CA3194162A1 - Antibodies against sars-cov-2 - Google Patents

Abstract

The instant disclosure provides antibodies and antigen-binding fragments thereof that can bind to a SARS-CoV-2 antigen and, in certain embodiments, are capable of neutralizing a SARS-CoV-2 infection. In certain embodiments, an antibody or antigen-binding fragment is capable of binding to a SARS-CoV-2 spike protein in the N-terminal domain (NTD). Also provided are polynucleotides that encode an antibody or antigen-binding fragment, vectors that comprise a polynucleotide, host cells that express an antibody or antigen-binding fragment, pharmaceutical compositions, and methods for treating or diagnosing a SARS-CoV-2 infection.

Description

STATEMENT REGARDING SEQUENCE LISTING
The Sequence Listing associated with this application is provided in text format in lieu of a paper copy, and is hereby incorporated by reference into the specification.
The name of the text file containing the Sequence Listing is 930585 418W0 SEQUENCE LISTING.txt. The text file is 330 KB, was created on September 26, 2021, and is being submitted electronically via EFS-Web.
BACKGROUND
A novel betacoronavirus emerged in Wuhan, China, in late 2019. As of September 22, 2021, approximately 230 million cases of infection by this virus (termed, among other names, SARS-CoV-2 and Wuhan coronavirus), were confirmed worldwide, and had resulted in over 4.7 million deaths. Therapies for preventing, treating, or diagnosing SARS-CoV-2 infection are needed.
BRIEF DESCRIPTION OF THE DRAWINGS
Figures 1A-1C show binding of certain antibodies of the present disclosure to SARS-CoV-2 Domain A. Human monoclonal antibodies isolated from donors were expressed recombinantly and were tested for binding by EL1SA. The boxes on the right side of each figure indicate the calculated EC50 value (ng/mL) for the indicated antibody.
Figure 2A shows frequency of antibodies specific for SARS CoV-2 RBD, Spike protein (non-RBD), or Domain A from sera of three donors. Figure 2B shows percent identity to IGHV germline sequence of certain antibodies. Figure 2C shows percent identity to IGLV germline sequence of certain antibodies.
Figures 3A-3E show additional functional characterization of certain antibodies. (3A) Neutralization on Domain A by ELISA. (3B, 3C) Neutralization of SARS CoV-2 pseudoparticles. (3D) Maximum neutralization plateau. (3E) Neutralization EC50.

Figures 4A-4B show neutralization by antibodies 418_i (4A) and 418 5 (4B) against authentic SARS-CoV-2 virus. Other antibodies were used as comparators.
Figures 5A-5C show results from epitope binning studies using biolayer interferometry (BLI). In (5B), RBD-specific antibodies (top) were used as comparators.
Figures 6A and 6B relate to binding of certain antibodies of the present disclosure to transiently transfected ExpiCHO cells expressing various sarbecoviruses (Clade 2, Clade 1, or Clade 3), embecoviruses, merbecovirus, or mock. Antibody S2X259 was included as a comparator in Figure 6A (flow cytometry study).
Figures 7A-7D show neutralization of infection by certain antibodies expressed as recombinant Fab or full IgG. Figure 7E shows results from binding on binding assays using ACE2 (left) or spike (bottom, right). In the bottom panel, data from comparator antibodies S309, S2E12, and S2M11 is also shown Figures 8A and 8B show effector functions of certain antibodies of the present disclosure, along with a comparator antibody. Figure 8C shows antibody-mediated shedding of CoV-2 Si protein from infected cells. In Figure 8C, antibodies S309, S2E12, and S2M11v1 were used as comparators Figures 9A-9C show data from neutralization experiments testing antibody combinations with antibody 418 4 and another antibody (S309, S2E12, or S2M11) against MLV pscudotypc with SARS-CoV2.
Figures 10A and 10B show binding of certain antibodies of the present disclosure to SARS-CoV-2 Domain A, as measured by ELISA.
Figure 11 shows data from neutralization experiments using certain antibodies of the present disclosure and SARS-CoV-2 pseudoparticles Figure 12 shows binding of certain antibodies of the present disclosure to to SARS-CoV-2 Domain A, as measured by ELISA.
Figure 13 shows neutralization of SARS CoV-2 pseudoparticles by certain antibodies of the present disclosure.
Figures 14A-14C show binding of certain antibodies of the present disclosure to SARS-CoV-2 Domain A, as measured by ELISA.
Figures 15A and 15B shows data from neutralization experiments using certain antibodies of the present disclosure and SARS CoV-2 pseudoparticles.

2 Figures 16A-16C show binding of certain antibodies of the present disclosure to SARS-CoV-2 spike protein and to SARS-CoV-2 Domain A, as measured by ELISA.
Figures 17A-17C show kinetics of binding by three antibodies of the present disclosure to SARS-CoV-2 spike protein. Calculated Kon, Koff, and KD values are shown in the boxes on the right side of each figure.
Figure 18 shows data from neutralization experiments using certain antibodies of the present disclosure and SARS-CoV-2 virus pseudoparticles.
Figures 19A-19F show kinetics of binding by certain antibodies of the present disclosure to SARS-CoV-2 Domain A, as measured by BLI.
Figure 20 shows the frequency of antibodies recognizing the SARS-CoV-2 N-terminal domain (NTD, also referred to herein as Domain A), RBD, or other S
regions for monoclonal antibodies cloned from IgG+ memory B cells of three donors.
Figure 21 shows binding and neutralization data for certain NTD-specific antibodies. The left panel shows binding of 41 anti-NTD antibodies to immobilized SARS-CoV-2 S protein, NTD, or RBD, as determined by ELISA. The center panel shows neutralization of infection by 1V1LV pseudotyped with SARS-CoV-2 S
protein for each of 15 NTD-specific antibodies. The right panel shows maximal neutralization plateau for the same 15 NTD-specific antibodies.
Figure 22 shows neutralization of authentic SARS-CoV-2-Nluc infection for certain antibodies assessed after 6 hours, using an MOT of 0.1. Error bars indicate standard deviation of triplicates.
Figure 23 shows neutralization of authentic SARS-CoV-2-Nluc infection for certain antibodies assessed after 24 hours, using an MOT of 0.01. Error bars indicate standard deviation of triplicates.
Figures 24A-24D show binding kinetic analysis of SARS-CoV-2 NTD to immobilized antibodies, as measured using BLI.
Figure 25 shows V gene usage for heavy chain (left panel) and light chain (right panel) of certain NTD-specific antibodies Figure 26 shows further characterization of certain NTD-specific antibodies.
The left panel shows nucleotide sequence identity of the antibodies relative to their

3 respective V germline genes. The right panel shows the HCDR3 amino acid length for the antibodies.
Figure 27 shows results from a cell-to-cell fusion inhibition assay using Vero E6 cells expressing SARS-CoV-2 S protein incubated with varying concentrations of each of four NTD-specific antibodies or RBD-specific antibody S2M11.
Figures 28A-28F show binding of each of 41 NTD-specific antibodies to immobilized SARS-CoV-2 S protein ("Spike"), NTD ("Dom A"), or RBD as measured by ELISA.
Figures 29A-29F show neutralization of infection by MLV pseudotyped with SARS-CoV-2 S protein for each of 41 NTD-specific antibodies.
Figure 30 shows six antigenic sites (i) ¨ (vi) identified by epitope binning of 41 NTD-specific antibodies based on competition binding assays using biolayer interferometry (BLI).
Figures 31A-31I show the results of competition binding assays for 41 NTD-specific antibodies using BLI. Results for antibodies identified as binding Site i are shown in Figures 31A-31C. Results for antibodies identified as binding Site ii are shown in Figure 31D. Results for antibodies identified as binding Site iii are shown in Figures 31E-31H. Results for antibodies identified as binding Site iv, Site v, and Site vi arc shown in Figure 3H.
Figure 32 shows competition by each of four NTD-specific antibodies and RBD-specific antibody S2E12 with ACE2 for binding to SARS-CoV-2 S protein as measured by BLI. ACE2 was immobilized at the surface of the biosensors before incubation with S protein alone or in complex with antibody. The vertical dashed line in each graph indicates the start of the association of S/antibody complex or free S with solid-phase ACE2.
Figure 33 shows neutralization of authentic SARS-CoV-2-Nluc infection by IgG or Fab of each of four NTD-specific antibodies and of comparator antibodies S309 and S2M11 Symbols are means SD of triplicates Dotted lines in each graph indicate IC50 and IC90 values.
Figure 34 shows results of SPR analysis of antibodies binding to SARS-CoV-2 S protein ectodomain trimer. The gray dashed line in each graph indicates a fit to a 1:1

