incubated for 60 min at 37 °C. Following incubation, virus titers of the mixtures were determined by plaque assay. Finally, 50% plaque reduction neutralization titer (PRNT50) values (the serum dilutions at which plaque formation was reduced by 50% relative to that of the untreated control) were calculated using a 4-parameter logistic curve fit to the percent neutralization data (GraphPad Software, La Jolla, CA).

[0210] Individual monoclonal antibody half maximal inhibitory concentration (IC50) against VSV-SARS-CoV-2 spike protein (S)-expressing pseudovirus encoding the Wuhan-Hu-1 (NCBI Accession Number MN908947.3) sequence of spike protein (S-wt) were determined in Vero cells (Table 26). The majority of antibodies displayed neutralization potency in the picomolar range (pM), with some exhibiting neutralization potency in nanomolar (nM) range.

[0211] While recombinant ACE2 was able to mediate neutralization of the VSV-spike pseudoparticles, as previously reported, its potency was far inferior to that of the monoclonal antibodies, with more than 1000-fold decrease in potency seen relative to the best neutralizing mAbs (Figure 10A). In addition, the potent neutralizing activity of mAb 10987, mAb 10989, mAb10933, and mAb10934 was confirmed in neutralization assays, including neutralization of SARS-CoV-2 in VeroE6 cells (Figure 10B). All neutralization assays generated similar potency across the four mAbs (mAb10987, mAb10989, mAb10933, and mAb10934) and no combinations demonstrated synergistic neutralization activity (Figure 10B). Table 26: mAb neutralization potency (IC50 (M)) against wild-type strain of VSV-SARS-CoV-2-S pseudoparticles in Vero cells Antibody IC50 (M) mAb10934 5.44E-11mAb10936 1.11E-10mAb10987 4.06E-11mAb10924 1.36E-10mAb10935 2.21E-10mAb10913 2.31E-10mAb10939 2.36E-10mAb10937 2.62E-10mAb10920 2.64E-10mAb10941 2.78E-10mAb10923 3.29E-10mAb10915 3.40E-10mAb10932 3.58E-10mAb10921 3.74E-10mAb10914 3.94E-10mAb10940 5.43E-10mAb10989 7.23E-12mAb10938 6.65E-10mAb10922 1.21E-10mAb10930 1.07E-09mAb10954 9.22E-11mAb10955 1.19E-10mAb10933 4.28E-11mAb10956 1.28E-10mAb10957 1.76E-10mAb10964 5.70E-11mAb10965 1.42E-10mAb10966 1.00E-10mAb10967 2.43E-10mAb10970 1.26E-10mAb10971 1.55E-10mAb10977 5.15E-11mAb10982 3.69E-10mAb10984 9.73E-11mAb10985 2.57E-10mAb10986 9.91E-11mAb10988 2.98E-10mAb10969 2.27E-09mAb10996 1.13E-08mAb10998 9.51E-09mAb11002 non-neutralizingmAb11000 2.79E-08mAb11004 6.00E-09mAb11006 1.40E-09mAb11008 2.05E-08mAb11010 non-neutralizing

[0212] Amino acid variants in spike (S) protein were identified from over 7000 publicly available SARS-CoV-2 sequences, representing globally circulating isolates, and cloned into VSV pseudoparticles. Neutralization assays with variant-encoding pseudoparticles were performed to assess the impact of each variant on neutralization potency of the monoclonal antibodies. Table 27 illustrates the relative neutralization potency of monoclonal antibodies against variant encoding pseudoparticles relative to SARS-CoV-2 spike (S-wt) at a single concentration of 5 µg / ml. Percent of neutralization relative to S-wt was captured for each individual antibody and variant. None of the antibodies demonstrated loss of neutralization potency at the 5 µg / ml concentration with the exception of mAb 10985 and the R408I variant. These data demonstrate broad functional neutralization coverage of monoclonal antibodies against globally circulating SARS-CoV-2 spike variants.

[0213] To further interrogate the impact of the S protein variants on neutralization potency of the monoclonal antibodies, full neutralization curves were run to determine the IC50 value of the most potent neutralizing antibodies against a subset of variants localized within the receptor binding domain (RBD) of the S protein. Table 28 shows the IC50 neutralization values for each variant psuedoparticle. Intrinsic variability of up to 3-fold can be observed between pseudoparticle neutralization assays and does not indicate a change in neutralization potency. These data demonstrate that the antibodies retained their neutralization potency against a diverse panel of S protein RBD variants. Example 13: Biacore binding kinetics of purified anti-SARS-CoV-2-S monoclonal antibodies

[0214] Equilibrium dissociation constant (K D ) for different SARS-COV-2 RBD reagents binding to purified CHOt anti-SARS-COV-2 monoclonal antibodies (mAbs) were determined using a real-time surface plasmon resonance based Biacore T200 / Biacore 8K biosensor. All binding studies were performed in 10mM HEPES, 150mM NaCl, 3mM EDTA, and 0.05% v / v Surfactant Tween-20, pH 7.4 (HBS-ET) running buffer at 25 °C and 37 °C. The Biacore CM5 sensor chip surface was first derivatized by amine coupling with either mouse anti-human Fc specific mAb (Regeneron, mAb2567) to capture anti-SARS-COV-2bmAbs. Binding studies were performed on human SARS-COV-2 RBD extracellular domain expressed with a C-terminal myc-myc-hexahistidine (SARS-COV-2 RBD-MMH) and SARS-COV-2 RBD extracellular domain expressed with a C-terminal mouse IgG2a (SARS-COV-2 RBD-mFc). Use of these reagents allowed for the testing of the antibodies' ability to bind monomeric and dimeric RBD peptides, respectively.

[0215] Different concentrations of hSARS-COV-2 RBD-MMH, (90nM-3.33nM, 3-fold dilution) and SARS-COV-2 RBD-mFc (30nM-1.11nM 3-fold dilution) prepared in HBS-ET running buffer, were injected for 3 minutes at a flow rate of 50µL / min while the dissociation of mAb bound different SARS-COV-2 RBD reagents was monitored for 6-10 minutes in HBS-ET running buffer. At the end of each cycle, the SARS-COV-2 RBD mAb capture surface was regenerated using either 12sec injection of 20mM phosphoric acid for mouse anti-human Fc specific mAb surface. The association rate (k a ) and dissociation rate (k d ) were determined by fitting the real-time binding sensorgrams to a 1:1 binding model with mass transport limitation using BiaEvaluation software v3.1 or Biacore Insight Evaluation software v2.0. or curve-fitting software. Binding dissociation equilibrium constant (K D ) and dissociative half-life (t½) were calculated from the kinetic rates as: K D M = kd ka , and t½ min = ln 2 60 ∗ kd