4 binding model. The equilibrium dissociation constant (KD) or apparent equilibrium dissociation constant (KD, app) are indicated on each graph. White and gray stripes on each graph indicate association and dissociation phases, respectively.
Figure 35 shows activation of FciRlla H131 (left panel) and FcyRIIIa V158 (right panel) induced by the NTD-specific antibodies indicated and by RBD-specific antibody S309.
Figure 36 shows a matrix assessing synergistic activity of S2X333 and S309 antibody cocktails for in vitro neutralization of authentic SARS-CoV-2-Nluc infection.
Data are from one representative example performed in triplicate.
Figures 37A-37D show data from Syrian hamsters injected with the indicated amount of S2X333 antibody 48 hours before intra-nasal challenge with SARS-CoV-2.
Figure 37A shows quantification of viral RNA in the lungs four days post-infection Figure 37B shows quantification of replicating virus in lung homogenates harvested four days post infection using a TCID50 assay. Figures 37C and 37D show viral RNA
load (Figure 37C) and replicating virus titers (Figure 37D) in the lung four days post infection plotted as a function of serum antibody concentration before infection (day 0) Figure 38 shows infection of HEK293T cells transfected to over-express ACE2 or one of a panel of selected lectins and receptor candidates by VSV-SARS-CoV-pscudovirus.
Figure 39 shows micrographs of stable TIEK293T cell lines overexpressing DC-SIGN, L-SIGN, SIGLEC1, or ACE2 infected with authentic SARS-CoV-2 (MOI of 0.1), then fixed and immunostained for 24 hours for SARS-CoV-2 nucleoprotein (red) Figure 40 shows quantification of luciferase levels in stable HEK293T cell lines overexpressing DC-SIGN, L-SIGN, SIGLEC1, or ACE2, as measured 24 hours after infection with SARS-CoV-2-Nluc.
Figure 41 shows quantification of luciferase levels in stable HEK293T cell lines overexpressing DC-SIGN, L-SIGN, SIGLEC1, or ACE2 after incubation with different concentrations of anti-SIGLEC1 monoclonal antibody (clone 7-239) and infection with SARS-CoV-2-Nluc.
Figure 42 shows infection of cells transiently transduced to overexpress DC-SIGN, L-SIGN, SIGLEC1, or ACE2 by VSV-SARS-CoV-2 pseudovirus. Results for

5 HEK293T cells (left panel), HeLa cells (center panel), and MRCS cells (right panel) are shown.
Figure 43 shows infection of stable HEK293T cell lines overexpressing DC-SIGN, L-SIGN, SIGLEC1, or ACE2 after treatment with ACE2 siRNA followed by infection with VSV-SARS-CoV-2 pseudovirus.
Figure 44 shows infection of stable HEK293T cell lines overexpressing DC-SIGN, L-SIGN, SIGLEC1, or ACE2 after treatment with different concentrations of anti-ACE2 antibody (polyclonal serum) followed by infection with VSV-SARS-CoV-pseudovirus.
Figure 45 shows the distribution and expression of ACE2, DC-SIGN (CD209), L-SIGN (CLEC4M), and SIGLEC1 in the human lung cell atlas.
Figure 46 shows analysis of major cell types with detectable SARS-CoV-2 genome in bronchoalveolar lavage fluid or sputum of severe COVID-19 patients.
The single cell gene expression profiles are shown as a t-SNE (t-distributed stochastic neighbor embedding) plot, sized by viral load.
Figure 47 shows analysis of major cell types with detectable SARS-CoV-2 genome in bronchoalveolar lavage fluid or sputum of severe COVID-19 patients.
The cumulative fraction of cells (y-axis) with detected viral RNA per cell up to the corresponding logCPM (log(counts per million); x-axis) is shown for each of the indicated cell types.
Figure 48 shows a heatmap matrix of counts for cells with detected transcripts for the receptor genes shown on the x-axis and SARS-CoV-2+ cell types on the y-axis.
Total n=3,085 cells from eight subjects. See Ren, X. et al. COVID-19 immune features revealed by a large-scale single cell transcriptome atlas. Cell, doi:10.1016/j.ce11.2021.01.053 (2021).
Figure 49 shows the correlation of receptor transcript counts (y-axis of each plot) with SARS-CoV-2 RNA counts (x-axis of each plot) in macrophages and in secretory cells Correlation is based on counts before log transformation from Ren et al Figure 50 shows the results of trans-infection with VSV-SARS-CoV-2. A
schematic of the trans-infection process is shown in the left panel. HeLa cells transduced with DC-SIGN, L-SIGN, or SIGLEC1 were incubated with VSV-SARS-

6 CoV-2, extensively washed, and co-cultured with Vero-E6-TMPRSS2 susceptible target cells. Results in the presence or absence of target cells are shown in the right panel.
Figure 51 shows the results of trans-infection, where VSV-SARS-CoV-2 viral adsorption was performed in the presence or absence of an anti-SIGLEC1 blocking antibody.
Figure 52 shows neutralization of SARS-CoV-2 infection of Vero-E6 cells by antibodies S309, S2E12, and S2X33.
Figure 53 shows neutralization of SARS-CoV-2 infection of Vero-E6-T1VIPRSS2 cells by antibodies S309, S2E12, and S2X33.
Figure 54 shows quantification of binding of purified, fluorescently-labeled SARS-CoV-2 spike protein or RED to the indicated cell lines, as measured by flow cytometry. "A" indicates cell line overexpressing ACE2; "T" indicates cell line overexpressing TMPRSS2.
Figure 55 shows quantification of cellular ACE2 and TIVfPRSS2 transcripts in the indicated cell lines, as measured by RT-qPCR "A" indicates cell line overexpressing ACE2, "T" indicates cell line overexpressing TMPRSS2.
Figure 56 shows neutralization of SARS-CoV-2-Nluc infection by antibodies S309, S2E12, or S2X333. Each of the seven cell lines indicated was tested.
Luciferase signal was quantified 24 hours post infection.
Figure 57 shows neutralization of VSV-SARS-CoV-2 pseudovirus infection by antibodies S309, S2E12, or S2X333. Each of the seven cell lines indicated was tested.
Luciferase signal was quantified 24 hours post infection.
Figure 58 shows S2E12-induced uni-directional fusion (also referred to as trans-fusion) of S-positive CHO-S cells with fluorescently labelled S-negative CHO
cells in the absence of ACE2. Nuclei were stained with Hoechst dye; cytoplasm was stained with CellTracker Green.
Figure 59 shows neutralization of infection of a stable HEK293T cell line overexpressing ACE2 by authentic SARS-CoV-2 pre-incubated with the indicated monoclonal antibodies. Infection was measured by immunostaining at 24 hours for the SARS-CoV-2 nucleoprotein.

7 Figure 60 shows neutralization of infection of a stable HEK293T cell line overexpressing SIGLEC1 by authentic SARS-CoV-2 pre-incubated with the indicated monoclonal antibodies. Infection was measured by immunostaining at 24 hours for the SARS-CoV-2 nucleoprotein.
Figure 61 shows neutralization of infection of a stable HEK293T cell line overexpressing DC-SIGN by authentic SARS-CoV-2 pre-incubated with the indicated monoclonal antibodies. Infection was measured by immunostaining at 24 hours for the SARS-CoV-2 nucleoprotein.
Figure 62 shows neutralization of infection of a stable HEK293T cell line overexpressing L-SIGN by authentic SARS-CoV-2 pre-incubated with the indicated monoclonal antibodies. Infection was measured by immunostaining at 24 hours for the SARS-CoV-2 nucleoprotein.
Figure 63 shows analysis of binding of antibodies targeting DC/L-SIGN, DC-SIGN, SIGLEC1, or ACE2 on HEK293T cells stably over-expressing the respective attachment receptor, as measured by flow cytometry.
Figure 64 shows analysis of binding of antibodies targeting DC/L-SIGN, DC-SIGN, SIGLEC1, or ACE2 on HEK293T cells stably over-expressing the respective attachment receptor, as measured by immunofluorescence.
Figure 65 shows infection of HEK293T cells stably over-expressing the indicated attachment receptor by VSV-SARS-CoV-2 pseudotyped with wild type spike protein (grey bars), or VSV-SARS-CoV-2 pseudotyped with spike protein bearing the mutations of the B1.1.7 lineage (red bars). Luminescence was analyzed one day post infection.
Figure 66 shows neutralization of SARS-CoV-2 infection of Vero-E6 or Vero-E6-TMPRS S2 cells by 10 mg/m1 of S309, S2E12, and S2X333. Cells were infected with SARS-CoV-2 (isolate USA-WA1/2020) at MOI 0.01 in the presence of the indicated antibodies. Cells were fixed 24h post infection and viral nucleocapsid protein was immunostained Figure 67 shows quantification of binding of purified, fluorescently labelled SARS-CoV-2 spike protein (left panels) or RBD (right panels) to the indicated cell lines, as measured by flow cytometry.

8

9 Figure 68 shows quantification of binding of purified, fluorescently labelled SARS-CoV-2 spike protein (left panels) or RBD (right panels) to the indicated cell lines, as measured by flow cytometry.
Figure 69 shows an analysis of the correlation between ACE2 transcript levels (x-axis) and maximum antibody-related neutralization of infection (y-axis) in SARS-CoV-2-susceptible cell lines for antibody S309 (left panel) and antibody S2X333 (right panel).
Figure 70 shows binding of immunocomplexes to hamster splenocytes. Alexa-488 fluorescent immunocomplexes (IC) were titrated (0-200 nM range) and incubated with total naive hamster splenocytes. Binding was revealed with a cytometer upon exclusion of dead/apoptotic cells and physical gating on bona fide monocyte population. Left panel shows the fluorescent intensity associated to hamster cells of IC
made with either hamster or human Fe antibodies. A single replicate of two is shown.
Right panel shows the relative Alexa-488 mean fluorescent intensity of the replicates measured on the entire monocyte population.
Figure 71 shows analysis of the role of host effector function in SARS-CoV-2 challenge. Syrian hamsters were injected with the indicated amount (mg/kg) of hamster IgG2a S309, either wt or Fe silenced (S309-N297A). Top panel shows quantification of viral RNA in the lung 4 days post infection. Center panel shows quantification of replicating virus in the lung 4 days post infection. Bottom panel shows histopathological score in the lung 4 days post infection. Control animals (white symbols) were injected with 4 mg/kg unrelated control isotype antibody. * p<
0.05, **
p< 0.01, *** p< 0.001, **** p< 0.0001 vs control animals, using Mann-Whitney test.
Figure 72 shows neutralization of SARS-CoV-2 infection of HEK293T cells stably expressing ACE2 (top panel) or DC-SIGN (bottom panel) in the presence of the indicated antibodies. Cells were infected at MOI of 0.02. Cells were fixed 24h post infection, viral nucleocapsid protein was immunostained and positive cells were quantified Figure 73 shows neutralization of SARS-CoV-2 infection of HEK293T cells stably expressing SIGLEC I (top panel) or L-SIGN (bottom panel) in the presence of the indicated antibodies. Cells were infected at MOI of 0.02. Cells were fixed 24h post infection, viral nucleocapsid protein was immunostained and positive cells were quantified.
DETAILED DESCRIPTION
Provided herein are antibodies and antigen-binding fragments that bind to SARS-CoV-2 coronavirus (e.g., a SARS-CoV-2 Domain A, in a SARS-CoV-2 virion and/or expressed on the surface of a cell infected by the SARS-CoV-2 coronavirus). In certain embodiments, presently disclosed antibodies and antigen-binding fragments can neutralize a SARS-CoV-2 infection in an in vitro model of infection and/or in a human subject. Also provided are polynucleotides that encode the antibodies and antigen-binding fragments, vectors, host cells, and related compositions, as well as methods of using the antibodies, nucleic acids, vectors, host cells, and related compositions to treat (e.g., reduce, delay, eliminate, or prevent) a SARS-CoV-2 infection in a subject and/or in the manufacture of a medicament for treating a SARS-CoV-2 infection in a subject.
Prior to setting forth this disclosure in more detail, it may be helpful to an understanding thereof to provide definitions of certain terms to be used herein.
Additional definitions are set forth throughout this disclosure.
As used herein, "SARS-CoV-2", also referred to herein as "Wuhan seafood market phenomia virus", or "Wuhan coronavirus" or "Wuhan CoV", or "novel CoV", or "nCoV", or "2019 nCoV", or "Wuhan nCoV" is a betacoronavirus believed to be of lineage B (sarbecovirus). SARS-CoV-2 was first identified in Wuhan, Hubei province, China, in late 2019 and spread within China and to other parts of the world by early 2020. Symptoms of SARS-CoV-2 infection include fever, dry cough, and dyspnea.
The genomic sequence of SARS-CoV-2 isolate Wuhan-Hu-1 is provided in SEQ
ID NO.:1 (see also GenBank MN908947.3, January 23, 2020), and the amino acid translation of the genome is provided in SEQ ID NO. :2 (see also GenBank QHD43416.1, January 23, 2020). Like other coronavinises (e.g., SARS- CoV-1), SARS-CoV-2 comprises a "spike" or surface ("S") type I transmembrane glycoprotein containing a receptor binding domain (RBD). RBD is believed to mediate entry of the lineage B SARS coronavirus to respiratory epithelial cells by binding to the cell surface receptor angiotensin-converting enzyme 2 (ACE2). In particular, a receptor binding motif (RBM) in the virus RBD is believed to interact with ACE2. SARS CoV-2 S
protein also includes, N-terminal to the RBD and C-terminal to the S protein signal peptide, domain A (also referred-to as the N-terminal Domain or "NTD").
Antibodies of the present disclosure are specific for domain A.
The amino acid sequence of the Wuhan-Hu-1 surface glycoprotein is provided in SEQ ID NO.:3. The amino acid sequence of SARS-CoV-2 RBD is provided in SEQ
ID NO.:4. SARS-CoV-2 S protein has approximately 73% amino acid sequence identity with SARS-CoV-1. The amino acid sequence of SARS-CoV-2 RBM is provided in SEQ ID NO.:5. SARS-CoV-2 RBD has approximately 75% to 77% amino acid sequence similarity to SARS¨CoV-1 RBD, and SARS-CoV-2 RBM has approximately 50% amino acid sequence similarity to SARS-CoV-1 RBM.
Unless otherwise indicated herein, SARS-CoV-2 Wuhan-Hu-1 refers to a virus comprising the amino acid sequence set forth in any one or more of SEQ ID
NOs.:2, or 3, optionally with the genomic sequence set forth in SEQ ID NO.:1.
There have been a number of emerging SARS-CoV-2 variants. Some SARS-CoV-2 variants contain an N439K mutation, which has enhanced binding affinity to the human ACE2 receptor (Thomson, E.C., et al., The circulating SARS-CoV-2 spike variant N439K maintains fitness while evading antibody-mediated immunity.
bioRxiv, 2020). Some SARS-CoV-2 variants contain an N501Y mutation, which is associated with increased transmissibility, including the lineages B.1.1.7 (also known as 20I/501Y.V1 and VOC 202012/01) and B.1.351 (also known as 20H/501Y.V2), which were discovered in the United Kingdom and South Africa, respectively (Tegally, H., et al., Emergence and rapid spread of a new severe acute respiratory syndrome-related coronavirus 2 (SARS-CoV-2) lineage with multiple spike mutations in South Africa.
medRxiv, 2020: p. 2020.12.21.20248640; Leung, K., et al., Early empirical assessment of the N50 IY mutant strains of SAPS-CoV-2 in the United Kingdom, October to November 2020. medRxiv, 2020: p. 2020.12 20.20248581). B.1.351 also include two other mutations in the RBD domain of SARS-CoV2 spike protein, K417N and E484K
(Tegally, H., et al., Emergence and rapid spread of a new severe acute respiratory syndrome-related coronavirus 2 (SARS-CoV-2) lineage with multiple spike mutations in South Africa. medRxiv, 2020: p. 2020.12.21.20248640). Other SARS-CoV-2 variants include the Lineage B.1.1.28, which was first reported in Brazil; the Variant P.1, lineage B.1.1.28 (also known as 20J/501Y.V3), which was first reported in Japan;
Variant L452R, which was first reported in California in the United States (Pan American Health Organization, Epidemiological update: Occurrence of variants of SARS-CoV-2 in the Americas, January 20, 2021, available at https://reliefweb.int/sites/reliefweb.int/files/resources/2021-jan-20-phe-epi-update-SARS-CoV-2.pdf). Other SARS-CoV-2 variants include a SARS CoV-2 of clade 19A;
SARS CoV-2 of clade 19B; a SARS CoV-2 of clade 20A; a SARS CoV-2 of clade 20B;
a SARS CoV-2 of clade 20C; a SARS CoV-2 of clade 20D; a SARS CoV-2 of clade 20E (EU1); a SARS CoV-2 of clade 20F; a SARS CoV-2 of clade 20G; and SARS
CoV-2 B1.1.207; and other SARS CoV-2 lineages described in Rambaut, A., et al., A
dynamic nomenclature proposal .for SARS-CoV-2 lineages to assist genomic epidemiology. Nat Microbiol 5, 1403-1407 (2020). The Alpha (B.1.1.7), Beta (B.1.351, B.1.351.2, B.1.351.3), Delta (B.1.617.2, AY.1, AY.2, AY.3), and Gamma (P.1, P.1.1, P.1.2) variants of SARS-CoV-2 circulating in the United States are classified as variants of concern by the U.S. Centers for Disease Control and Prevention (see https://www.cdc.gov/coronavirus/2019-ncov/variants/variant-info.html).
Treating a SARS CoV-2 infection in accordance with the present disclosure includes treating infection by any one or more of the aforementioned SARS-CoV-2 viruses. In certain embodiments, treating a SARS-CoV-2 infection comprises treating any one or more of:
SARS CoV-2 Wuhan-Hu-1; a SARS-CoV-2 variant comprising a N439K mutation; a SARS-CoV-2 variant comprising a N501 Y mutation; a SARS-CoV-2 variant comprising a K417N mutation and/or a E484K mutation; a SARS-CoV-2 comprising a L452R mutation; B.1.1.28; B.1.1.7 (also referred-to as the "alpha" variant);
B.1.351 (also referred-to as the "beta" variant); P.1 (also referred-to as the "gamma"
variant);
B.1.617.1 (also referred-to as the "kappa" variant); B.1.429 (also referred-to as the "epsilon" variant); B.1.525 (also referred-to as the "eta" variant); B.1.526 (also referred-to as the "iota" variant); B.1.258; a variant of Wuhan-Hu-1 comprising a N440K
mutation; B.1.243.1; B.1.258 with a K417N mutation; A.27.1; R.1; P.2; R.2;
B.1.1.519;
A.23.1; B.1.318; B.1.619; A.VOI.V2; B.1.618; a variant of Wuhan-Hu-1 comprising N440K and E484K mutations; B.1.617.2 (also referred-to as the "delta"
variant);
B.1.1.298; B.1.617.2-AY.1; B.1.617.2-AY.2; C.37 (also referred-to as the "lambda"
variant); a SARS CoV-2 of clade 19A; SARS CoV-2 of clade 19B; a SARS CoV-2 of clade 20A; a SARS CoV-2 of clade 20B; a SARS CoV-2 of clade 20C; a SARS CoV-2 of clade 20D; a SARS CoV-2 of clade 20E (EU1); a SARS CoV-2 of clade 20F; and a SARS CoV-2 of clade 20G. Other coronaviruses are believed to enter cells by binding to other receptors (e.g., 9-0-Ac-Sia receptor analog; DPP4; APN).
In the present description, any concentration range, percentage range, ratio range, or integer range is to be understood to include the value of any integer within the recited range and, when appropriate, fractions thereof (such as one tenth and one hundredth of an integer), unless otherwise indicated. Also, any number range recited herein relating to any physical feature, such as polymer subunits, size or thickness, are to be understood to include any integer within the recited range, unless otherwise indicated. As used herein, the term "about" means 20% of the indicated range, value, or structure, unless otherwise indicated. In particular embodiments, "about"
comprises 5%, 10%, or 15%.
It should be understood that the terms "a" and "an" as used herein refer to "one or more" of the enumerated components. The use of the alternative (e.g., "or") should be understood to mean either one, both, or any combination thereof of the alternatives.
As used herein, the terms "include," "have," and "comprise" are used synonymously, which terms and variants thereof are intended to be construed as non-limiting.