[0216] Binding kinetics parameters for different SARS-COV-2 mAbs binding to different anti-SARS-COV-2 RBD reagents of the invention at 25 °C and 37 °C are shown in Tables 29 through 32, respectively. Table 29: Binding kinetics parameters of SARS-COV-2 RBD-MMH binding to anti-SARS-COV-2-S monoclonal antibodies at 25 °C. mAb Captured (mAb#) mAb Capture Level (RU) 90nM Ag Bound (RU) k a (1 / Ms) k d (1 / s) KD (M) t½ (min) mAb10913287 ± 355.94.04E+052.12E-025.26E-080.5mAb10914310±251.18.81E+043.76E-034.26E-083.1mAb10915310±263.29.61E+041.08E-041.13E-09106.9mAb 10920307 ± 373.94.52E+051.30E-022.87E-080.9mAb10921307 ± 361.41.01E+054.75E-044.71E-0924.3mAb 10922312.2±1.7120.26.14E+051.48E-032.41E-097.8mAb 10923283 ± 280.44.66E+056.17E-031.32E-081.9mAb 10924319 ± 294.62.07E+051.74E-038.40E-096.6mAb10930284.7±0.759.61.24E+053.34E-032.70E-083.5mAb10932315 ± 379.48.99E+041.21E-041.35E-0995.5mAb10933280 ± 199.81.52E+062.78E-031.83E-094.2mAb10934280 ± 1103.44.82E+065.77E-031.20E-092.0mAb10935337 ± 2107.83.93E+054.19E-031.07E-082.8mAb10936311 ± 2107.35.45E+051.07E-031.97E-0910.8mAb10937311±2102.25.72E+054.76E-038.34E-092.4mAb10938338 ± 361.57.27E+041.75E-042.41E-0966.0mAb10939343 ± 282.31.63E+052.84E-031.74E-084.1mAb 10940338 ± 3103.58.01E+052.51E-033.13E-094.6mAb 10941327±192.11.20E+054.12E-043.43E-0928.0mAb 10954286.9±3110.54.04E+053.64E-048.99E-1031.7mAb10955298.3±2.588.81.61E+052.12E-031.32E-085.4mAb10956293.7±0.686.62.22E+054.06E-031.82E-082.8mAb10957286.7±293.01.38E+052.53E-041.84E-0945.7mAb 10964259.6±1.299.91.65E+063.90E-042.36E-1029.6mAb 10965253.1±1.963.61.24E+052.92E-032.35E-084.0mAb 10966266.6±397.42.37E+053.65E-041.54E-0931.6mAb10967260.2±0.970.71.24E+056.28E-055.08E-10183.9mAb 10969272.2±1.387.12.45E+053.80E-031.55E-083.0mAb10970307.3±1.3102.82.27E+051.10E-034.85E-0910.5mAb10971263.1±1.189.32.15E+053.75E-041.74E-0930.8mAb 10977305±398.52.43E+052.57E-041.06E-0944.9mAb 10982267.8±0.569.31.23E+052.06E-031.68E-085.6mAb 10984334±2.1117.92.04E+054.26E-042.09E-0927.1mAb10985306.9±2.1113.41.44E+061.55E-031.08E-097.5mAb 10986268.8±0.9104.34.64E+051.49E-043.21E-1077.5mAb10987270.8±1.378.05.60E+051.20E-022.14E-081.0mAb10988279.2±2.363.68.29E+052.71E-023.27E-080.4mAb10989316.7±1.6114.31.86E+062.78E-031.50E-094.2mAb 10996414.2±2.837.51.41E+052.28E-021.61E-070.5mAb 10998212.3±117.73.54E+051.84E-025.21E-080.6mAb11000322.6±3.573.61.09E+061.14E-031.04E-0910.1mAb11002291.7±2.713.81.65E+056.73E-034.07E-081.7mAb11004232.9±0.676.43.79E+053.24E-038.54E-093.6mAb11006277.2±1.166.99.67E+044.40E-044.55E-0926.3mAb11008214.9±1.540.89.30E+043.27E-033.52E-083.5mAb11010221.8±1.376.81.11E+062.74E-032.47E-094.2mAb1932205±0.85.3NBNBNBNB Table 30: Binding kinetics parameters of SARS-COV-2 RBD-MMH binding to anti-SARS-COV-2-S monoclonal antibodies at 37 °C. mAb Captured (mAb#) mAb Capture Level (RU) 90nM Ag Bound (RU) k a (1 / Ms) k d (1 / s) KD (M) t½ (min) mAb10913366 ± 6495.29E+055.56E-021.05E-070.2mAb10914401 + 3632.51E+051.58E-026.27E-080.7mAb10915401 ± 3931.57E+057.57E-044.84E-0915.3mAb 10920394 ± 3736.10E+053.41E-025.60E-080.3mAb10921394 ± 3871.60E+052.07E-031.29E-085.6mAb 10922405.6±1.71301.04E+069.27E-038.89E-091.2mAb 10923355 ± 3846.15E+052.76E-024.48E-080.4mAb 10924406 ± 51102.99E+056.18E-032.07E-081.9mAb10930373.9±3.5422.30E+051.87E-028.14E-080.6mAb10932406 ± 41191.43E+056.55E-044.57E-0917.6mAb10933368 ± 31242.37E+068.28E-033.49E-091.4mAb10934368 ± 31174.62E+062.32E-025.02E-090.5mAb10935430 ± 5754.37E+053.74E-028.56E-080.3mAb10936402 ± 31269.75E+055.51E-035.65E-092.1mAb10937402 ± 31079.68E+052.43E-022.51E-080.5mAb10938434 ± 31001.06E+051.12E-031.05E-0810.3mAb10939439 ± 5902.40E+059.46E-033.95E-081.2mAb 10940434 ± 31241.42E+061.23E-028.70E-090.9mAb10941418±31341.97E+051.75E-038.87E-096.6mAb10954371.8±21315.68E+051.35E-032.38E-098.6mAb10955384.1±6.3812.85E+051.26E-024.43E-080.9mAb10956383±2.3893.56E+051.30E-023.65E-080.9mAb10957322±2.11242.44E+056.19E-042.54E-0918.7mAb 10964333.3±4.61213.68E+062.08E-035.64E-105.6mAb 10965326.8±1.2672.23E+059.19E-034.12E-081.3mAb 10966350.2±2.91184.40E+051.67E-033.79E-096.9mAb10967336±2.21081.91E+052.62E-041.38E-0944.1mAb 10969349.5±3864.07E+051.59E-023.92E-080.7mAb10970393.8±3.41043.33E+057.58E-032.28E-081.5mAb10971347±1.91163.92E+059.79E-042.50E-0911.8mAb 10977341±1.41224.35E+051.31E-033.01E-098.8mAb 10982347.5±1.3671.94E+059.42E-034.85E-081.2mAb 10984422.5±0.71443.28E+051.82E-035.55E-096.3mAb10985395.5±2.51342.57E+064.23E-031.65E-092.7mAb 10986349.3±1.51298.24E+055.83E-047.07E-1019.8mAb10987354±5.3828.38E+052.51E-023.00E-080.5mAb10988364.4±2.6529.19E+055.78E-026.29E-080.2mAb10989405.6±1.91282.97E+061.16E-023.90E-091.0mAb 10996524.3±2.8431.06E+051.25E-021.19E-070.9mAb 10998271.1±0.6152.81E+057.54E-032.68E-081.5mAb11000418.2±1872.89E+059.10E-033.14E-081.3mAb11002370.1±2.5122.81E+057.54E-032.68E-081.5mAb11004297.8±0.4791.75E+061.48E-038.48E-107.8mAb11006350.2± 1.2926.28E+051.48E-022.35E-080.8mAb11008289.4±2.7381.42E+051.51E-031.06E-087.6mAb11010286.3±0.5961.67E+051.45E-028.71E-080.8mAb1932265.3±1.45NBNBNBNB Table 31: Binding kinetics parameters of SARS-COV-2 RBD-mFc binding to anti-SARS-COV-2-S monoclonal antibodies at 25 °C. mAb Captured (mAb#) mAb Capture Level (RU) 30nM Ag Bound (RU) k a (1 / Ms) k d (1 / s) KD (M) t½ (min) mAb10913107 + 0.4655.00E+062.77E-045.53E-1141.7mAb10914116 + 0.8442.59E+051.40E-045.40E-1082.5mAb10915103 + 0.2412.83E+059.13E-063.23E-111265.1mAb 10920116 + 0.9695.08E+062.55E-045.02E-1145.3mAb10921104 + 0.2392.66E+053.34E-051.25E-10345.8mAb 10922111.4 ± 0.8803.20E+065.64E-051.76E-11204.8mAb 10923110 + 1.0713.69E+061.35E-043.67E-1185.6mAb 10924121 + 0.5748.09E+057.63E-059.43E-11151.4mAb10930104.2 ± 0.9619.43E+051.71E-041.81E-1067.5mAb10932121 + 0.8602.95E+052.85E-059.67E-11405.3mAb10933108 + 0.5726.16E+066.10E-059.89E-12189.3mAb10934113 + 0.5701.12E+071.56E-041.39E-1174.0mAb10935128 + 0.8881.35E+061.07E-047.94E-11107.9mAb10936117 + 0.4741.78E+065.04E-052.83E-11229.2mAb10937106 + 0.3671.78E+065.40E-053.04E-11213.9mAb10938128 + 1.5472.42E+051.69E-057.02E-11683.4mAb10939127 + 0.8677.22E+058.74E-051.21E-10132.2mAb 10940102 + 0.4673.72E+064.66E-051.25E-11247.9mAb10941125 + 0.2683.70E+053.48E-059.43E-11331.9mAb10954108.8 ± 1862.35E+064.78E-052.03E-11241.6mAb10955109.8 ± 0.8761.20E+069.22E-057.71E-11125.3mAb10956104.1 ± 0.5741.46E+061.30E-048.87E-1188.8mAb10957104.7 ± 0.5771.02E+063.35E-053.27E-11344.8mAb 1096493.3 ± 0.3709.30E+063.69E-053.97E-12313.0mAb 1096594.2 ± 0.8636.94E+051.56E-042.25E-1074.0mAb 10966100.2 ± 0.4731.50E+063.37E-052.24E-11342.7mAb1096793.3 ± 0.2606.64E+051.35E-052.03E-11855.6mAb 10969111.4 ± 0.8804.64E+051.00E-042.16E-10115.5mAb10970113.4 ± 0.7852.19E+064.05E-041.85E-1028.5mAb 1097199 ± 0.5721.40E+064.09E-052.92E-11282.4mAb 10977109.1 ± 0.4731.82E+062.29E-051.26E-11504.4mAb 1098294.8 ± 0.1599.10E+058.06E-058.86E-11143.3mAb 10984121 ± 0.6891.39E+063.97E-052.86E-11290.9mAb10985112.7 ± 0.3778.09E+068.51E-051.05E-11135.7mAb 1098694.2 ± 0.5662.70E+062.40E-058.88E-12481.3mAb1098798 ± 0.7733.19E+064.24E-041.33E-1027.2mAb10988101.6 ± 0.6694.96E+065.08E-041.02E-1022.7mAb10989112.1 ± 0.4771.08E+079.63E-058.95E-12119.9mAb 10996104.2 ± 0.9615.62E+058.02E-041.43E-0914.4mAb 1099894.8 ± 0.1591.47E+063.58E-032.44E-093.2mAb11000112.7 ± 0.3771.11E+061.27E-041.15E-1090.9mAb11002121 ± 0.6895.54E+052.47E-034.46E-094.7mAb1100494.2 ± 0.5666.95E+056.40E-059.21E-11180.5mAb1100698 ± 0.7733.30E+055.21E-051.58E-10221.7mAb11008101.6 ± 0.6693.90E+051.92E-044.92E-1060.2mAb11010112.1 ± 0.4771.14E+068.99E-057.89E-11128.5mAb193297.8 ± 0.23NBNBNBNB Table 32: Binding kinetics parameters of SARS-COV-2 RBD-mFc binding to anti-SARS-COV-2-S monoclonal antibodies at 37 °C mAb Captured (mAb#) mAb Capture Level (RU) 30nM Ag Bound (RU) k a (1 / Ms) k d (1 / s) KD (M) t½ (min) mAb10913147 + 0.8756.32E+061.73E-032.74E-106.7mAb10914163 + 1.2706.91E+052.20E-043.18E-1052.5mAb10915141+0.6634.41E+056.89E-051.56E-10167.6mAb 10920155 + 1.1836.31E+067.53E-041.19E-1015.3mAb10921135 + 0.3624.58E+051.25E-042.73E-1092.4mAb 10922149.1 ± 1974.60E+061.60E-043.49E-1172.2mAb 10923144 + 0.8885.53E+061.85E-043.36E-1162.4mAb 10924160 + 1.1981.17E+061.31E-041.12E-1088.2mAb10930142.9 ± 0.4721.49E+065.97E-043.99E-1019.3mAb10932164+ 1.5894.48E+056.86E-051.53E-10168.4mAb10933152+0.9897.30E+067.94E-051.09E-11145.5mAb10934151 + 0.7871.36E+072.93E-042.16E-1139.4mAb10935171 +0.81015.68E+064.94E-048.69E-1123.4mAb10936161+ 1.0943.81E+066.75E-051.77E-11171.1mAb10937141+0.6854.47E+065.74E-051.29E-11201.2mAb10938172 + 1.2763.78E+056.56E-051.73E-10176.1mAb10939169 + 0.6921.06E+061.65E-041.55E-1070.0mAb 10940136 + 0.6855.54E+065.04E-059.10E-12229.2mAb10941164 + 0.81008.02E+058.01E-051.00E-10144.2mAb10954142.4 ± 0.81053.02E+061.12E-043.69E-11103.1mAb10955146.8 ± 0.7911.92E+063.88E-042.02E-1029.8mAb10956136.6 ± 0.4912.17E+063 .42E-041.58E-1033.8mAb10957137.7 ± 1.21001.55E+067.19E-054.63E-11160.6mAb 10964122.5 ± 0.3841.05E+071.26E-041.20E-1191.7mAb 10965125.7 ± 1811.42E+063.38E-042.37E-1034.2mAb 10966137.3 ± 1.1922.45E+069.93E-054.05E-11116.3mAb10967123.3 ± 0.9811.45E+063.33E-052.29E-11346.8mAb 10969149.1 ± 1978.11E+051.41E-041.74E-1081.9mAb10970149.9 ± 0.61022.18E+064.20E-041.92E-1027.5mAb10971136.1 ± 0.8902.37E+069.41E-053.97E-11122.7mAb 10977145.8 ± 0.7932.50E+061.07E-044.28E-11107.9mAb 10982125.5 ± 0.8741.23E+062.58E-042.10E-1044.8mAb 10984158.4 ± 0.71102.07E+068.36E-054.04E-11138.2mAb10985151.8 ± 0.7879.36E+063.75E-044.01E-1130.8mAb 10986125 ± 0.7834.59E+065.79E-051.26E-11199.5mAb10987131.5 ± 0.7875.04E+063.90E-047.75E-1129.6mAb10988138.6 ± 0.5828.34E+067.90E-049.47E-1114.6mAb10989146.1 ± 0.6921.38E+073.65E-042.65E-1131.6mAb 10996142.9 ± 0.4729.35E+052.47E-032.64E-094.7mAb 10998125.5 ± 0.8748.79E+051.97E-022.24E-080.6mAb11000151.8 ± 0.7871.63E+062.71E-041.66E-1042.6mAb11002158.4 ± 0.71105.06E+051.65E-023.26E-080.7mAb11004125 ± 0.7831.01E+061.18E-041.17E-1097.9mAb11006131.5 ± 0.7873.88E+057.65E-051.97E-10151.0mAb11008138.6 ± 0.5824.64E+054.05E-048.72E-1028.5mAb11010146.1 ± 0.6921.59E+068.02E-055.05E-11144.0mAb1932128 ± 0.35NBNBNBNB Example 14: Anti-SARS-CoV-2 Antibodies block RBD binding to hACE2 as determined by ELISA

[0217] An ELISA-based blocking assay was used to determine the ability of anti-SARS-CoV-2 antibodies to block the binding of the SARS-COV-2 Spike protein receptor binding domain (RBD) to its receptor, human angiotensin converting enzyme 2 (hACE2).

[0218] The SARS-CoV-2 protein used in this assay was comprised of the receptor binding domain (RBD) portion of the SARS-CoV-2 Spike protein (amino acids Arg319-Phe541) expressed with the Fc portion of the human IgG1 at the c-terminus (SARS-CoV-2 RBD-hFc) The human ACE2 protein used in the experiments was purchased from R&D Systems and was comprised of amino acids Gln18-Ser740 with a C-terminal 10X-Histidine tag (hACE2-His; NCBI Accession No. Q9BYF1).

[0219] Experiments were carried out using the following procedure. A monoclonal anti-Penta-His antibody (Qiagen) was coated at 1µg / ml in PBS on a 96-well microtiter plate overnight at 4 °C. The hACE2-His receptor was added at 0.2ug / ml in PBS and bound for two hours at room temperature (RT). Nonspecific binding sites were subsequently blocked using a 0.5% (w / v) solution of BSA in PBS. In other microtiter plates, a constant amount of 100pM of SARS-CoV-2 RBD-hFc protein was bound with anti-SARS-COV-2 antibodies and an isotype IgG1 antibody control at dilutions from 0.0008nM to 50nM in PBS +0.5% BSA. After a one-hour incubation, the mixture solutions were transferred to the microtiter plate coated hACE2-His. After 1.5 hours of incubation at RT, the wells were washed, and plate-bound SARS-COV2 was detected with goat-anti-human IgG antibody conjugated with horseradish peroxidase (HRP) (Jackson). The plates were then developed using TMB substrate solution (BD Biosciences, #555214) according to manufacturer's recommendation and absorbance at 450nm was measured on a Victor X5 plate reader.

[0220] Binding data were analyzed using a sigmoidal dose-response model within Prism™ software (GraphPad). The calculated IC50 value, defined as the concentration of antibody required to block 50% of SARS-CoV-2 RBD-hFc binding to plate-coated hACE2-His, was used as an indicator of blocking potency. Percent blocking was defined based on the background-corrected binding signal observed at the highest antibody concentration tested using this formula and reported for all tested antibodies: % Blocking = 100 − Experimental Signal highest Ab conc − Background Signal buffer Maximum Signal hEGF . mFc alone − Background Signal buffer × 100

[0221] Antibodies that blocked binding less than or equal to 50% at the highest concentration tested were classified as non-blockers and IC50 values were not reported for those antibodies.

[0222] The ability of anti-SARS-CoV-2 antibodies to block SARS-CoV-2 RBD binding to human ACE2 was assessed using a blocking ELISA. In this assay 100pM SARS-COV-2 RBD-hFc was titrated with a wide range of the concentrations of the anti-SARS-CoV-2-S antibody and the inhibition of the presence of the antibody on RBD binding to hACE2-His was evaluated. The plate-bound RBD-hFc was detected with an HRP conjugated anti-hFc antibody.

[0223] The blocking IC50s and maximum blocking at the highest tested concentrations of the anti-SARS-CoV-2-S antibodies are summarized in Table 33, and the blocking curves shown in Figures 1-8. Of the 46 antibodies tested, 44 displayed antibody concentration-dependent blocking of RBD.hFc binding to hACE-2. IC50 values ranged from 41pM to 4.5nM and maximum blocking ranging from 55% to about 100% at the highest antibody concentration tested. Two antibodies out of 46 tested showed no blocking activities under the assay conditions. The irrelevant isotype control antibody showed no blocking activity, as expected. Table 33: Blocking potency of Anti-SAR-COV-2 Antibodies on Spike RBD-hFc Binding to Immobilized Human ACE-2 mAb Assay Run # Blocking 100pM (RBD).hFc to ACE2 IC 50 ,M Blocking 100pM (RBD).hFc to ACE2 % Blocking mAb1091312.17E-1080mAb1091419.80E-1093mAb1091513.21E-1099mAb1092013.38E-1095mAb1092031.39E-1087mAb1092114.33E-1099mAb1092135.07E-1094mAb1092226.65E-1197mAb1092311.49E-1094mAb1092331.84E-1085mAb1092411.63E-1098mAb1092421.27E-1098mAb1093022.82E-1086mAb1093213.73E-1099mAb1093317.07E-1199mAb1093336.53E-1195mAb1093325.22E-11101mAb1093416.60E-1196mAb1093435.97E-1198mAb1093424.80E-1196mAb1093511.02E-1099mAb1093526.94E-1198mAb1093618.75E-1195mAb1093627.10E-1197mAb1093716.49E-1199mAb1093812.75E-1099mAb1093911.75E-1097mAb1093932.63E-1093mAb1094016.52E-1192mAb1094112.27E-10100mAb1094122.06E-10100mAb1095427.11E-1195mAb1095521.41E-1097mAb1095621.85E-1099mAb1095721.69E-1099mAb1096436.83E-1193mAb1096426.25E-1195mAb1096522.13E-1097mAb1096621.60E-1099mAb1096722.80E-1098mAb1096932.15E-1095mAb1097021.07E-1097mAb1097121.49E-1098mAb1097738.71E-1177mAb1097727.11E-1165mAb1098221.16E-1093mAb1098427.75E-1190mAb1098536.96E-1197mAb1098524.11E-1199mAb1098627.54E-1198mAb1098732.85E-1093mAb1098721.81E-1095mAb1098828.64E-1195mAb1098935.91E-1196mAb1098924.28E-1198mAb1099636.10E-0971mAb1099834.30E-0955mAb1100034.50E-0975mAb110023NBD7mAb110043NBD9mAb1100632.20E-1085mAb1100831.49E-0993mAb1101031.47E-1083mAb193250 IgG1 Control1---8mAb193250 IgG1 Control3---19mAb193250 IgG1 Control2---15Note: RBD-hFc at 100 pM was titrated with anti-SARS-COV-2-S antibodies in serial dilutions from 50nM and bound RBD-hFc on immobilized hACE2 with a 10x histidine tag, and detected with HRP-conjugated anti-hFc antibody. NBD; no blocking detected. Example 15: Cross-competition between mAb10987, mAb10989, mAb10933, and mAb10934

[0224] mAb10987, mAb10989, mAb10933, and mAb10934 were examined in cross-competition binding assays (Figure 11), identifying several pairs of non-competing mAbs with picomolar neutralization potency that could potentially be combined to form antibody cocktails, e.g., mAb10987 and mAb0933.