"Optional" or "optionally" means that the subsequently described element, component, event, or circumstance may or may not occur, and that the description includes instances in which the element, component, event, or circumstance occurs and instances in which they do not.
In addition, it should be understood that the individual constructs, or groups of constructs, derived from the various combinations of the structures and subunits described herein, are disclosed by the present application to the same extent as if each construct or group of constructs was set forth individually. Thus, selection of particular structures or particular subunits is within the scope of the present disclosure.

The term "consisting essentially of' is not equivalent to "comprising" and refers to the specified materials or steps of a claim, or to those that do not materially affect the basic characteristics of a claimed subject matter. For example, a protein domain, region, or module (e.g., a binding domain) or a protein "consists essentially of' a particular amino acid sequence when the amino acid sequence of a domain, region, module, or protein includes extensions, deletions, mutations, or a combination thereof (e.g., amino acids at the amino- or carboxy-terminus or between domains) that, in combination, contribute to at most 20% (e.g., at most 15%, 10%, 8%, 6%, 5%, 4%, 3%, 2% or 1%) of the length of a domain, region, module, or protein and do not substantially affect (i.e., do not reduce the activity by more than 50%, such as no more than 40%, 30%, 25%, 20%, 15%, 10%, 5%, or 1%) the activity of the domain(s), region(s), module(s), or protein (e.g., the target binding affinity of a binding protein).
As used herein, "amino acid" refers to naturally occurring and synthetic amino acids, as well as amino acid analogs and amino acid mimetics that function in a manner similar to the naturally occurring amino acids. Naturally occurring amino acids are those encoded by the genetic code, as well as those amino acids that are later modified, e.g., hydroxyproline, y-carboxyglutamate, and 0-phosphoserine. Amino acid analogs refer to compounds that have the same basic chemical structure as a naturally occurring amino acid, i.e., an a-carbon that is bound to a hydrogen, a carboxyl group, an amino group, and an R group, e.g., homoserine, norleucine, methionine sulfoxide, methionine methyl sulfonium. Such analogs have modified R groups (e.g., norleucine) or modified peptide backbones, but retain the same basic chemical structure as a naturally occurring amino acid. Amino acid mimetics refer to chemical compounds that have a structure that is different from the general chemical structure of an amino acid, but that functions in a manner similar to a naturally occurring amino acid.
As used herein, "mutation" refers to a change in the sequence of a nucleic acid molecule or polypeptide molecule as compared to a reference or wild-type nucleic acid molecule or polypepti de molecule, respectively A mutation can result in several different types of change in sequence, including substitution, insertion or deletion of nucleotide(s) or amino acid(s).

A "conservative substitution" refers to amino acid substitutions that do not significantly affect or alter binding characteristics of a particular protein.
Generally, conservative substitutions are ones in which a substituted amino acid residue is replaced with an amino acid residue having a similar side chain. Conservative substitutions include a substitution found in one of the following groups: Group 1: Alanine (Ala or A), Glycine (Gly or G), Serine (Ser or S), Threonine (Thr or T); Group 2:
Aspartic acid (Asp or D), Glutamic acid (Glu or Z); Group 3: Asparagine (Asn or N), Glutamine (Gln or Q); Group 4: Arginine (Arg or R), Lysine (Lys or K), Histidine (His or H);
Group 5:
Isoleucine (Ile or I), Leucine (Leu or L), Methionine (Met or M), Valine (Val or V); and Group 6: Phenylalanine (Phe or F), Tyrosine (Tyr or Y), Tryptophan (Trp or W).
Additionally or alternatively, amino acids can be grouped into conservative substitution groups by similar function, chemical structure, or composition (e.g., acidic, basic, aliphatic, aromatic, or sulfur-containing). For example, an aliphatic grouping may include, for purposes of substitution, Gly, Ala, Val, Leu, and Ile. Other conservative substitutions groups include: sulfur-containing: Met and Cysteine (Cys or C);
acidic:
Asp, Glu, Asn, and Gln; small aliphatic, nonpolar or slightly polar residues:
Ala, Ser, Thr, Pro, and Gly, polar, negatively charged residues and their amides: Asp, Asn, Glu, and Gln; polar, positively charged residues: His, Arg, and Lys; large aliphatic, nonpolar residues: Met, Leu, Ile, Val, and Cys; and large aromatic residues: Phe, Tyr, and Trp.
Additional information can be found in Creighton (1984) Proteins, W.H Freeman and Company.
As used herein, "protein" or "polypeptide" refers to a polymer of amino acid residues. Proteins apply to naturally occurring amino acid polymers, as well as to amino acid polymers in which one or more amino acid residue is an artificial chemical mimetic of a corresponding naturally occurring amino acid, and non-naturally occurring amino acid polymers. Variants of proteins, peptides, and polypeptides of this disclosure are also contemplated. In certain embodiments, variant proteins, peptides, and polypeptides comprise or consist of an amino acid sequence that is at least 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 99.9%
identical to an amino acid sequence of a defined or reference amino acid sequence as described herein.

"Nucleic acid molecule" or "polynucleotide" or "polynucleic acid" refers to a polymeric compound including covalently linked nucleotides, which can be made up of natural subunits (e.g., purine or pyrimidine bases) or non-natural subunits (e.g., morpholine ring). Purine bases include adenine, guanine, hypoxanthine, and xanthine, and pyrimidine bases include uracil, thymine, and cytosine. Nucleic acid molecules include polyribonucleic acid (RNA), which includes mRNA, microRNA, siRNA, viral genomic RNA, and synthetic RNA, and polydeoxyribonucleic acid (DNA), which includes cDNA, genomic DNA, and synthetic DNA, either of which may be single or double stranded. If single-stranded, the nucleic acid molecule may be the coding strand or non-coding (anti-sense) strand. A nucleic acid molecule encoding an amino acid sequence includes all nucleotide sequences that encode the same amino acid sequence.
Some versions of the nucleotide sequences may also include intron(s) to the extent that the intron(s) would be removed through co- or post-transcriptional mechanisms.
In other words, different nucleotide sequences may encode the same amino acid sequence as the result of the redundancy or degeneracy of the genetic code, or by splicing.
Variants of nucleic acid molecules of this disclosure are also contemplated.
Variant nucleic acid molecules are at least 70%, 75%, 80%, 85%, 90%, and are preferably 95%, 96%, 97%, 98%, 99%, or 99.9% identical a nucleic acid molecule of a defined or reference polynucleotide as described herein, or that hybridize to a polynucleotide under stringent hybridization conditions of 0.015M sodium chloride, 0.0015M sodium citrate at about 65-68 C or 0.015M sodium chloride, 0.0015M
sodium citrate, and 50% formamide at about 42 C. Nucleic acid molecule variants retain the capacity to encode a binding domain thereof having a functionality described herein, such as binding a target molecule.
"Percent sequence identity" refers to a relationship between two or more sequences, as determined by comparing the sequences. Preferred methods to determine sequence identity are designed to give the best match between the sequences being compared For example, the sequences are aligned for optimal comparison purposes (e.g., gaps can be introduced in one or both of a first and a second amino acid or nucleic acid sequence for optimal alignment). Further, non-homologous sequences may be disregarded for comparison purposes. The percent sequence identity referenced herein is calculated over the length of the reference sequence, unless indicated otherwise.
Methods to determine sequence identity and similarity can be found in publicly available computer programs. Sequence alignments and percent identity calculations may be performed using a BLAST program (e.g., BLAST 2.0, BLASTP, BLASTN, or BLASTX). The mathematical algorithm used in the BLAST programs can be found in Altschul et al., Nucleic Acids Res. 25:3389-3402, 1997. Within the context of this disclosure, it will be understood that where sequence analysis software is used for analysis, the results of the analysis are based on the "default values" of the program referenced. "Default values" mean any set of values or parameters which originally load with the software when first initialized.
The term "isolated" means that the material is removed from its original environment (e.g., the natural environment if it is naturally occurring). For example, a naturally occurring nucleic acid or polypeptide present in a living animal is not isolated, but the same nucleic acid or polypeptide, separated from some or all of the co-existing materials in the natural system, is isolated. Such nucleic acid could be part of a vector and/or such nucleic acid or polypeptide could be part of a composition (e.g., a cell lysate), and still be isolated in that such vector or composition is not part of the natural environment for the nucleic acid or polypeptide. "Isolated" can, in some embodiments, also describe an antibody, antigen-binding fragment, polynucleotide, vector, host cell, or composition that is outside of a human body.
The term "gene" means the segment of DNA or RNA involved in producing a polypeptide chain; in certain contexts, it includes regions preceding and following the coding region (e.g., 5' untranslated region (UTR) and 3' UTR) as well as intervening sequences (introns) between individual coding segments (exons).
A "functional variant" refers to a polypeptide or polynucleotide that is structurally similar or substantially structurally similar to a parent or reference compound of this disclosure, but differs slightly in composition (e.g., one base, atom or functional group is different, added, or removed), such that the polypeptide or encoded polypeptide is capable of performing at least one function of the parent polypeptide with at least 50% efficiency, preferably at least 55%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, 99.9%, or 100% level of activity of the parent polypeptide.