[0225] Epitope binning of the anti-SARS-CoV-2-S mAbs was conducted in a pre-mix sandwich format involving competing mAbs against one another in a pairwise combinatorial manner for binding to SARS-CoV-2 RBD-MMH protein using a ForteBio Octet HTX biolayer interferometry instrument (Molecular Devices ForteBio LLC, Fremont, CA) with running buffer of 10 mM HEPES, 150 mM NaCl, 0.05% (v / v) Tween-20, pH 7.4, 1 mg / mL BSA. Assays were performed at 30 °C with continuous agitation at 1000 rpm. After obtaining an initial baseline in running buffer 20 µg / mL of anti-COVID19 mAbs was captured onto anti-human Fc (AHC) biosensor tips for 300 s. To block remaining free unsaturated binding sites on AHC biosensor tips, all sensors were exposed for 240 s to blocking solution well containing 100 µg / mL irrelevant IgG1. Following this process, biosensors were immersed into wells containing pre-mix solution of 100nM SARS CoV-2 RBD-MMH protein and 600 nM of anti-COVID19 mAb binding site of a second mAbs for 300 s. Binding response at each step was recorded and specific signal was normalized by subtracting self-blocking mAb competing control from dataset. Data analysis was performed with Octet Data Analysis HT 10.0 software using the Epitope Binning.

[0226] Comparing the cross-competition binding assays with the HDX-MS results described above provides structural insights into the mechanism by which non-competing pairs of antibodies can simultaneously bind the RBD, and can thus be ideal partners for a therapeutic antibody cocktail. mAb 10987 and mAb 10933 represent such a pair of antibodies. mAb10933 targets the spike-like loop region on one edge of the ACE2 interface. Within that region, the residues that show the most significant HDX protection by mAb 10933 face upward, suggesting that the Fab region of mAb 10933 binds the RBD from the top direction, where mAb 10933 will have significant collisions with ACE2. In order to avoid competition with mAb10933, mAb 10987 only binds to the HDX-defined protected regions from the front or the lower left side (in the front view of mAb10987 in Figure 12). This is consistent with the neutralization data described above, as mAb 10987 would orient it in a position that has high probability to interfere with ACE2.Example 16: Structure determination of antibody-bound spike protein

[0227] To better understand the binding of mAb 10933 and mAb 10987 to the spike protein RBD, structural analysis was performed via cryo-electron microscopy (cryoEM). Fab fragments of mAb10933 and mAb10987 were isolated using FabALACTICA kit (Genovis). 600 µg of the mAb10933 Fab and 600 µg of mAb10987 Fab were mixed with 300 µg of SARS-CoV-2-S RBD and incubated on ice for ∼1 hour then injected into a Superdex 200 increase gel filtration column equilibrated to 50 mM Tris pH 7.5, 150 mM NaCl. Peak fractions containing the mAb10933 Fab - mAb 10987 Fab - RBD complex were collected and concentrated using a 10 kDa MWCO centrifugal filter. For cryoEM grid preparation, the protein sample was diluted to 1.5 mg / mL and 0.15% PMAL-C8 amphipol was added. 3.5 µL of protein was deposited onto a freshly plasma cleaned UltrAufoil grid (1.2 / 1.3, 300 mesh). Excess solution was blotted away using filter paper and plunge frozen into liquid ethane using a Vitrobot Mark IV. The cryoEM grid was transferred to a Titan Krios (Thermo Fisher) equipped with a K3 detector (Gatan). Movies were collected using EPU (Thermo Fisher) at 105,000x magnification, corresponding to a pixel size of 0.85 Å. A dose rate of 15 electrons per pixel per second was used and each movie was 2 seconds, corresponding to a total dose of ∼40 electrons per Å2.

[0228] All cryoEM data processing was carried out using cryoSPARC v2.14.2. 2,821 movies were aligned using patch motion correction and patch CTF estimation. 2,197 aligned micrographs were selected for further processing on the basis of estimated defocus values and CTF fit resolutions. An initial set of particles picked using blob picker were subjected to 2D classification to generate templates for template picking. 989,553 particles picked by template picking were subjected to multiple rounds of 2D classification to remove unbound fabs and particles containing an incomplete complex. Ab initio reconstruction with three classes generated a single class containing 61,707 particles that corresponded to the mAb10933 Fab - mAb10987 Fab - RBD complex. Heterogenous refinement of the particles in this class followed by nonuniform refinement resulted in a 3.9 Å resolution (FSC=0.143) map containing 48,140 particles that was used for model building. Into this map, models of the RBD (taken from PDB code 6M17) and the two Fabs (taken from prior antibody structures, except for the lambda light chain of mAb10987 which came from PDB code 5U15), were manually placed. These models were then manually rebuilt using Coot and real-space refined against the map using Phenix.

[0229] Confirming the above-described data, single-particle cryoEM of the complex of SARS-CoV-2 spike RBD bound to Fab fragments of mAb10933 and mAb10987 shows that the two antibodies in this cocktail can simultaneously bind to distinct regions of the RBD (Figure 13A, Figure 13B, and Figure 14). A 3D reconstructed map of the complex with nominal resolution of 3.9Å shows that the both Fab fragments bind at different epitopes on the RBD, confirming that they are non-competing antibodies. mAb10933 binds at the top of the RBD, extensively overlapping the binding site for ACE2. On the other hand, the epitope for mAb10987 is located on the side of the RBD, well away from the mAb10933 epitope, and has little to no overlap with the ACE2 binding site.Example 17: Cross-competition between anti-SARS-CoV-2-S mAbs

[0230] Binding competition between anti-SARS-CoV-2-S monoclonal antibodies (mAbs) was determined using a real time, label-free bio-layer interferometry (BLI) assay on the Octet HTX biosensor platform (Pall ForteBio Corp.). The entire experiment was performed at 25 °C in 10mM HEPES, 150mM NaCl, 3mM EDTA, and 0.05% v / v Surfactant Tween-20, 1mg / mL BSA, pH 7.4 (HBS-EBT) buffer with the plate shaking at a speed of 1000rpm. To assess whether two mAbs were able to compete with one another for binding to their respective epitopes on the SARS-COV-2-S RBD extracellular domain expressed with a C-terminal myc-myc-hexahistidine (SARS-COV-2 RBD-MMH), ∼0.51nm of SARS-COV-2-S RBD-MMH was first captured onto anti-Penta-His antibody coated Octet biosensor tips (Fortebio Inc, # 18-5122) by submerging the biosensor tips for 1 minute in wells containing a 10 µg / mL solution of SARS-COV-2-S RBD-MMH. The SARS-COV-2-S RBD-MMH captured biosensor tips were then saturated with a first anti-SARS-CoV-2-S monoclonal antibody (subsequently referred to as mAb-1) by dipping into wells containing 50µg / mL solution of mAb-1 for 5 minutes. The biosensor tips were then subsequently dipped into wells containing 50µg / mL solution of a second anti-SARS-CoV-2 monoclonal antibody (subsequently referred to as mAb-2) for 5 minutes. The biosensor tips were washed in HBS-ETB buffer in between every step of the experiment. The real-time binding response was monitored during the entire course of the experiment and the binding response at the end of every step was recorded. The response of mAb-2 binding to SARS-COV-2 RBD-MMH pre-complexed with mAb-1 was compared and competitive / non-competitive behavior of different anti-SARS-CoV-2 monoclonal antibodies was determined as shown in Table 34. Table 34: Cross-competition between anti-SARS-CoV-2-S antibodies mAb-1mAb-2 Competing with mAb-1mAb10924mAb10989mAb10920mAb10913mAb10923mAb10977mAb10930mAb10969mAb10988mAb10964mAb10996mAb10966mAb10998mAb10984mAb11006mAb10921mAb10971mAb10938mAb10932mAb10970mAb10957mAb10956mAb10941mAb10939mAb10935mAb10914mAb10982mAb11008mAb11002mAb10933mAb10940mAb10922mAb11004mAb10937mAb10936mAb10934mAb10924mAb10977mAb10989mAb10920mAb10913mAb10923mAb10930mAb10969mAb10988mAb10964mAb10996mAb10966mAb10998mAb10984mAb11006mAb10921mAb10971mAb10938mAb10932mAb10970mAb10957mAb10956mAb10941mAb10939mAb10935mAb10914mAb10982mAb11008mAb10915mAb10965mAb10967mAb10986mAb10955mAb10954mAb10933mAb11000mAb10985mAb10937mAb10936mAb10934mAb10989mAb10977mAb10924mAb10920mAb10913mAb10923mAb10930mAb10969mAb10988mAb10964mAb10996mAb10966mAb10998mAb10984mAb11006mAb10921mAb10971mAb10938mAb10932mAb10970mAb10957mAb10956mAb10941mAb10939mAb10935mAb10914mAb10982mAb11008mAb10915mAb10965mAb10967mAb10986mAb10955mAb10954mAb10933mAb10987mAb10940mAb10922mAb11004mAb10937mAb10936mAb10934mAb10920mAb10977mAb10924mAb10989mAb10913mAb10923mAb10930mAb10969mAb10988mAb10964mAb10996mAb10966mAb10998mAb10984mAb11006mAb10921mAb10971mAb10938mAb10932mAb10970mAb10957mAb10956mAb10941mAb10939mAb10935mAb10914mAb10982mAb11008mAb10915mAb10965mAb10967mAb10986mAb10955mAb10954mAb11002mAb10933mAb10987mAb10940mAb10922mAb11004mAb10937mAb10936mAb10934mAb10913mAb10977mAb10924mAb10989mAb10920mAb10923mAb10930mAb10969mAb10988mAb10964mAb10996mAb10966mAb10998mAb10984mAb11006mAb10921mAb10971mAb10938mAb10932mAb10970mAb10957mAb10956mAb10941mAb10939mAb10935mAb10914mAb10982mAb11008mAb10915mAb10965mAb10967mAb10986mAb10955mAb10954mAb11002mAb10933mAb11004mAb10937mAb10936mAb10934mAb10923mAb10977mAb10924mAb10989mAb10920mAb10913mAb10930mAb10969mAb10988mAb10964mAb10996mAb10966mAb10998mAb10984mAb11006mAb10921mAb10971mAb10938mAb10932mAb10970mAb10957mAb10956mAb10941mAb10939mAb10935mAb10914mAb10982mAb11008mAb10915mAb10965mAb10967mAb10986mAb10955mAb10954mAb11002mAb10933mAb11004mAb10937mAb10936mAb10934mAb10930mAb10977mAb10924mAb10989mAb10920mAb10913mAb10923mAb10969mAb10988mAb10964mAb10996mAb10966mAb10998mAb10984mAb11006mAb10921mAb10971mAb10938mAb10932mAb10970mAb10957mAb10956mAb10941mAb10939mAb10935mAb10914mAb10982mAb11008mAb10915mAb10965mAb10967mAb10986mAb10955mAb10954mAb11002mAb10933mAb10937mAb10936mAb10934mAb10969mAb10977mAb10924mAb10989mAb10920mAb10913mAb10923mAb10930mAb10988mAb10964mAb10966mAb10998mAb10984mAb11006mAb10921mAb10971mAb10938mAb10932mAb10970mAb10957mAb10956mAb10941mAb10939mAb10935mAb10914mAb10982mAb11008mAb10915mAb10965mAb10967mAb10986mAb10955mAb10954mAb11002mAb10933mAb10985mAb10937mAb10936mAb10934mAb10988mAb10977mAb10924mAb10989mAb10920mAb10913mAb10923mAb10930mAb10969mAb10964mAb10996mAb10966mAb10998mAb10984mAb11006mAb10921mAb10971mAb10938mAb10932mAb10970mAb10957mAb10956mAb10941mAb10939mAb10935mAb10914mAb10982mAb11008mAb10915mAb10965mAb10967mAb10986mAb10955mAb10954mAb10933mAb10936mAb10934mAb10964mAb10977mAb10924mAb10989mAb10920mAb10913mAb10923mAb10930mAb10969mAb10988mAb10996mAb10966mAb10998mAb10984mAb11006mAb10921mAb10971mAb10938mAb10932mAb10970mAb10957mAb10956mAb10941mAb10939mAb10935mAb10914mAb10982mAb11008mAb10915mAb10965mAb10967mAb10986mAb10955mAb10954mAb11002mAb10933mAb10936mAb10934mAb10996mAb10977mAb10924mAb10989mAb10920mAb10913mAb10923mAb10930mAb10969mAb10988mAb10964mAb10966mAb10998mAb10984mAb11006mAb10921mAb10971mAb10938mAb10932mAb10970mAb10957mAb10956mAb10941mAb10939mAb10935mAb10914mAb10982mAb11008mAb10915mAb10965mAb10967mAb10986mAb10955mAb10954mAb11002mAb10933mAb10985mAb10934mAb10966mAb10977mAb10924mAb10989mAb10920mAb10913mAb10923mAb10930mAb10969mAb10988mAb10964mAb10996mAb10998mAb10984mAb11006mAb10921mAb10971mAb10938mAb10932mAb10970mAb10957mAb10956mAb10941mAb10939mAb10935mAb10914mAb10982mAb11008mAb10915mAb10965mAb10967mAb10986mAb10955mAb10954mAb11002mAb10933mAb10985mAb10934mAb10998mAb10977mAb10924mAb10989mAb10920mAb10913mAb10923mAb10930mAb10969mAb10988mAb10964mAb10996mAb10966mAb10984mAb11006mAb10921mAb10971mAb10938mAb10932mAb10970mAb10957mAb10956mAb10941mAb10939mAb10935mAb10914mAb10982mAb11008mAb10915mAb10965mAb10967mAb10986mAb10955mAb10954mAb11002mAb10985mAb10936mAb10984mAb10977mAb10924mAb10989mAb10920mAb10913mAb10923mAb10930mAb10969mAb10988mAb10964mAb10996mAb10966mAb10998mAb11006mAb10921mAb10971mAb10938mAb10932mAb10970mAb10957mAb10956mAb10941mAb10939mAb10935mAb10914mAb10982mAb11008mAb10915mAb10965mAb10967mAb10986mAb10955mAb10954mAb11002mAb10985mAb11006mAb10977mAb10924mAb10989mAb10920mAb10913mAb10923mAb10930mAb10969mAb10988mAb10964mAb10996mAb10966mAb10998mAb10984mAb10921mAb10971mAb10938mAb10932mAb10970mAb10957mAb10956mAb10941mAb10939mAb10935mAb10914mAb10982mAb11008mAb10915mAb10965mAb10967mAb10986mAb10955mAb10954mAb11002mAb10933mAb10985mAb10921mAb10977mAb10924mAb10989mAb10920mAb10913mAb10923mAb10930mAb10969mAb10988mAb10964mAb10996mAb10966mAb10998mAb10984mAb11006mAb10971mAb10938mAb10932mAb10970mAb10957mAb10956mAb10941mAb10939mAb10935mAb10914mAb10982mAb11008mAb10915mAb10965mAb10967mAb10986mAb10955mAb10954mAb11002mAb10985mAb10971mAb10977mAb10924mAb10989mAb10920mAb10913mAb10923mAb10930mAb10969mAb10988mAb10964mAb10996mAb10966mAb10998mAb10984mAb11006mAb10921mAb10938mAb10932mAb10970mAb10957mAb10956mAb10941mAb10939mAb10935mAb10914mAb10982mAb11008mAb10915mAb10965mAb10967mAb10986mAb10955mAb10954mAb11002mAb10985mAb10938mAb10977mAb10924mAb10989mAb10920mAb10913mAb10923mAb10930mAb10969mAb10988mAb10964mAb10996mAb10966mAb10998mAb10984mAb11006mAb10921mAb10971mAb10932mAb10970mAb10957mAb10956mAb10941mAb10939mAb10935mAb10914mAb10982mAb11008mAb10915mAb10965mAb10967mAb10986mAb10955mAb10954mAb11002mAb10985mAb10932mAb10977mAb10924mAb10989mAb10920mAb10913mAb10923mAb10930mAb10969mAb10988mAb10964mAb10996mAb10966mAb10998mAb10984mAb11006mAb10921mAb10971mAb10938mAb10970mAb10957mAb10956mAb10941mAb10939mAb10935mAb10914mAb10982mAb11008mAb10915mAb10965mAb10967mAb10986mAb10955mAb10954mAb11002mAb10985mAb10970mAb10977mAb10924mAb10989mAb10920mAb10913mAb10923mAb10930mAb10969mAb10988mAb10964mAb10996mAb10966mAb10998mAb10984mAb11006mAb10921mAb10971mAb10938mAb10932mAb10957mAb10956mAb10941mAb10939mAb10935mAb10914mAb10982mAb11008mAb10915mAb10965mAb10967mAb10986mAb10955mAb10954mAb11002mAb10985mAb10936mAb10957mAb10977mAb10924mAb10989mAb10920mAb10913mAb10923mAb10930mAb10969mAb10988mAb10964mAb10996mAb10966mAb10998mAb10984mAb11006mAb10921mAb10971mAb10938mAb10932mAb10970mAb10956mAb10941mAb10939mAb10935mAb10914mAb10982mAb11008mAb10915mAb10965mAb10967mAb10986mAb10955mAb10954mAb11002mAb10985mAb10956mAb10977mAb10924mAb10989mAb10920mAb10913mAb10923mAb10930mAb10969mAb10988mAb10964mAb10996mAb10966mAb10998mAb10984mAb11006mAb10921mAb10971mAb10938mAb10932mAb10970mAb10957mAb10941mAb10939mAb10935mAb10914mAb10982mAb11008mAb10915mAb10965mAb10967mAb10986mAb10955mAb10954mAb11002mAb10985mAb10941mAb10977mAb10924mAb10989mAb10920mAb10913mAb10923mAb10930mAb10969mAb10988mAb10964mAb10996mAb10966mAb10998mAb10984mAb11006mAb10921mAb10971mAb10938mAb10932mAb10970mAb10957mAb10956mAb10939mAb10935mAb10914mAb10982mAb11008mAb10915mAb10965mAb10967mAb10986mAb10955mAb10954mAb11002mAb10985mAb10939mAb10977mAb10924mAb10989mAb10920mAb10913mAb10923mAb10930mAb10969mAb10988mAb10964mAb10996mAb10966mAb10998mAb10984mAb11006mAb10921mAb10971mAb10938mAb10932mAb10970mAb10957mAb10956mAb10941mAb10935mAb10914mAb10982mAb11008mAb10915mAb10965mAb10967mAb10986mAb10955mAb10954mAb11002mAb10935mAb10977mAb10924mAb10989mAb10920mAb10913mAb10923mAb10930mAb10969mAb10988mAb10964mAb10996mAb10966mAb10998mAb10984mAb11006mAb10921mAb10971mAb10938mAb10932mAb10970mAb10957mAb10956mAb10941mAb10939mAb10914mAb10982mAb11008mAb10915mAb10965mAb10967mAb10986mAb10955mAb10954mAb11002mAb10985mAb10914mAb10977mAb10924mAb10989mAb10920mAb10913mAb10923mAb10930mAb10969mAb10988mAb10964mAb10996mAb10966mAb10998mAb10984mAb11006mAb10921mAb10971mAb10938mAb10932mAb10970mAb10957mAb10956mAb10941mAb10939mAb10935mAb10982mAb11008mAb10915mAb10965mAb10967mAb10986mAb10955mAb10954mAb11002mAb10933mAb10985mAb10982mAb10977mAb10924mAb10989mAb10920mAb10913mAb10923mAb10930mAb10969mAb10988mAb10964mAb10996mAb10966mAb10998mAb10984mAb11006mAb10921mAb10971mAb10938mAb10932mAb10970mAb10957mAb10956mAb10941mAb10939mAb10935mAb10914mAb11008mAb10915mAb10965mAb10967mAb10986mAb10955mAb10954mAb11002mAb10985mAb11008mAb10977mAb10924mAb10989mAb10920mAb10913mAb10923mAb10930mAb10969mAb10988mAb10964mAb10996mAb10966mAb10998mAb10984mAb11006mAb10921mAb10971mAb10938mAb10932mAb10970mAb10957mAb10956mAb10941mAb10939mAb10935mAb10914mAb10982mAb10915mAb10965mAb10967mAb10986mAb10955mAb10954mAb11002mAb10933mAb10985mAb10915mAb10924mAb10989mAb10920mAb10913mAb10923mAb10930mAb10969mAb10988mAb10964mAb10996mAb10966mAb10998mAb10984mAb11006mAb10921mAb10971mAb10938mAb10932mAb10970mAb10957mAb10956mAb10941mAb10939mAb10935mAb10914mAb10982mAb11008mAb10965mAb10967mAb10986mAb10955mAb10954mAb11002mAb10985mAb10965mAb10924mAb10989mAb10920mAb10913mAb10923mAb10930mAb10969mAb10988mAb10964mAb10996mAb10966mAb10998mAb10984mAb11006mAb10921mAb10971mAb10938mAb10932mAb10970mAb10957mAb10956mAb10941mAb10939mAb10935mAb10914mAb10982mAb11008mAb10915mAb10967mAb10986mAb10955mAb10954mAb11002mAb10985mAb10967mAb10924mAb10989mAb10920mAb10913mAb10923mAb10930mAb10969mAb10988mAb10964mAb10996mAb10966mAb10998mAb10984mAb11006mAb10921mAb10971mAb10938mAb10932mAb10970mAb10957mAb10956mAb10941mAb10939mAb10935mAb10914mAb10982mAb11008mAb10915mAb10965mAb10986mAb10955mAb10954mAb11002mAb10985mAb10986mAb10924mAb10989mAb10920mAb10913mAb10923mAb10930mAb10969mAb10988mAb10964mAb10996mAb10966mAb10998mAb10984mAb11006mAb10921mAb10971mAb10938mAb10932mAb10970mAb10957mAb10956mAb10941mAb10939mAb10935mAb10914mAb10982mAb11008mAb10915mAb10965mAb10967mAb10955mAb10954mAb11002mAb10985mAb10955mAb10924mAb10989mAb10920mAb10913mAb10923mAb10930mAb10969mAb10988mAb10964mAb10996mAb10966mAb10998mAb10984mAb11006mAb10921mAb10971mAb10938mAb10932mAb10970mAb10957mAb10956mAb10941mAb10939mAb10935mAb10914mAb10982mAb11008mAb10915mAb10965mAb10967mAb10986mAb10954mAb11002mAb10985mAb10954mAb10924mAb10989mAb10920mAb10913mAb10923mAb10930mAb10969mAb10988mAb10964mAb10996mAb10966mAb10998mAb10984mAb11006mAb10921mAb10971mAb10938mAb10932mAb10970mAb10957mAb10956mAb10941mAb10939mAb10935mAb10914mAb10982mAb11008mAb10915mAb10965mAb10967mAb10986mAb10955mAb11002mAb10985mAb11002mAb10977mAb10920mAb10913mAb10923mAb10930mAb10969mAb10964mAb10996mAb10966mAb10998mAb10984mAb11006mAb10921mAb10971mAb10938mAb10932mAb10970mAb10957mAb10956mAb10941mAb10939mAb10935mAb10914mAb10982mAb11008mAb10915mAb10965mAb10967mAb10986mAb10955mAb10954mAb10933mAb10985mAb10936mAb10933mAb10977mAb10924mAb10989mAb10920mAb10913mAb10923mAb10930mAb10969mAb10988mAb10964mAb10996mAb10966mAb11006mAb10914mAb11008mAb11002mAb11000mAb10937mAb10936mAb10934mAb11000mAb10924mAb10933mAb10985mAb11010mAb10985mAb10924mAb10969mAb10996mAb10966mAb10998mAb10984mAb11006mAb10921mAb10971mAb10938mAb10932mAb10970mAb10957mAb10956mAb10941mAb10935mAb10914mAb10982mAb11008mAb10915mAb10965mAb10967mAb10986mAb10955mAb10954mAb11002mAb11000mAb11010mAb11010mAb11000mAb10985mAb10987mAb10989mAb10920mAb10940mAb10922mAb11004mAb10937mAb10936mAb10934mAb10940mAb10977mAb10989mAb10920mAb10987mAb10922mAb11004mAb10937mAb10936mAb10934mAb10922mAb10977mAb10989mAb10920mAb10987mAb10940mAb11004mAb10937mAb10936mAb10934mAb11004mAb10977mAb10989mAb10920mAb10913mAb10923mAb10987mAb10940mAb10922mAb10937mAb10936mAb10934mAb10937mAb10977mAb10924mAb10989mAb10920mAb10913mAb10923mAb10930mAb10969mAb10933mAb10987mAb10940mAb10922mAb11004mAb10936mAb10934mAb10936mAb10977mAb10924mAb10989mAb10920mAb10913mAb10923mAb10930mAb10969mAb10988mAb10964mAb10998mAb10970mAb11002mAb10933mAb10987mAb10940mAb10922mAb11004mAb10937mAb10934mAb10934mAb10977mAb10924mAb10989mAb10920mAb10913mAb10923mAb10930mAb10969mAb10988mAb10964mAb10996mAb10966mAb10933mAb10987mAb10940mAb10922mAb11004mAb10937mAb10936 Example 18: pH sensitivity of anti-SARS-CoV-2-S monoclonal antibodies binding to monomeric SARS-CoV-2-S RBD reagents measured at 37 °C

[0231] The dissociation rate constants (k d ) for different anti-SARS-CoV-2-S monoclonal antibodies in pH 7.4, pH 6.0, and pH5.0 buffers were determined using a real-time surface plasmon resonance (SPR)-based Biacore T200 biosensor. All binding studies were performed at 37 °C using three running buffers, (i) PBS, 0.05% v / v Surfactant Tween-20, pH7.4 (PBS-T-pH7.4) (ii) PBS, 0.05% v / v Surfactant Tween-20, pH6.0 (PBS-T-pH6.0), and (iii) PBS, 0.05% v / v Surfactant Tween-20, pH5.0 (PBS-T-pH5.0). The Biacore CM5 sensor chip surface was first derivatized by amine coupling with a mouse anti-human Fc specific mAb (Regeneron) to capture anti-SARS-CoV-2-S monoclonal antibodies. Binding studies were performed on human SARS-COV-2-S RBD extracellular domain expressed with a C-terminal myc-myc-hexahistidine (SARS-COV-2 RBD-MMH), Single concentrations of SARS-COV-2-S RBD-MMH (90nM) prepared in PBS-T-pH 7.4 buffer were injected at a flow rate of 25µL / min for 3 minutes followed by the dissociation of bound SARS-COV-2-S RBD-MMH in PBS-T-pH 7.4, PBS-T-pH 6.0 or PBS-T PBS-T-pH 5.0 running buffers for 5 minutes.