In other words, a functional variant of a polypeptide or encoded polypeptide of this disclosure has "similar binding," "similar affinity" or "similar activity"
when the functional variant displays no more than a 50% reduction in performance in a selected assay as compared to the parent or reference polypeptide, such as an assay for measuring binding affinity (e.g., Biacore or tetramer staining measuring an association (Ka) or a dissociation (Ku) constant).
As used herein, a "functional portion" or "functional fragment" refers to a polypeptide or polynucleotide that comprises only a domain, portion or fragment of a parent or reference compound, and the polypeptide or encoded polypeptide retains at least 50% activity associated with the domain, portion or fragment of the parent or reference compound, preferably at least 55%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, 99.9%, or 100% level of activity of the parent polypeptide, or provides a biological benefit (e.g., effector function). A "functional portion" or "functional fragment" of a polypeptide or encoded polypeptide of this disclosure has "similar binding" or "similar activity" when the functional portion or fragment displays no more than a 50% reduction in performance in a selected assay as compared to the parent or reference polypeptide (preferably no more than 20% or 10%, or no more than a log difference as compared to the parent or reference with regard to affinity).
As used herein, the term "engineered," "recombinant," or "non-natural" refers to an organism, microorganism, cell, nucleic acid molecule, or vector that includes at least one genetic alteration or has been modified by introduction of an exogenous or heterologous nucleic acid molecule, wherein such alterations or modifications are introduced by genetic engineering (i.e., human intervention). Genetic alterations include, for example, modifications introducing expressible nucleic acid molecules encoding functional RNA, proteins, fusion proteins or enzymes, or other nucleic acid molecule additions, deletions, substitutions, or other functional disruption of a cell's genetic material. Additional modifications include, for example, non-coding regulatory regions in which the modifications alter expression of a polynucleotide, gene, or operon.
As used herein, "heterologous" or "non-endogenous" or "exogenous" refers to any gene, protein, compound, nucleic acid molecule, or activity that is not native to a host cell or a subject, or any gene, protein, compound, nucleic acid molecule, or activity native to a host cell or a subject that has been altered. Heterologous, non-endogenous, or exogenous includes genes, proteins, compounds, or nucleic acid molecules that have been mutated or otherwise altered such that the structure, activity, or both is different as between the native and altered genes, proteins, compounds, or nucleic acid molecules.
In certain embodiments, heterologous, non-endogenous, or exogenous genes, proteins, or nucleic acid molecules (e.g., receptors, ligands, etc.) may not be endogenous to a host cell or a subject, but instead nucleic acids encoding such genes, proteins, or nucleic acid molecules may have been added to a host cell by conjugation, transformation, transfection, electroporation, or the like, wherein the added nucleic acid molecule may integrate into a host cell genome or can exist as extra-chromosomal genetic material (e.g., as a plasmid or other self-replicating vector) The term "homologous" or "homolog" refers to a gene, protein, compound, nucleic acid molecule, or activity found in or derived from a host cell, species, or strain. For example, a heterologous or exogenous polynucleotide or gene encoding a polypeptide may be homologous to a native polynucleotide or gene and encode a homologous polypeptide or activity, but the polynucleotide or polypeptide may have an altered structure, sequence, expression level, or any combination thereof. A non-endogenous polynucleotide or gene, as well as the encoded polypeptide or activity, may be from the same species, a different species, or a combination thereof.
In certain embodiments, a nucleic acid molecule or portion thereof native to a host cell will be considered heterologous to the host cell if it has been altered or mutated, or a nucleic acid molecule native to a host cell may be considered heterologous if it has been altered with a heterologous expression control sequence or has been altered with an endogenous expression control sequence not normally associated with the nucleic acid molecule native to a host cell. In addition, the term "heterologous" can refer to a biological activity that is different, altered, or not endogenous to a host cell As described herein, more than one heterologous nucleic acid molecule can be introduced into a host cell as separate nucleic acid molecules, as a plurality of individually controlled genes, as a polycistronic nucleic acid molecule, as a single nucleic acid molecule encoding a fusion protein, or any combination thereof.
When As used herein, the term "endogenous" or "native" refers to a polynucleotide, gene, protein, compound, molecule, or activity that is normally present in a host cell or a subject.
The term "expression", as used herein, refers to the process by which a polypeptide is produced based on the encoding sequence of a nucleic acid molecule, such as a gene. The process may include transcription, post-transcriptional control, post-transcriptional modification, translation, post-translational control, post-translational modification, or any combination thereof An expressed nucleic acid molecule is typically operably linked to an expression control sequence (e.g., a promoter).
The term "operably linked" refers to the association of two or more nucleic acid molecules on a single nucleic acid fragment so that the function of one is affected by the other. For example, a promoter is operably linked with a coding sequence when it is capable of affecting the expression of that coding sequence (i.e., the coding sequence is under the transcriptional control of the promoter). "Unlinked" means that the associated genetic elements are not closely associated with one another and the function of one does not affect the other.
As described herein, more than one heterologous nucleic acid molecule can be introduced into a host cell as separate nucleic acid molecules, as a plurality of individually controlled genes, as a polycistronic nucleic acid molecule, as a single nucleic acid molecule encoding a protein (e.g., a heavy chain of an antibody), or any combination thereof. When two or more heterologous nucleic acid molecules are introduced into a host cell, it is understood that the two or more heterologous nucleic acid molecules can be introduced as a single nucleic acid molecule (e.g., on a single vector), on separate vectors, integrated into the host chromosome at a single site or multiple sites, or any combination thereof The number of referenced heterologous nucleic acid molecules or protein activities refers to the number of encoding nucleic acid molecules or the number of protein activities, not the number of separate nucleic acid molecules introduced into a host cell.

The term "construct" refers to any polynucleotide that contains a recombinant nucleic acid molecule (or, when the context clearly indicates, a fusion protein of the present disclosure). A (polynucleotide) construct may be present in a vector (e.g., a bacterial vector, a viral vector) or may be integrated into a genome. A
"vector" is a nucleic acid molecule that is capable of transporting another nucleic acid molecule.
Vectors may be, for example, plasmids, cosmids, viruses, a RNA vector or a linear or circular DNA or RNA molecule that may include chromosomal, non-chromosomal, semi-synthetic or synthetic nucleic acid molecules. Vectors of the present disclosure also include transposon systems (e.g., Sleeping Beauty, see, e.g., Geurts et al., Mol.
Ther. 8:108, 2003: Mates et al, Nat. Genet. 41:753, 2009). Exemplary vectors are those capable of autonomous replication (episomal vector), capable of delivering a polynucleotide to a cell genome (e.g., viral vector), or capable of expressing nucleic acid molecules to which they are linked (expression vectors).
As used herein, "expression vector" or "vector" refers to a DNA construct containing a nucleic acid molecule that is operably linked to a suitable control sequence capable of effecting the expression of the nucleic acid molecule in a suitable host. Such control sequences include a promoter to effect transcription, an optional operator sequence to control such transcription, a sequence encoding suitable mRNA
ribosome binding sites, and sequences which control termination of transcription and translation.
The vector may be a plasmid, a phage particle, a virus, or simply a potential genomic insert. Once transformed into a suitable host, the vector may replicate and function independently of the host genome, or may, in some instances, integrate into the genome itself or deliver the polynucleotide contained in the vector into the genome without the vector sequence. In the present specification, "plasmid," "expression plasmid," "virus,"
and "vector" are often used interchangeably.
The term "introduced" in the context of inserting a nucleic acid molecule into a cell, means "transfection", "transformation," or "transduction" and includes reference to the incorporation of a nucleic acid molecule into a eukaryotic or prokaryotic cell wherein the nucleic acid molecule may be incorporated into the genome of a cell (e.g., chromosome, plasmid, plastid, or mitochondrial DNA), converted into an autonomous replicon, or transiently expressed (e.g., transfected mRNA).