[0232] The dissociation rate constants (k d ) in four pH running buffers were determined by fitting the real-time binding sensorgrams to a 1:1 binding model using Scrubber 2.0c curve-fitting software. The dissociative half-life (t½) was calculated from the k d values as: t½ min = ln 2 60 ∗ kd

[0233] The k d and t½ values for SARS-COV-2-S RBD-MMH binding to different anti-SARS-CoV-2-S monoclonal antibodies in PBS-T-pH 7.4 followed by dissociation in PBS-T-pH 7.4 and PBS-T-pH 6.0 at 37 °C are shown in Table 35. The k d and t½ values for SARS-COV-2-S RBD-MMH binding to different anti-SARS-CoV-2-S monoclonal antibodies in PBS-T-pH 7.4 followed by dissociation in PBS-T-pH 7.4 and PBS-T-pH 5.0 at 37 °C are shown in Table 36. The comparison of the dissociative half-life (t½) of SARS-COV-2 RBD-MMH in pH 7.4, pH 6.0 and pH 5.0 buffers. Table 35: Binding of SARS-COV-2-S RBD-MMH to anti-SARS-CoV-2-S monoclonal antibodies in PBS-T-pH 7.4 buffer and dissociation in PBS-T-pH 7.4 & pH 6.0 buffer at 37°C.Running Buffer: PBS-T, pH7.4 @ 37C°Running Buffer: PBS-T, Chase in pH6.0 @ 37C°t½ RatiomAb CapturedmAb Capture Level (RU)90nM RBD.mmh Bound (RU)kd (1 / s)t½ (min)mAb Capture Level (RU)90nM RBD.mmh Bound (RU)kd (1 / s)t½ (min)pH7.4 / pH6.0mAb10913427694.30E-020.3421674.38E-020.31mAb10914388699.41E-031.2386621.16E-0211.2mAb10915319847.43E-0415.5312881.51E-037.72mAb109324321336.60E-0417.54381311.28E-0391.9mAb109333601247.85E-031.53531198.53E-031.41.1mAb109343411071.74E-020.73341082.11E-020.51.2mAb10935407762.78E-020.4404721.71E-020.70.6mAb109363811245.29E-032.23751208.69E-031.31.6mAb10937330942.09E-020.6323982.09E-020.61mAb109243851115.69E-0323791101.20E-0212.1mAb10938407951.05E-0311407902.99E-033.92.8mAb109403431191.08E-021.13391271.04E-021.11mAb109413981291.65E-0373961272.04E-035.71.2mAb10920383792.47E-020.5380735.39E-020.22.2mAb10921345891.79E-036.5339922.01E-020.611.3mAb10923355872.35E-020.5349882.43E- 020.51mAb10939410909.48E-031.2412831.18E-0211.2mAb10922251859.07E-031.3240929.61E- 031.21.1mAb10930377501.92E-020.6372461.67E-020.70.9mAb10982389799.90E-031.2387747.72E-031.50.8mAb109843781331.71E-036.83701351.94E-035.91.1mAb109854571723.63E-033.24641723.19E-033.60.9mAb109864131556.29E-0418.44111521.24E-039.32mAb109873791052.37E-020.53721091.83E-020.60.8mAb109884671094.35E-020.34691035.37E-020.21.2mAb109893821269.32E-031.23751197.36E-031.60.8mAb10970340937.65E-031.5334966.37E-031.80.8mAb109713501259.44E-0412.23421251.27E-039.11.3mAb109643801401.94E-0363791372.51E-034.61.3mAb10965290658.66E-031.3281709.47E-031.21.1mAb109664171521.60E-037.24091491.41E- 038.20.9mAb109673721182.98E- 0438.83671153.45E-0433.51.2mAb109543361181.74E-036.63311242.70E-034.31.6mAb109554041001.22E-020.9403971.46E-020.81.2mAb109564521141.25E-020.94461061.50E-020.81.2mAb109573881365.80E-0419.93821407.67E-0415.11.3mAb10977293441.59E-020.7285443.39E-020.32.1mAb10969340721.86E-020.6336711.01E-021.10.5mAb10996408354.69E-020.2405374.37E-020.30.9mAb10998308202.86E-020.4307192.84E-020.41mAb11002373102.60E-020.436845.91E-0320.2mAb11000404881.48E-037.8403902.85E-034.11.9mAb11004356971.47E-020.8353962.09E-020.61.4mAb110063981051.46E-037.9398981.98E-035.81.4mAb110083411121.33E-038.73381181.28E-0391mAb110104321573.90E-0334311567.51E-031.51.9Isotype Control4304NBNB4279NBNBNB Table 36: Binding of SARS-COV-2-S RBD-MMH to anti-SARS-CoV-2 monoclonal antibodies in PBS-T-pH7.4 buffer and the dissociation in PBS-T-pH 7.4 & pH 5.0 buffer at 37°C. Running Buffer: PBS-T, pH7.4 @ 37C°Running Buffer: PBS-T, Chase in pH5.0 @ 37C°t½ RatiomAb CapturedmAb Capture Level (RU)90nM RBD.mmh Bound (RU)kd (1 / s)t½ (min)mAb Capture Level (RU)90nM RBD.mmh Bound (RU)kd (1 / s)t½ (min)pH7.4 / pH5.0mAb10913427694.30E-020.3430653.53E-020.30.8mAb10914388699.41E-031.2391571.00E-021.21.1mAb10915319847.43E-0415.5316942.05E-035.62.8mAb109324321336.60E-0417.54521312.11E-035.53.2mAb109333601247.85E-031.53531141.14E-0211.5mAb109343411071.74E-020.73381091.71E-020.71mAb10935407762.78E-020.4413701.28E-020.90.5mAb109363811245.29E-032.23791161.60E-020.73mAbl0937330942.09E-020.63261041.55E-020.70.7mAb109243851115.69E-0323901131.48E- 020.82.6mAb10938407951.05E-0311417827.61E-031.57.2mAb109403431191.08E-021.13411358.23E-031.40.8mAb109413981291.65E-0374071282.21E-035.21.3mAb10920383792.47E-020.5382682.93E-020.41.2mAb10921345891.79E-036.53451005.46E-020.230.6mAb10923355872.35E-020.5357902.13E-020.50.9mAb10939410909.48E-031.2419781.14E-0211.2mAb10922251859.07E-031.32401028.08E-031.40.9mAb10930377501.92E-020.6383441.20E-0210.6mAb10982389799.90E-031.2391666.27E-031.80.6mAb109843781331.71E-036.83781402.33E-0351.4mAb109854571723.63E-033.24711703.36E-033.40.9mAb109864131556.29E-0418.44171483.18E-033.65.1mAb109873791052.37E-020.53771158.80E-3-01.30.4mAb109884671094.35E-020.34921036.98E-020.21.6mAb109893821269.32E-031.23791056.13E-031.90.7mAb10970340937.65E-031.53411026.02E-031.90.8mAb109713501259.44E-0412.23521291.70E-036.81.8mAb109643801401.94E-0363791323.02E-033.81.6mAb10965290658.66E-031.3284777.40E-031.60.9mAb109664171521.60E-037.24221511.25E-039.20.8mAb109673721182.98E-0438.83771144.05E-0428.51.4mAb109543361181.74E-036.63351325.33E-032.23.1mAb109554041001.22E-020.9416961.85E-020.61.5mAb109564521141.25E-020.94621012.18E-020.51.7mAb109573881365.80E-0419.93901467.93E-0414.61.4mAb10977293441.59E-020.7287464.81E-020.23mAb10969340721.86E-020.6344691.33E-020.90.7mAb10996408354.69E-020.2415429.02E-020.11.9mAb10998308202.86E-020.4311212.32E-020.50.8mAb11002373102.60E-020.437117.15E-0416.20mAb11000404881.48E-037.8411962.46E-034.71.7mAb11004356971.47E-020.8362982.70E-020.41.8mAb110063981051.46E-037.9411932.10E-035.51.4mAb110083411121.33E-038.73401271.10E-0310.50.8mAb110104321573.90E-0334401567.15E-031.61.8Isotype Control4304NBNB43515NBNBNB Example 19: Anti-SARS-CoV-2-S antibodies binding to virus-like particles

[0234] To investigate the ability of a panel of anti-SARS-CoV-2 monoclonal antibodies to bind SARS-CoV-2 Spike glycoprotein, an in vitro binding assay utilizing vesicular stomatitis virus (VSV) pseudotyped with SARS-CoV-2 Spike protein in an electrochemiluminescence based detection platform (MSD) was developed.

[0235] Pseudotyped vesicular stomatitis virus (VSV) viral like particles (VLPs) were generated from HEK293T cells to transiently express the SARS-CoV-2 Spike Protein (Accession number MN908947.3, amino acids 16-1211). VLPs expressing VSV only were also generated as a negative binding control.

[0236] Experiments were carried out according to following procedure. VLPs from the two sources described above were diluted in PBS, seeded into the 96-well carbon electrode plates (MULTI-ARRAY high bind plate, MSD) and incubated overnight at 4 °C to allow the VLPs to adhere. Nonspecific binding sites were blocked by 2% BSA (w / v) in PBS for 1 hour at room temperature. To the plate-bound particles, anti-SARS-CoV-2 antibodies and a non-binding human IgG1 control, diluted in PBS + 0.5% BSA at a range of concentrations from 0.0008nM to 50nM, and buffer with no antibody were added in duplicate and the plates incubated for 1 hour at room temperature with shaking. The plates were then washed with 1X PBS to remove the unbound antibodies using an AquaMax2000 plate washer (MDS Analytical Technologies). The plate-bound antibodies were detected with a SULFO-TAGTM-conjugated anti-human IgG antibody (Jackson Immunoresearch) for 1 hour at room temperature. After washes, the plates were developed with the Read Buffer (MSD) according to manufacturer's recommended procedure and the luminescent signals were recorded with a SECTOR Imager 600 (Meso Scale Development) instrument. The direct binding signals (in RLU) were captured for SARS-CoV-2-expressing VLPs and VSV only VLPs.

[0237] The ability of the anti-SARS-CoV-2-S monoclonal antibodies to bind to SARS-CoV-2-S expressing VLPs compared with binding to irrelevant VSV expressing VLPs was assessed using an immunobinding assay. Binding to the immobilized VLPs on 96-well High Bind plates (MSD) was performed with a series of antibody dilutions and the bound antibodies were detected using SULFO-TAGTM-conjugated anti-human IgG. The binding signals from electrochemiluminescence were recorded on a Sector Imager 600 (MSD). RLU values were determined for the antibody binding to VLPs. All antibodies displayed a concentration-dependent binding and the ratios of binding on the SARS-COV-2-S expressing VLPs to VSV only were analyzed at 5.5nM and 0.20nM.

[0238] The binding results of anti-SARS-CoV-2-S mAbs at the two concentrations to VSV / spike and VSV-only VLPs are summarized in Table 37. Of 46 antibodies tested, 44 antibodies bound specifically to VSV / spike with a ratio to VSV of 3 or higher at either concentration. At 0.2nM antibody, the ratio of VSV / spike to VSV ranged from 3 to 56, and at 5nM the ratio ranged from 3 to 303. Although two antibodies (mAb10998 and mAb11002) displayed weak binding to the VSV / Spike VLPs, with ratios of less than 3 to the VSV VLPs, the signals at 5nM were higher on the VSV / spike than the VSV. An irrelevant IgG1 isotype antibody showed minimal binding, as expected. Table 37: Specificity of anti-SARS-COV-2-S antibodies binding to spike protein-expressing VSV VLPs vs VSV by ElectrochemiluminescenceAntibody Binding Signal (RLU) Ratio Antibody Concentration 5.5nM 0.20nM 5.5nM 0.20nM mAb# VSV / Spike VSV VSV / Spike VSV VSV / Spike :VSV VSV / Spike :VSV Expt# mAb10913114030243451491mAb 1091461391823911853111mab109151676370228687724371mAb 109207757253613321023133mAb 1092181747059388912113mAb 109221458129562391162mAb1092314441324463311143mAb 10924192235337557571mAb10930148829142938542mAb1093211774105128235113371mAb1093363182446298161mAb109341099124648299221mAb109352526387611477131mAb10936508722817024122421mAb10937105620437443591mAb109381141839512233729331mAb109394656637948997103mAb 10940947583843416111mAb 1094172976995817106561mAb1095497272052114484782mAb10955218927039755862mAb10956100637326371362mAb109571062412716066884112mAb 109641425247948626303242mAb109651039872795812142mAb 1096691769714068895152mAb 10967107441221090328882mAb 10969116333426242363mAb 1097056407610615074132mAb 10971799560137227134202mAb 109772689542839330165622mAb 10982187522042736962mAb 1098491421952270334792mAb 10985149790529651782mAb 109861115517723156563112mAb 10987114616869953782mAb 1098896716343839642mAb10989219512815336617132mAb 109968123098265313mAb 1099822531590122104113mAb 110005801399447423mAb 110024192834750113mAb 110041061563862819143mAb 1100626528629971592474293mAb 110085084880281133mAb 11010349649630533IgGl Isotype Control113843221121IgGl Isotype Control1671277535123IgGl Isotype Control94999931112 Example 20: Anti-SARS-CoV-2-S antibodies binding to spike protein-expressing cells

[0239] To investigate the ability of a panel of anti-SARS-CoV-2-S monoclonal antibodies to bind to SARS-CoV-2-S expressing cells, an in vitro binding assay utilizing SARS-CoV-2-S expressing cells in an electrochemiluminescence based detection platform (MSD) was developed.

[0240] Jurkat / Tet3G / hCD20 / Tet-3G inducible cells were engineered to transiently express the SARS-CoV-2 Spike Protein (Accession number MN908947.3, amino acids 16-1211, Jurkat / Tet3G / hCD20 / Tet-On 3G Inducible COVID-19 Spike Protein High Sorted), and flow cytometry sorted for selection of high expression of the SARS-CoV-2 protein. Parental Jurkat / Tet3G / hCD20 / Tet-3G were also included in the experiments as a negative binding control.

[0241] Experiments were carried out according to following procedure. Cells from the two lines described above were induced with 1 µg / ml doxycycline at 37 °C for 36 hours prior to harvest, spun down, washed with PBS, then diluted in PBS, seeded into the 96-well carbon electrode plates (MULTI-ARRAY high bind plate, MSD), and incubated overnight at 4 °C to allow the cells to adhere. Nonspecific binding sites were blocked by 2% BSA (w / v) in PBS for one hour at room temperature. To the plate-bound cells, anti-SARS-CoV-2 antibodies and a non-binding human IgGl control, diluted in PBS + 0.5% BSA at a range of concentrations from 0.0008nM to 50nM, and buffer with no antibody were added in duplicate and the plates incubated for one hour at room temperature with shaking. The plates were then washed with 1X PBS to remove the unbound antibodies using an AquaMax2000 plate washer (MDS Analytical Technologies). The plate-bound antibodies were detected with a SULFO-TAGTM-conjugated anti-human IgG antibody (Jackson Immunoresearch) for one hour at room temperature. After washes, the plates were developed with the Read Buffer (MSD) according to manufacturer's recommended procedure and the luminescent signals were recorded with a SECTOR Imager 600 (Meso Scale Development) instrument. The direct binding signals (in RLU) were captured for SARS-CoV-2-S expressing cells and a negative control cell line.

[0242] The ability of the anti-SARS-CoV-2 monoclonal antibodies to bind to SARS-CoV-2 Spike Protein expressing cells compared with binding to parental cells was assessed using an immunobinding assay. Binding to the immobilized cells on 96-well high bind plates (MSD) was performed with a series of antibody dilutions and the bound antibodies were detected using SULFO-TAGTM-conjugated anti-human IgG. The binding signals from electrochemiluminescence were recorded on a Sector Imager 600 (MSD). All antibodies displayed a concentration-dependent binding and the ratio of the binding on spike expressing cells to the parental cells were analyzed at the concentration of 5.5nM and 0.20nM.

[0243] The binding results of the anti-SARS-COV-2-S mAbs at the two concentrations to Spike protein expressing and parental Jurkat cells are summarized in Table 38. Of the 46 antibodies tested, 44 antibodies bound specifically to Jurkat / spike cells (Jurkat / Tet3G / hCD20 / Tet-On 3G Inducible SARS-CoV-2 Spike Protein High Sorted cells) with a ratio to the parental cells of 4 or higher at either concentration. At 0.2nM, the ratios of the binding signals on Jurkat / spike cells to the parental cells ranged from 4 to 36, and at 5 nM the ratio ranged from 4 to 63. Although the two antibodies (mAb 10998 and mAb 11002) displayed weak binding to Jurkat / spike cells with binding ratio to the parental cells less than 4, at 5 nM the binding signals were higher on Jurket / spike than on the parental cells. An irrelevant IgG1 isotype antibody showed minimal binding, as expected. Table 38: Specificity of anti-SARS-CoV-2-S antibodies binding to spike protein-expressing Jurkat cells vs parental cells by electrochemiluminescenceAntibody Binding Signal (RLU) Ratio Antibody Concentration 5.5nM 0.2nM 5.5nM 0.2nM mAb# Jurkat / Spike Parental Jurkat / Spike Parental Jurkat / Spike: Parental Jurkat / Spike:P arental mab1091390717457636516mAb 1091416245692626434mab1091518142172694286mAb 109203501597197080625mAb 10921374627243660147mAb 109223996322522610mAb1092325611031137462525mAb 109241418121336241214mAb109306731511752547mAb1093215256520629237mAb109338981716717359mAb1093476214669746515mAb10935157220951328819mAb1093699511656728921mAb109378679538830913mAb10938167816519530107mAb109393195292901119118mAb 1094065751291191316mAb 1094111963719233336mAb109549291103274687mAb1095575013427428610mAb109568011362144265mAb109578467621148114mAb 1096489637724202436mAb109656814913569142mAb 109669696524553155mAb 109679281211682686mAb 109692793124774352322mAb 109707435924657134mAb 1097183942263232012mAb 1097720319756047628mAb 109827371172112568mAb 109848899528226911mAb 10985527631796583mAb 10986105092341331110mAb 109876328347131815mAb 10988367832724147mAb1098977862778381320mAb 1099613991721852787mAb 1099812773931286532mAb 11000174570261222512mAb 11002241160303621mAb 11004203148748344322mAb 1100650521055104470515mAb 1100823823823750635mAb 11010387521403384IgGl isotype control9534622233IgGl isotype control5865214810IgGl isotype control64731186212

[0244] All references cited herein are incorporated by reference to the same extent as if each individual publication, database entry (e.g., Genbank sequences or GeneID entries), patent application, or patent, was specifically and individually indicated to be incorporated by reference. This statement of incorporation by reference is intended by Applicants to relate to each and every individual publication, database entry (e.g., Genbank sequences or GeneID entries), patent application, or patent identified even if such citation is not immediately adjacent to a dedicated statement of incorporation by reference. The inclusion of dedicated statements of incorporation by reference, if any, within the specification does not in any way weaken this general statement of incorporation by reference. Citation of the references herein is not intended as an admission that the reference is pertinent prior art, nor does it constitute any admission as to the contents or date of these publications or documents.ASPECTS OF THE INVENTION