In certain embodiments, polynucleotides of the present disclosure may be operatively linked to certain elements of a vector. For example, polynucleotide sequences that are needed to effect the expression and processing of coding sequences to which they are ligated may be operatively linked. Expression control sequences may include appropriate transcription initiation, termination, promoter, and enhancer sequences; efficient RNA processing signals such as splicing and polyadenylation signals; sequences that stabilize cytoplasmic mRNA; sequences that enhance translation efficiency (i.e., Kozak consensus sequences); sequences that enhance protein stability;
and possibly sequences that enhance protein secretion. Expression control sequences may be operatively linked if they are contiguous with the gene of interest and expression control sequences that act in trans or at a distance to control the gene of interest.
In certain embodiments, the vector comprises a plasmid vector or a viral vector (e.g., a lentiviral vector or a y-retroviral vector). Viral vectors include retrovirus, 1.5 adenovirus, parvovirus (e.g., adeno-associated viruses), coronavirus, negative strand RNA viruses such as ortho-myxovirus (e.g., influenza virus), rhabdovirus (e.g., rabies and vesicular stomatitis virus), paramyxovirus (e.g., measles and Sendai), positive strand RNA viruses such as picornavirus and alphavirus, and double-stranded DNA
viruses including adenovirus, herpesvirus (e.g., Herpes Simplex virus types 1 and 2, Epstein-Barr virus, cytomegalovirus), and poxvirus (e.g., vaccini a, fowlpox, and canarypox). Other viruses include, for example, Norwalk virus, togavirus, flavivirus, reoviruses, papovavirus, hepadnavirus, and hepatitis virus. Examples of retroviruses include avian leukosis-sarcoma, mammalian C-type, B-type viruses, D type viruses, HTLV-BLV group, lentivirus, spumavirus (Coffin, J. M., Retroviridae: The viruses and their replication, In Fundamental Virology, Third Edition, B. N. Fields et al., Eds., Lippincott-Raven Publishers, Philadelphia, 1996).
"Retroviruses" are viruses having an RNA genome, which is reverse-transcribed into DNA using a reverse transcriptase enzyme, the reverse-transcribed DNA is then incorporated into the host cell genome. "Gammaretrovirus" refers to a genus of the retroviridae family. Examples of gammaretroviruses include mouse stem cell virus, murine leukemia virus, feline leukemia virus, feline sarcoma virus, and avian reticuloendotheliosis viruses.
"Lentiviral vectors" include HIV-based lentiviral vectors for gene delivery, which can be integrative or non-integrative, have relatively large packaging capacity, and can transduce a range of different cell types. Lentiviral vectors are usually generated following transient transfection of three (packaging, envelope, and transfer) or more plasmids into producer cells. Like HIV, lentiviral vectors enter the target cell through the interaction of viral surface glycoproteins with receptors on the cell surface.
On entry, the viral RNA undergoes reverse transcription, which is mediated by the viral reverse transcriptase complex. The product of reverse transcription is a double-stranded linear viral DNA, which is the substrate for viral integration into the DNA of infected cells.
In certain embodiments, the viral vector can be a gammaretrovirus, e.g., Moloney murine leukemia virus (MLV)-derived vectors. In other embodiments, the viral vector can be a more complex retrovirus-derived vector, e.g., a lentivirus-derived vector. HIV-1-derived vectors belong to this category. Other examples include lentivirus vectors derived from HIV-2, Fly, equine infectious anemia virus, Sly, and Maedi-Visna virus (ovine lentivirus). Methods of using retroviral and lentiviral viral vectors and packaging cells for transducing mammalian host cells with viral particles containing transgenes are known in the art and have been previous described, for example, in: U.S. Patent 8,119,772; Walchli et at., PLoS One 6:327930, 2011;
Zhao et at., J. Immunol. /74:4415, 2005; Engels et at., Hum. Gene Ther. 14:1155, 2003;
Frecha et at., Mot. Ther. 18:1748, 2010; and Verhoeyen et at., Methods Mot. Biol.
506:97, 2009. Retroviral and lentiviral vector constructs and expression systems are also commercially available. Other viral vectors also can be used for polynucleotide delivery including DNA viral vectors, including, for example adenovirus-based vectors and adeno-associated virus (AAV)-based vectors; vectors derived from herpes simplex viruses (HSVs), including amplicon vectors, replication-defective HSV and attenuated HSV (Krisky et at., Gene Ther. 5:1517 , 1998).
Other vectors that can be used with the compositions and methods of this disclosure include those derived from baculoviruses and a-viruses. (Jolly, D
J. 1999.

Emerging Viral Vectors. pp 209-40 in Friedmann T. ed. The Development of Human Gene Therapy. New York: Cold Spring Harbor Lab), or plasmid vectors (such as sleeping beauty or other transposon vectors).
When a viral vector genome comprises a plurality of polynueleotides to be expressed in a host cell as separate transcripts, the viral vector may also comprise additional sequences between the two (or more) transcripts allowing for bicistronic or multicistronic expression. Examples of such sequences used in viral vectors include internal ribosome entry sites (IRES), furin cleavage sites, viral 2A peptide, or any combination thereof.
Plasmid vectors, including DNA-based antibody or antigen-binding fragment-encoding plasmid vectors for direct administration to a subject, are described further herein.
As used herein, the term "host" refers to a cell or microorganism targeted for genetic modification with a heterologous nucleic acid molecule to produce a polypeptide of interest (e.g., an antibody of the present disclosure).
A host cell may include any individual cell or cell culture which may receive a vector or the incorporation of nucleic acids or express proteins. The term also encompasses progeny of the host cell, whether genetically or phenotypically the same or different. Suitable host cells may depend on the vector and may include mammalian cells, animal cells, human cells, simian cells, insect cells, yeast cells, and bacterial cells.
These cells may be induced to incorporate the vector or other material by use of a viral vector, transformation via calcium phosphate precipitation, DEAE-dextran, el ectroporati on, mi croinj ecti on, or other methods. See, for example, Sambrook etal., Molecular Cloning: A Laboratory Manual 2d ed. (Cold Spring Harbor Laboratory, 1989).
In the context of a SARS-CoV-2 infection, a "host" refers to a cell or a subject infected with the SARS-CoV-2 coronavirus.
"Antigen" or "Ag", as used herein, refers to an immunogenic molecule that provokes an immune response. This immune response may involve antibody production, activation of specific immunologically-competent cells, activation of complement, antibody dependent cytotoxicicity, or any combination thereof An antigen (immunogenic molecule) may be, for example, a peptide, glycopeptide, polypeptide, glycopolypeptide, polynucleotide, polysaccharide, lipid, or the like. It is readily apparent that an antigen can be synthesized, produced recombinantly, or derived from a biological sample. Exemplary biological samples that can contain one or more antigens include tissue samples, stool samples, cells, biological fluids, or combinations thereof. Antigens can be produced by cells that have been modified or genetically engineered to express an antigen. Antigens can also be present in a SARS-CoV-2 coronavirus (e.g., a surface glycoprotein or portion thereof), such as present in a virion, or expressed or presented on the surface of a cell infected by SARS-CoV-2.
The term "epitope" or "antigenic epitope" includes any molecule, structure, amino acid sequence, or protein determinant that is recognized and specifically bound by a cognate binding molecule, such as an immunoglobulin, or other binding molecule, domain, or protein. Epitopic determinants generally contain chemically active surface groupings of molecules, such as amino acids or sugar side chains, and can have specific three-dimensional structural characteristics, as well as specific charge characteristics.
Where an antigen is or comprises a peptide or protein, the epitope can be comprised of consecutive amino acids (e.g., a linear epitope), or can be comprised of amino acids from different parts or regions of the protein that are brought into proximity by protein folding (e.g., a discontinuous or conformational epitope), or non-contiguous amino acids that are in close proximity irrespective of protein folding.
Antibodies, Antigen-Binding Fragments, and Compositions In one aspect, the present disclosure provides an isolated antibody, or an antigen-binding fragment thereof, that comprises a heavy chain variable domain (VH) comprising a CDRH1, a CDRH2, and a CDRH3, and a light chain variable domain (VL) comprising a CDRL1, a CDRL2, and a CDRL3, and is capable of binding to a surface glycoprotein of SARS-CoV-2, in particular in an epitope that is at least partially comprised in or defined by Domain A. In certain embodiments, the antibody or antigen-binding fragment is capable of binding to a surface glycoprotein of SARS-CoV-2 expressed on a cell surface of a host cell and/or on a SARS-CoV-2 virion In certain embodiments, an antibody or antigen-binding fragment of the present disclosure associates with or unites with a SARS-CoV-2 surface glycoprotein Domain A epitope or antigen comprising the epitope, while not significantly associating or uniting with any other molecules or components in a sample.
In certain embodiments, an antibody or antigen binding fragment of the present disclosure is cross-reactive for SARS-CoV-2 and one or more additional sarbecovirus of clade 2, but not of clade 1 or clade 3. In certain embodiments, an antibody or antigen binding fragment of the present disclosure is not cross-reactive against an embecovirus, a merbecovirus, or both.
In certain embodiments, an antibody or antigen-binding fragment of the present disclosure specifically binds to a SARS-CoV-2 surface glycoprotein. As used herein, "specifically binds" refers to an association or union of an antibody or antigen-binding fragment to an antigen with an affinity or Ka (i.e., an equilibrium association constant of a particular binding interaction with units of 1/M) equal to or greater than 1051\44 (which equals the ratio of the on-rate [Km] to the off rate [Karr] for this association reaction), while not significantly associating or uniting with any other molecules or components in a sample. Alternatively, affinity may be defined as an equilibrium dissociation constant (Ka) of a particular binding interaction with units of M
(e.g., 10-5 M to 10-13 M). Antibodies may be classified as "high-affinity" antibodies or as "low-affinity" antibodies. "High-affinity" antibodies refer to those antibodies having a Ka of at least 107M4, at least 1081\41, at least 109 M-1, at least 1010 M-1, at least 1011 M4, at least 1012M4, or at least 10'3 M4 "Low-affinity" antibodies refer to those antibodies having a Ka of up to 107M4, up to 106 M-", up to 105 M1. Alternatively, affinity may be defined as an equilibrium dissociation constant (Ka) of a particular binding interaction with units of M (e.g., le M to I 013 M) In some contexts, antibody and antigen-binding fragments may be described with reference to affinity and/or to avidity for antigen. Unless otherwise indicated, avidity refers to the total binding strength of an antibody or antigen-binding fragment thereof to antigen, and reflects binding affinity, valency of the antibody or antigen-binding fragment (e.g., whether the antibody or antigen-binding fragment comprises one, two, three, four, five, six, seven, eight, nine, ten, or more binding sites), and, for example, whether another agent is present that can affect the binding (e.g., a non-competitive inhibitor of the antibody or antigen-binding fragment).