[0245] 1. An isolated antibody or antigen-binding fragment thereof that binds a SARS-CoV-2 spike protein comprising the amino acid sequence set forth in SEQ ID NO: 832, wherein said isolated antibody or antigen-binding fragment comprises three heavy chain complementarity determining regions (CDRs) (HCDR1, HCDR2 and HCDR3) contained within a heavy chain variable region (HCVR) comprising the amino acid sequence set forth in SEQ ID NO: 202, and three light chain complementarity determining regions (CDRs) (LCDR1, LCDR2 and LCDR3) contained within a light chain variable region (LCVR) comprising the amino acid sequence set forth in SEQ ID NO: 210. 2. The isolated antibody or antigen-binding fragment thereof of aspect 1, wherein HCDR1 comprises the amino acid sequence set forth in SEQ ID NO: 204, HCDR2 comprises the amino acid sequence set forth in SEQ ID NO: 206, HCDR3 comprises the amino acid sequence set forth in SEQ ID NO: 208, LCDR1 comprises the amino acid sequence set forth in SEQ ID NO: 212, LCDR2 comprises the amino acid sequence set forth in SEQ ID NO: 55, and LCDR3 comprises the amino acid sequence set forth in SEQ ID NO: 214. 3. The isolated antibody or antigen-binding fragment thereof of aspect 2, comprising an HCVR that comprises the amino acid sequence set forth in SEQ ID NO: 202. 4. The isolated antibody or antigen-binding fragment thereof of aspect 2, comprising an LCVR that comprises the amino acid sequence set forth in SEQ ID NO: 210. 5. The isolated antibody or antigen-binding fragment thereof of aspect 2, comprising an HCVR that comprises the amino acid sequence set forth in SEQ ID NO: 202 and an LCVR that comprises the amino acid sequence set forth in SEQ ID NO: 210. 6. An isolated antibody that binds a SARS-CoV-2 spike protein comprising the amino acid sequence set forth in SEQ ID NO: 832, wherein said isolated antibody comprises an immunoglobulin constant region, three heavy chain complementarity determining regions (CDRs) (HCDR1, HCDR2 and HCDR3) contained within a heavy chain variable region (HCVR) comprising the amino acid sequence set forth in SEQ ID NO: 202, and three light chain complementarity determining regions (CDRs) (LCDR1, LCDR2 and LCDR3) contained within a light chain variable region (LCVR) comprising the amino acid sequence set forth in SEQ ID NO: 210. 7. The isolated antibody of aspect 6, wherein HCDR1 comprises the amino acid sequence set forth in SEQ ID NO: 204, HCDR2 comprises the amino acid sequence set forth in SEQ ID NO: 206, HCDR3 comprises the amino acid sequence set forth in SEQ ID NO: 208, LCDR1 comprises the amino acid sequence set forth in SEQ ID NO: 212, LCDR2 comprises the amino acid sequence set forth in SEQ ID NO: 55, and LCDR3 comprises the amino acid sequence set forth in SEQ ID NO: 214. 8. The isolated antibody of aspect 6, comprising an HCVR that comprises the amino acid sequence set forth in SEQ ID NO: 202 and an LCVR that comprises the amino acid sequence set forth in SEQ ID NO: 210. 9. The isolated antibody of aspect 6, wherein said isolated antibody comprises a heavy chain comprising the amino acid sequence set forth in SEQ ID NO: 216 and a light chain comprising the amino acid sequence set forth in SEQ ID NO: 218. 10. The isolated antibody of aspect 6, wherein said immunoglobulin constant region is an IgG1 constant region. 11. The isolated antibody of aspect 6 which is a recombinant antibody. 12. The isolated antibody of aspect 6 which is multispecific. 13. A pharmaceutical composition comprising the isolated antibody of aspect 6 and a pharmaceutically acceptable carrier or diluent. 14. The pharmaceutical composition of aspect 13, further comprising a second therapeutic agent. 15. The pharmaceutical composition of aspect 14, wherein the second therapeutic agent is selected from the group consisting of: a second antibody, or an antigen-binding fragment thereof, that binds a SARS-CoV-2 spike protein comprising the amino acid sequence set forth in SEQ ID NO: 832, an anti-inflammatory agent, an antimalarial agent, and an antibody or antigen-binding fragment thereof that binds TMPRSS2. 16. The pharmaceutical composition of aspect 14, wherein the second therapeutic agent is a second antibody, or an antigen-binding fragment thereof, that binds a SARS-CoV-2 spike protein comprising the amino acid sequence set forth in SEQ ID NO: 832. 17. The pharmaceutical composition of aspect 16, wherein the second antibody or antigen-binding fragment thereof comprises three heavy chain CDRs (HCDR1, HCDR2 and HCDR3) contained within an HCVR comprising the amino acid sequence set forth in SEQ ID NO: 640, and three light chain CDRs (LCDR1, LCDR2 and LCDR3) contained within an LCVR comprising the amino acid sequence set forth in SEQ ID NO: 646. 18. The pharmaceutical composition of aspect 17, wherein the second antibody or antigen-binding fragment thereof comprises: HCDR1, comprising the amino acid sequence set forth in SEQ ID NO: 642; HCDR2, comprising the amino acid sequence set forth in SEQ ID NO: 499; HCDR3, comprising the amino acid sequence set forth in SEQ ID NO: 644; LCDR1, comprising the amino acid sequence set forth in SEQ ID NO: 648; LCDR2, comprising the amino acid sequence set forth in SEQ ID NO: 650; and LCDR3, comprising the amino acid sequence set forth in SEQ ID NO: 652. 19. The pharmaceutical composition of aspect 18, wherein the second antibody or antigen-binding fragment thereof comprises an HCVR comprising the amino acid sequence set forth in SEQ ID NO: 640 and an LCVR comprising the amino acid sequence set forth in SEQ ID NO: 646. 20. The pharmaceutical composition of aspect 19, wherein the second antibody or antigen-binding fragment thereof comprises a heavy chain comprising the amino acid sequence set forth in SEQ ID NO: 654 and a light chain comprising the amino acid sequence set forth in SEQ ID NO: 656. 21. An isolated antibody or antigen-binding fragment thereof that binds a SARS-CoV-2 spike protein comprising the amino acid sequence set forth in SEQ ID NO: 832, wherein said isolated antibody or antigen-binding fragment comprises three heavy chain complementarity determining regions (CDRs) (HCDR1, HCDR2 and HCDR3) contained within a heavy chain variable region (HCVR) comprising the amino acid sequence set forth in SEQ ID NO: 640, and three light chain complementarity determining regions (CDRs) (LCDR1, LCDR2 and LCDR3) contained within a light chain variable region (LCVR) comprising the amino acid sequence set forth in SEQ ID NO: 646. 22. The isolated antibody or antigen-binding fragment thereof of aspect 21, wherein HCDR1 comprises the amino acid sequence set forth in SEQ ID NO: 642, HCDR2 comprises the amino acid sequence set forth in SEQ ID NO: 499, HCDR3 comprises the amino acid sequence set forth in SEQ ID NO: 644, LCDR1 comprises the amino acid sequence set forth in SEQ ID NO: 648, LCDR2 comprises the amino acid sequence set forth in SEQ ID NO: 650, and LCDR3 comprises the amino acid sequence set forth in SEQ ID NO: 652. 23. The isolated antibody or antigen-binding fragment thereof of aspect 21, comprising an HCVR that comprises an amino acid sequence set forth in SEQ ID NO: 640. 24. The isolated antibody or antigen-binding fragment thereof of aspect 21, comprising an LCVR that comprises an amino acid sequence set forth in SEQ ID NO: 646. 25. The isolated antibody or antigen-binding fragment thereof of aspect 21, comprising an HCVR that comprises an amino acid sequence set forth in SEQ ID NO: 640 and an LCVR that comprises an amino acid sequence set forth in SEQ ID NO: 646. 26. An isolated antibody that binds a SARS-CoV-2 spike protein comprising the amino acid sequence set forth in SEQ ID NO: 832, wherein said isolated antibody comprises an immunoglobulin constant region, three heavy chain complementarity determining regions (CDRs) (HCDR1, HCDR2 and HCDR3) contained within a heavy chain variable region (HCVR) comprising the amino acid sequence set forth in SEQ ID NO: 640, and three light chain complementarity determining regions (CDRs) (LCDR1, LCDR2 and LCDR3) contained within a light chain variable region (LCVR) comprising the amino acid sequence set forth in SEQ ID NO: 646. 27. The isolated antibody of aspect 26, wherein HCDR1 comprises the amino acid sequence set forth in SEQ ID NO: 642, HCDR2 comprises the amino acid sequence set forth in SEQ ID NO: 499, HCDR3 comprises the amino acid sequence set forth in SEQ ID NO: 644, LCDR1 comprises the amino acid sequence set forth in SEQ ID NO: 648, LCDR2 comprises the amino acid sequence set forth in SEQ ID NO: 650, and LCDR3 comprises the amino acid sequence set forth in SEQ ID NO: 652. 28. The isolated antibody of aspect 26, comprising an HCVR that comprises the amino acid sequence set forth in SEQ ID NO: 640 and an LCVR that comprises the amino acid sequence set forth in SEQ ID NO: 646. 29. The isolated antibody of aspect 26, wherein said isolated antibody comprises a heavy chain comprising the amino acid sequence set forth in SEQ ID NO: 654 and a light chain comprising the amino acid sequence set forth in SEQ ID NO: 656. 30. The isolated antibody of aspect 26, wherein said immunoglobulin constant region is an IgG1 constant region. 31. The isolated antibody of aspect 26 which is a recombinant antibody. 32. The isolated antibody of aspect 26 which is multispecific. 33. A pharmaceutical composition comprising the isolated antibody of aspect 26 and a pharmaceutically acceptable carrier or diluent. 34. The pharmaceutical composition of aspect 33, further comprising a second therapeutic agent. 35. The pharmaceutical composition of aspect 34, wherein the second therapeutic agent is selected from the group consisting of: a second antibody, or an antigen-binding fragment thereof, that binds a SARS-CoV-2 spike protein comprising the amino acid sequence set forth in SEQ ID NO: 832, an anti-inflammatory agent, an antimalarial agent, and an antibody or antigen-binding fragment thereof that binds TMPRSS2. 36. The pharmaceutical composition of aspect 34, wherein the second therapeutic agent is a second antibody, or an antigen-binding fragment thereof, that binds a SARS-CoV-2 spike protein comprising the amino acid sequence set forth in SEQ ID NO: 832. 37. The pharmaceutical composition of aspect 36, wherein the second antibody or antigen-binding fragment thereof comprises three heavy chain CDRs (HCDR1, HCDR2 and HCDR3) contained within an HCVR comprising the amino acid sequence set forth in SEQ ID NO: 202, and three light chain CDRs (LCDR1, LCDR2 and LCDR3) contained within an LCVR comprising the amino acid sequence set forth in SEQ ID NO: 210. 38. The pharmaceutical composition of aspect 37, wherein the second antibody or antigen-binding fragment thereof comprises: HCDR1, comprising the amino acid sequence set forth in SEQ ID NO: 204; HCDR2, comprising the amino acid sequence set forth in SEQ ID NO: 206; HCDR3, comprising the amino acid sequence set forth in SEQ ID NO: 208 LCDR1, comprising the amino acid sequence set forth in SEQ ID NO: 212; LCDR2, comprising the amino acid sequence set forth in SEQ ID NO: 55; and LCDR3, comprising the amino acid sequence set forth in SEQ ID NO: 214. 39. The pharmaceutical composition of aspect 38, wherein the second antibody or antigen-binding fragment thereof comprises an HCVR comprising the amino acid sequence set forth in SEQ ID NO: 202 and an LCVR comprising the amino acid sequence set forth in SEQ ID NO: 210. 40. The pharmaceutical composition of aspect 39, wherein the second antibody or antigen-binding fragment thereof comprises a heavy chain comprising the amino acid sequence set forth in SEQ ID NO: 216 and a light chain comprising the amino acid sequence set forth in SEQ ID NO: 218. 41. An isolated recombinant antibody or antigen-binding fragment thereof that specifically binds to a coronavirus spike protein (CoV-S), wherein the antibody has one or more of the following characteristics: (a) binds to CoV-S with an EC 50 of less than about 10 -9< M; (b) demonstrates an increase in survival in a coronavirus-infected animal after administration to said coronavirus-infected animal, as compared to a comparable coronavirus-infected animal without said administration; and / or (c) comprises three heavy chain complementarity determining regions (CDRs) (CDR-H1, CDR-H2, and CDR-H3) contained within a heavy chain variable region (HCVR) comprising an amino acid sequence having at least about 90% sequence identity to an HCVR of Table 1; and three light chain CDRs (CDR-L1, CDR-L2, and CDR-L3) contained within a light chain variable region (LCVR) comprising an amino acid sequence having at least about 90% sequence identity to an LCVR Table 1. 42. The antibody or antigen-binding fragment of aspect 41, comprising: (a) an immunoglobulin heavy chain variable region comprising the CDR-H1, CDR-H2, and CDR-H3 of an antibody of Table 1; and / or (b) an immunoglobulin light chain variable region comprising the CDR-L1, CDR-L2, and CDR-L3 of an antibody of Table 1. 43. The antibody or antigen-binding fragment of aspect 41 or 42, comprising: (a) a heavy chain immunoglobulin variable region comprising an amino acid sequence having at least 90% amino acid sequence identity to an HCVR sequence of Table 1; and / or (b) a light chain immunoglobulin variable region comprising an amino acid sequence having at least 90% amino acid sequence identity to an LCVR sequence of Table 1. 44. The antibody or antigen-binding fragment of any one of aspects 41-43, wherein said antibody or antigen-binding fragment comprises the CDR-H1, CDR-H2, CDR-H3, CDR-L1, CDR-L2, and CDR-L3 of a single antibody of Table 1. 45. The antibody or antigen-binding fragment of any one of aspects 41-44, comprising an immunoglobulin that comprises the HCVR and the LCVR of a single antibody of Table 1. 46. An antigen-binding protein that competes with the antibody or antigen-binding fragment of any one of aspects 41-45 for binding to CoV-S. 47. An antigen-binding protein that binds to the same epitope as, or to an overlapping epitope on, CoV-S as the antibody or antigen-binding fragment of any one of aspects 41-46. 48. The antibody or antigen-binding fragment of any one of aspects 41-47 which is multispecific. 49. The antibody or antigen-binding fragment of any one of aspects 41-48 which comprises one or more of the following properties: (a) inhibits growth of coronavirus; (b) binds to the surface of a coronavirus; (c) limits spread of coronavirus infection of cells in vitro; and (d) protects mice engineered to express the human ACE2 or TMPRSS2 protein from death and / or weight loss caused by coronavirus infection. 50. The antibody or antigen-binding fragment of any one of aspects 41-49, wherein said CoV-S is SARS-CoV-2-S. 51. A complex comprising an antibody or antigen-binding fragment of any one of aspects 41-50 bound to a CoV-S polypeptide. 52. The complex of aspect 51, wherein said CoV-S is SARS-CoV-2-S. 53. A method for making an antibody or antigen-binding fragment of any one of aspects 41-50, comprising: (a) introducing into a host cell one or more polynucleotides encoding said antibody or antigen-binding fragment; (b) culturing the host cell under conditions favorable to expression of the one or more polynucleotides; and (c) optionally, isolating the antibody or antigen-binding fragment from the host cell and / or a medium in which the host cell is grown. 54. The method of aspect 53, wherein the host cell is a Chinese hamster ovary cell. 55. An antibody or antigen-binding fragment which is a product of the method of aspect 53 or 54. 56. A polypeptide comprising: (a) CDR-H1, CDR-H2, and CDR-H3 of an HCVR domain of an antibody or antigen-binding fragment that comprises an HCVR amino acid sequence set forth in Table 1; or (b) CDR-L1, CDR-L2, and CDR-L3 of an LCVR domain of an immunoglobulin chain that comprises an LCVR amino acid sequence set forth in Table 1. 57. A polynucleotide encoding the polypeptide of aspect 56. 58. A vector comprising the polynucleotide of aspect 57. 59. A host cell comprising the antibody or antigen-binding fragment of any one of aspects 41-50, or polypeptide or polynucleotide or vector of any one of aspects 55-58. 60. A composition or kit comprising the antibody or antigen-binding fragment of any one of aspects 41-50 and 55 in association with a further therapeutic agent. 61. A pharmaceutical composition comprising the antigen-binding protein of any one of aspects 41-50 and 55 and pharmaceutically acceptable carrier and, optionally, a further therapeutic agent. 62. The composition or kit of aspect 60 or 61 in association with a further therapeutic agent which is an anti-viral drug or a vaccine. 63. The composition or kit of any one of aspects 60-62 wherein the further therapeutic agent is selected from the group consisting of: an anti-inflammatory agent, an antimalarial agent, an antibody or antigen-binding fragment thereof that specifically binds TMPRSS2, and an antibody or antigen-binding fragment thereof that specifically binds to CoV-S. 64. The composition or kit of aspect 63, wherein said antimalarial agent is chloroquine or hydroxychloroquine. 65. The composition or kit of aspect 63, wherein said anti-inflammatory agent is an antibody. 66. The composition or kit of aspect 65, wherein said antibody is sarilumab, tocilizumab, or gimsilumab. 67. The composition or kit of aspect 63, wherein the antibody or antigen-binding fragment comprises the HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3 sequences of a second antibody of Table 1. 68. A vessel or injection device comprising the antigen-binding protein or composition of any one of aspects 41-51, 55, and 60-63. 69. A method for treating or preventing infection with a coronavirus, in a subject in need thereof, comprising administering a therapeutically effective amount of antigen-binding protein of any one of aspects 41-50 and 55. 70. The method of aspect 68, wherein said coronavirus is selected from the group consisting of SARS-CoV-2, SARS-CoV, and MERS-CoV. 71. The method of aspect 69 or 70, wherein the subject is administered one or more further therapeutic agents. 72. The method of aspect 71, wherein the one or more further therapeutic agents is an anti-viral drug or a vaccine. 73. The method of aspect 71, wherein the one or more further therapeutic agents is selected from the group consisting of: an anti-inflammatory agent, an antimalarial agent, an antibody or antigen-binding fragment thereof that specifically binds TMPRSS2, and an antibody or antigen-binding fragment thereof that specifically binds to CoV-S. 74. The method of aspect 73, wherein said antimalarial agent is chloroquine or hydroxychloroquine. 75. The method of aspect 73, wherein said anti-inflammatory agent is an antibody. 76. The method of aspect 75, wherein said antibody is sarilumab, tocilizumab, or gimsilumab. 77. The method of aspect 73, wherein the antibody or antigen-binding fragment comprises the HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3 sequences of a second antibody of Table 1. 78. A method for administering an antibody or antigen-binding fragment of any one of aspects 41-50 and 55 into the body of a subject comprising injecting the antibody or antigen-binding fragment into the body of the subject. 79. The method of aspect 78, wherein the antibody or antigen-binding fragment is injected into the body of the subject subcutaneously, intravenously or intramuscularly. 80. The antibody or antigen-binding fragment, composition, kit, complex, polypeptide, polynucleotide, vector, cell, or method of any one of aspects 41-79, wherein said antibody or antigen-binding binding fragment comprises a VH3-66 or Vk1-33 variable domain sequence.

Claims

1. A pharmaceutical composition comprising: (i) an antibody or antigen-binding fragment thereof that binds a SARS-CoV-2 spike protein comprising the amino acid sequence set forth in SEQ ID NO: 832, wherein said antibody or antigen-binding fragment comprises three heavy chain complementarity determining regions (CDRs) (HCDR1, HCDR2 and HCDR3) contained within a heavy chain variable region (HCVR) comprising the amino acid sequence set forth in SEQ ID NO: 202, and three light chain complementarity determining regions (CDRs) (LCDR1, LCDR2 and LCDR3) contained within a light chain variable region (LCVR) comprising the amino acid sequence set forth in SEQ ID NO: 210; and (ii) an antibody or antigen-binding fragment thereof that binds a SARS-CoV-2 spike protein comprising the amino acid sequence set forth in SEQ ID NO: 832, wherein said antibody or antigen-binding fragment comprises three heavy chain complementarity determining regions (CDRs) (HCDR1, HCDR2 and HCDR3) contained within a heavy chain variable region (HCVR) comprising the amino acid sequence set forth in SEQ ID NO: 640, and three light chain complementarity determining regions (CDRs) (LCDR1, LCDR2 and LCDR3) contained within a light chain variable region (LCVR) comprising the amino acid sequence set forth in SEQ ID NO: 646; and (iii) a pharmaceutically acceptable carrier or diluent2. The pharmaceutical composition of claim 1, wherein the antibody or antigen-binding fragment thereof of (i) comprises: (a) a HCDR1 comprising the amino acid sequence set forth in SEQ ID NO: 204, a HCDR2 comprising the amino acid sequence set forth in SEQ ID NO: 206, a HCDR3 comprising the amino acid sequence set forth in SEQ ID NO: 208, a LCDR1 comprising the amino acid sequence set forth in SEQ ID NO: 212, a LCDR2 comprising the amino acid sequence set forth in SEQ ID NO: 55, and a LCDR3 comprising the amino acid sequence set forth in SEQ ID NO: 214; (b) an HCVR that comprises the amino acid sequence set forth in SEQ ID NO: 202; (c) an LCVR that comprises the amino acid sequence set forth in SEQ ID NO: 210; or (d) an HCVR that comprises the amino acid sequence set forth in SEQ ID NO: 202 and an LCVR that comprises the amino acid sequence set forth in SEQ ID NO: 210.

3. The pharmaceutical composition of claim 1 or 2, wherein the antibody or antigen-binding fragment thereof of (ii) comprises: (a) a HCDR1 comprising the amino acid sequence set forth in SEQ ID NO: 642, a HCDR2 comprising the amino acid sequence set forth in SEQ ID NO: 499, a HCDR3 comprising the amino acid sequence set forth in SEQ ID NO: 644, a LCDR1 comprising the amino acid sequence set forth in SEQ ID NO: 648, a LCDR2 comprising the amino acid sequence set forth in SEQ ID NO: 650, and LCDR3 comprising the amino acid sequence set forth in SEQ ID NO: 652; (b) an HCVR that comprises the amino acid sequence set forth in SEQ ID NO: 640; (c) an LCVR that comprises the amino acid sequence set forth in SEQ ID NO: 646; or (d) an HCVR that comprises the amino acid sequence set forth in SEQ ID NO: 640 and an LCVR that comprises the amino acid sequence set forth in SEQ ID NO: 646.

4. The pharmaceutical composition of claim 1, 2 or 3, wherein the antibody of (i) and / or the antibody of (ii) comprises: (a) an immunoglobulin constant region; (b) an IgG1 constant region; or (c) a human IgG1 constant region.

5. The pharmaceutical composition of any one of the preceding claims, wherein: (a) the antibody of (i) comprises a heavy chain comprising the amino acid sequence set forth in SEQ ID NO: 216 and a light chain comprising the amino acid sequence set forth in SEQ ID NO: 218; and / or (b) the antibody of (ii) comprises a heavy chain comprising the amino acid sequence set forth in SEQ ID NO: 654 and a light chain comprising the amino acid sequence set forth in SEQ ID NO: 656.

6. The pharmaceutical composition of any one of the preceding claims, wherein the antibody or antigen-binding fragment of (i) and / or the antibody or antigen-binding fragment of (ii): (a) is multispecific; and / or (b) is a recombinant antibody or antigen-binding fragment.

7. The pharmaceutical composition of any one of the preceding claims, comprising a further therapeutic agent, optionally wherein the further therapeutic agent is selected from the group consisting of: a further antibody, or an antigen-binding fragment thereof, that binds a SARS-CoV-2 spike protein comprising the amino acid sequence set forth in SEQ ID NO: 832, an anti-inflammatory agent, for example an anti-inflammatory antibody such as sarilumab, tocilizumab, or gimsilumab, an antimalarial agent such as chloroquine or hydroxychloroquine, an antibody or antigen-binding fragment thereof that binds TMPRSS2, and an anti-viral drug.

8. The pharmaceutical composition of any one of the preceding claims, for use in a method of treating or preventing a SARS-CoV-2 infection in a subject in need thereof, said method comprising administering the pharmaceutical composition to the subject.

9. The pharmaceutical composition for use according to claim 8, which is administered in combination with a further therapeutic agent, optionally wherein the further therapeutic agent is selected from the group consisting of: a further antibody, or an antigen-binding fragment thereof, that binds a SARS-CoV-2 spike protein comprising the amino acid sequence set forth in SEQ ID NO: 832, an anti-inflammatory agent, for example an anti-inflammatory antibody such as sarilumab, tocilizumab, or gimsilumab, an antimalarial agent such as chloroquine or hydroxychloroquine, an antibody or antigen-binding fragment thereof that binds TMPRSS2, an anti-viral drug and a vaccine.

10. A first antibody or antigen-binding fragment thereof that binds a SARS-CoV-2 spike protein comprising the amino acid sequence set forth in SEQ ID NO: 832 for use in a method of treating or preventing a SARS-CoV-2 infection in a subject in need thereof, in combination with a second antibody or antigen-binding fragment thereof that binds a SARS-CoV-2 spike protein comprising the amino acid sequence set forth in SEQ ID NO: 832, wherein: (i) the first antibody or antigen-binding fragement thereof comprises three heavy chain complementarity determining regions (CDRs) (HCDR1, HCDR2 and HCDR3) contained within a heavy chain variable region (HCVR) comprising the amino acid sequence set forth in SEQ ID NO: 202, and three light chain complementarity determining regions (CDRs) (LCDR1, LCDR2 and LCDR3) contained within a light chain variable region (LCVR) comprising the amino acid sequence set forth in SEQ ID NO: 210; and (ii) the second antibody or antigen-binding fragement thereof comprises three heavy chain complementarity determining regions (CDRs) (HCDR1, HCDR2 and HCDR3) contained within a heavy chain variable region (HCVR) comprising the amino acid sequence set forth in SEQ ID NO: 640, and three light chain complementarity determining regions (CDRs) (LCDR1, LCDR2 and LCDR3) contained within a light chain variable region (LCVR) comprising the amino acid sequence set forth in SEQ ID NO: 646.

11. The antibody or antigen-binding fragment thereof for use according to claim 10, wherein the first antibody or antigen-binding fragment thereof comprises: (a) a HCDR1 comprising the amino acid sequence set forth in SEQ ID NO: 204, a HCDR2 comprising the amino acid sequence set forth in SEQ ID NO: 206, a HCDR3 comprising the amino acid sequence set forth in SEQ ID NO: 208, a LCDR1 comprising the amino acid sequence set forth in SEQ ID NO: 212, a LCDR2 comprising the amino acid sequence set forth in SEQ ID NO: 55, and a LCDR3 comprising the amino acid sequence set forth in SEQ ID NO: 214; (b) an HCVR that comprises the amino acid sequence set forth in SEQ ID NO: 202; (c) an LCVR that comprises the amino acid sequence set forth in SEQ ID NO: 210; or (d) an HCVR that comprises the amino acid sequence set forth in SEQ ID NO: 202 and an LCVR that comprises the amino acid sequence set forth in SEQ ID NO: 210.

12. The antibody or antigen-binding fragment thereof for use according to claim 10 or 11, wherein the second antibody or antigen-binding fragment thereof comprises: (a) a HCDR1 comprising the amino acid sequence set forth in SEQ ID NO: 642, a HCDR2 comprising the amino acid sequence set forth in SEQ ID NO: 499, a HCDR3 comprising the amino acid sequence set forth in SEQ ID NO: 644, a LCDR1 comprising the amino acid sequence set forth in SEQ ID NO: 648, a LCDR2 comprising the amino acid sequence set forth in SEQ ID NO: 650, and LCDR3 comprising the amino acid sequence set forth in SEQ ID NO: 652; (b) an HCVR that comprises the amino acid sequence set forth in SEQ ID NO: 640; (c) an LCVR that comprises the amino acid sequence set forth in SEQ ID NO: 646; or (d) an HCVR that comprises the amino acid sequence set forth in SEQ ID NO: 640 and an LCVR that comprises the amino acid sequence set forth in SEQ ID NO: 646.

13. The antibody or antigen-binding fragment thereof for use according to claim 10, 11 or 12, wherein the first and / or second antibody comprises: (a) an immunoglobulin constant region; (b) an IgG1 constant region; or (c) a human IgG1 constant region.

14. The antibody or antigen-binding fragment thereof for use according to any one of claims 10-13, wherein: (a) the first antibody comprises a heavy chain comprising the amino acid sequence set forth in SEQ ID NO: 216 and a light chain comprising the amino acid sequence set forth in SEQ ID NO: 218; and / or (b) the second antibody comprises a heavy chain comprising the amino acid sequence set forth in SEQ ID NO: 654 and a light chain comprising the amino acid sequence set forth in SEQ ID NO: 65615. A first antibody or antigen-binding fragment thereof that binds a SARS-CoV-2 spike protein comprising the amino acid sequence set forth in SEQ ID NO: 832 for use in a method of treating or preventing a SARS-CoV-2 infection in a subject in need thereof in combination with a second antibody or antigen-binding fragment thereof that binds a SARS-CoV-2 spike protein comprising the amino acid sequence set forth in SEQ ID NO: 832, wherein: (i) the first antibody or antigen-binding fragement thereof comprises three heavy chain complementarity determining regions (CDRs) (HCDR1, HCDR2 and HCDR3) contained within a heavy chain variable region (HCVR) comprising the amino acid sequence set forth in SEQ ID NO: 640, and three light chain complementarity determining regions (CDRs) (LCDR1, LCDR2 and LCDR3) contained within a light chain variable region (LCVR) comprising the amino acid sequence set forth in SEQ ID NO: 646; and (ii) the second antibody or antigen-binding fragement thereof comprises three heavy chain complementarity determining regions (CDRs) (HCDR1, HCDR2 and HCDR3) contained within a heavy chain variable region (HCVR) comprising the amino acid sequence set forth in SEQ ID NO: 202, and three light chain complementarity determining regions (CDRs) (LCDR1, LCDR2 and LCDR3) contained within a light chain variable region (LCVR) comprising the amino acid sequence set forth in SEQ ID NO: 210.

16. The antibody or antigen-binding fragment thereof for use according to claim 15, wherein the first antibody or antigen-binding fragment thereof comprises: (a) a HCDR1 comprising the amino acid sequence set forth in SEQ ID NO: 642, a HCDR2 comprising the amino acid sequence set forth in SEQ ID NO: 499, a HCDR3 comprising the amino acid sequence set forth in SEQ ID NO: 644, a LCDR1 comprising the amino acid sequence set forth in SEQ ID NO: 648, a LCDR2 comprising the amino acid sequence set forth in SEQ ID NO: 650, and LCDR3 comprising the amino acid sequence set forth in SEQ ID NO: 652; (b) an HCVR that comprises the amino acid sequence set forth in SEQ ID NO: 640; (c) an LCVR that comprises the amino acid sequence set forth in SEQ ID NO: 646; or (d) an HCVR that comprises the amino acid sequence set forth in SEQ ID NO: 640 and an LCVR that comprises the amino acid sequence set forth in SEQ ID NO: 646.

17. The antibody or antigen-binding fragment thereof for use according to claim 15 or 16, wherein the second antibody or antigen-binding fragment thereof comprises: (a) a HCDR1 comprising the amino acid sequence set forth in SEQ ID NO: 204, a HCDR2 comprising the amino acid sequence set forth in SEQ ID NO: 206, a HCDR3 comprising the amino acid sequence set forth in SEQ ID NO: 208, a LCDR1 comprising the amino acid sequence set forth in SEQ ID NO: 212, a LCDR2 comprising the amino acid sequence set forth in SEQ ID NO: 55, and a LCDR3 comprising the amino acid sequence set forth in SEQ ID NO: 214; (b) an HCVR that comprises the amino acid sequence set forth in SEQ ID NO: 202; (c) an LCVR that comprises the amino acid sequence set forth in SEQ ID NO: 210; or (d) an HCVR that comprises the amino acid sequence set forth in SEQ ID NO: 202 and an LCVR that comprises the amino acid sequence set forth in SEQ ID NO: 210.

18. The antibody or antigen-binding fragment thereof for use according to claim 15, 16 or 17, wherein the first and / or second antibody comprises: (a) an immunoglobulin constant region; (b) an IgG1 constant region; or (c) a human IgG1 constant region.

19. The antibody or antigen-binding fragment thereof for use according to any one of claims 15-18, wherein: (a) the first antibody comprises a heavy chain comprising the amino acid sequence set forth in SEQ ID NO: 654 and a light chain comprising the amino acid sequence set forth in SEQ ID NO: 656; and / or (a) the second antibody comprises a heavy chain comprising the amino acid sequence set forth in SEQ ID NO: 216 and a light chain comprising the amino acid sequence set forth in SEQ ID NO: 218.

20. The antibody or antigen-binding fragment thereof for use according to any one of claims 10-19, wherein the first and / or second antibody is a recombinant antibody and / or a multispecific antibody.

21. The antibody or antigen-binding fragment thereof for use according to any one of claims 10-20, which is administered in combination with a further therapeutic agent, optionally wherein the further therapeutic agent is selected from the group consisting of: a further antibody, or an antigen-binding fragment thereof, that binds a SARS-CoV-2 spike protein comprising the amino acid sequence set forth in SEQ ID NO: 832, an anti-inflammatory agent, for example an anti-inflammatory antibody such as sarilumab, tocilizumab, or gimsilumab, an antimalarial agent such as chloroquine or hydroxychloroquine, an antibody or antigen-binding fragment thereof that binds TMPRSS2, an anti-viral drug and a vaccine.

22. The antibody or antigen-binding fragment thereof for use according to any one of claims 10-21, wherein the first and second antibody are administered concurrently or sequentially.

23. An isolated antibody or antigen-binding fragment thereof that binds a SARS-CoV-2 spike protein comprising the amino acid sequence set forth in SEQ ID NO: 832, wherein said isolated antibody or antigen-binding fragment comprises three heavy chain complementarity determining regions (CDRs) (HCDR1, HCDR2 and HCDR3) contained within a heavy chain variable region (HCVR) comprising the amino acid sequence set forth in SEQ ID NO: 202, and three light chain complementarity determining regions (CDRs) (LCDR1, LCDR2 and LCDR3) contained within a light chain variable region (LCVR) comprising the amino acid sequence set forth in SEQ ID NO: 210.

24. The isolated antibody or antigen-binding fragment thereof of claim 23, wherein: (a) HCDR1 comprises the amino acid sequence set forth in SEQ ID NO: 204, HCDR2 comprises the amino acid sequence set forth in SEQ ID NO: 206, HCDR3 comprises the amino acid sequence set forth in SEQ ID NO: 208, LCDR1 comprises the amino acid sequence set forth in SEQ ID NO: 212, LCDR2 comprises the amino acid sequence set forth in SEQ ID NO: 55, and LCDR3 comprises the amino acid sequence set forth in SEQ ID NO: 214; (b) the antibody comprises an HCVR that comprises the amino acid sequence set forth in SEQ ID NO: 202; (c) the antibody comprises an LCVR that comprises the amino acid sequence set forth in SEQ ID NO: 210; or (d) the antibody comprises an HCVR that comprises the amino acid sequence set forth in SEQ ID NO: 202 and an LCVR that comprises the amino acid sequence set forth in SEQ ID NO: 210.

25. The antibody of claim 23 or 24, which comprises: (a) an immunoglobulin constant region; (b) an IgG1 constant region; (c) a human IgG1 constant region; or (d) a heavy chain comprising the amino acid sequence set forth in SEQ ID NO: 216 and / or a light chain comprising the amino acid sequence set forth in SEQ ID NO: 218.

26. An isolated antibody or antigen-binding fragment thereof that binds a SARS-CoV-2 spike protein comprising the amino acid sequence set forth in SEQ ID NO: 832, wherein said isolated antibody or antigen-binding fragment comprises three heavy chain complementarity determining regions (CDRs) (HCDR1, HCDR2 and HCDR3) contained within a heavy chain variable region (HCVR) comprising the amino acid sequence set forth in SEQ ID NO: 640, and three light chain complementarity determining regions (CDRs) (LCDR1, LCDR2 and LCDR3) contained within a light chain variable region (LCVR) comprising the amino acid sequence set forth in SEQ ID NO: 646.

27. The antibody or antigen-binding fragment thereof of claim 26, wherein: (a) HCDR1 comprises the amino acid sequence set forth in SEQ ID NO: 642, HCDR2 comprises the amino acid sequence set forth in SEQ ID NO: 499, HCDR3 comprises the amino acid sequence set forth in SEQ ID NO: 644, LCDR1 comprises the amino acid sequence set forth in SEQ ID NO: 648, LCDR2 comprises the amino acid sequence set forth in SEQ ID NO: 650, and LCDR3 comprises the amino acid sequence set forth in SEQ ID NO: 652; (b) the antibody comprises an HCVR comprising an amino acid sequence set forth in SEQ ID NO: 640; (c) the antibody comprises an LCVR comprising an amino acid sequence set forth in SEQ ID NO: 646; or (d) the antibody comprises an HCVR comprising an amino acid sequence set forth in SEQ ID NO: 640 and an LCVR comprising an amino acid sequence set forth in SEQ ID NO: 646.

28. The antibody of claim 26 or 27, which comprises: (a) an immunoglobulin constant region; (b) an IgG1 constant region; (c) a human IgG1 constant region; or (d) a heavy chain comprising the amino acid sequence set forth in SEQ ID NO: 654 and / or a light chain comprising the amino acid sequence set forth in SEQ ID NO: 656.