A variety of assays are known for identifying antibodies of the present disclosure that bind a particular target, as well as determining binding domain or binding protein affinities, such as Western blot, ELISA (e.g., direct, indirect, or sandwich), analytical ultracentrifugation, spectroscopy, and surface plasmon resonance (Biacoreg) analysis (see, e.g., Scatchard et al., Ann. N.Y. Acad. Sci. 51:660, 1949;
Wilson, Science 295:2103, 2002; Wolff et al., Cancer Res. 53:2560, 1993; and U.S.
Patent Nos. 5,283,173, 5,468,614, or the equivalent). Assays for assessing affinity or apparent affinity or relative affinity are also known.
In certain examples, binding can be determined by recombinantly expressing a SARS-CoV-2 antigen in a host cell (e.g., by transfection) and immunostaining the (e.g., fixed, or fixed and permeabilized) host cell with antibody and analyzing binding by flow cytometry (e.g., using a ZE5 Cell Analyzer (BioRadg) and FlowJo software (TreeStar). In some embodiments, positive binding can be defined by differential staining by antibody of SARS-CoV-2 -expressing cells versus control (e.g., mock) cells.
In some embodiments an antibody or antigen-binding fragment of the present disclosure binds to SARS-CoV-2 S protein, as measured using biolayer interferometry.
In certain embodiments, an antibody or antigen-binding fragment of the present disclosure binds to SARS-CoV-2 S protein with a KD of less than about 4.5x109 M, less than about 5x109 M, less than about 1x104 M, less than about 5x104 M, less than about 1x10-11 M, less than about 5x10-11 M, less than about 1x10'2 M, or less than about 5x1012 M.
Certain characteristics of presently disclosed antibodies or antigen-binding fragments may be described using IC50 or EC50 values. In certain embodiments, the IC50 is the concentration of a composition (e.g., antibody) that results in half-maximal inhibition of the indicated biological or biochemical function, activity, or response. In certain embodiments, the EC50 is the concentration of a composition that provides the half-maximal response in the assay. In some embodiments, e.g., for describing the ability of a presently disclosed antibody or antigen-binding fragment to neutralize infection by SARS-CoV-2, IC50 and EC50 are used interchangeably.
In certain embodiments, an antibody of the present disclosure is capable of neutralizing infection by SARS-CoV-2. As used herein, a "neutralizing antibody" is one that can neutralize, i.e., prevent, inhibit, reduce, impede, or interfere with, the ability of a pathogen to initiate and/or perpetuate an infection in a host.
Neutralization may be quantified by, for example, assessing SARS-CoV-2 RNA levels in a(n e.g.

lung) sample, assessing SARS-CoV-2 viral load in a(n e.g. lung) sample, assessing histopathology of a(n e.g. lung) sample, or the like. The terms "neutralizing antibody"
and "an antibody that neutralizes" or "antibodies that neutralize" are used interchangeably herein. In any of the presently disclosed embodiments, the antibody or antigen-binding fragment is capable of preventing and/or neutralizing a SARS-CoV-2 infection in an in vitro model of infection and/or in an in vivo animal model of infection (e.g., using a Syrian hamster model with intranasal delivery of SARS-CoV-2) and/or in a human.
In certain embodiments, the antibody or antigen-binding fragment (i) recognizes an epitope in the Domain A of SARS-CoV-2; (ii) is capable of neutralizing a SARS
CoV-2 infection; (iii) is capable of eliciting at least one immune effector function against SARS CoV-2; (iv) is capable of preventing shedding, from a cell infected with SARS CoV-2, of Si protein; or (v) any combination of (i)-(iv).
Terms understood by those in the art of antibody technology are each given the meaning acquired in the art, unless expressly defined differently herein. For example, the term "antibody" refers to an intact antibody comprising at least two heavy (H) chains and two light (L) chains inter-connected by disulfide bonds, as well as any antigen-binding portion or fragment of an intact antibody that has or retains the ability to bind to the antigen target molecule recognized by the intact antibody, such as an scFv, Fab, or Fab'2 fragment. Thus, the term "antibody" herein is used in the broadest sense and includes polyclonal and monoclonal antibodies, including intact antibodies and functional (antigen-binding) antibody fragments thereof, including fragment antigen binding (Fab) fragments, F(abl)2 fragments, Fab fragments, Fv fragments, recombinant IgG (rIgG) fragments, single chain antibody fragments, including single chain variable fragments (scFv), and single domain antibodies (e.g., sdAb, sdFv, nanobody) fragments. The term encompasses genetically engineered and/or otherwise modified forms of immunoglobulins, such as intrabodies, peptibodies, chimeric antibodies, fully human antibodies, humanized antibodies, and heteroconjugate antibodies, multispecific, e.g., bispecific antibodies, diabodies, triabodies, tetrabodies, tandem di-scFv, and tandem tri-scFv. Unless otherwise stated, the term "antibody"
should be understood to encompass functional antibody fragments thereof. The term also encompasses intact or full-length antibodies, including antibodies of any class or sub-class, including IgG and sub-classes thereof (IgGl, IgG2, IgG3, IgG4), IgM, IgE, IgA, and IgD.
The terms "VL" or "VL" and "VH" or "VH" refer to the variable binding region from an antibody light chain and an antibody heavy chain, respectively. In certain embodiments, a VL is a kappa (lc) class (also "VK" herein). In certain embodiments, a VL is a lambda (X) class. The variable binding regions comprise discrete, well-defined sub-regions known as "complementarity determining regions" (CDRs) and "framework regions" (FRs). The terms "complementarity determining region," and "CDR," are synonymous with "hypervariable region" or "HVR," and refer to sequences of amino acids within antibody variable regions, which, in general, together confer the antigen specificity and/or binding affinity of the antibody, wherein consecutive CDRs (i.e., CDR1 and CDR2, CDR2 and CDR3) are separated from one another in primary structure by a framework region. There are three CDRs in each variable region (HCDR1, HCDR2, HCDR3; LCDR1, LCDR2, LCDR3; also referred to as CDRHs and CDRLs, respectively). In certain embodiments, an antibody VH comprises four FRs and three CDRs as follows: FR1-HCDR1-FR2-HCDR2-FR3-HCDR3-FR4; and an antibody VL comprises four FRs and three CDRs as follows: FR1-LCDR1-FR2-LCDR2-FR3-LCDR3-FR4. In general, the VH and the VL together form the antigen-binding site through their respective CDRs.
As used herein, a "variant" of a CDR refers to a functional variant of a CDR
sequence having up to 1-3 amino acid substitutions (e.g., conservative or non-conservative substitutions), deletions, or combinations thereof.
Numbering of CDR and framework regions may be according to any known method or scheme, such as the Kabat, Chothia, EU, IMGT, and AHo numbering schemes (see, e.g., Kabat et al., " Sequences of Proteins of Immunological Interest, US
Dept. Health and Human Services, Public Health Service National Institutes of Health, 1991, 5th ed.; Chothia and Lesk, J. Mol. Biol. 196:901-917 (1987)); Lefranc et al., Dev.

Comp. Immunol. 27:55, 2003; Honegger and Pltickthun, J. Mol. Bio. 309:657-670 (2001)). Equivalent residue positions can be annotated and for different molecules to be compared using Antigen receptor Numbering And Receptor Classification (ANARCI) software tool (2016, Bioinformatics 15:298-300). Accordingly, identification of CDRs of an exemplary variable domain (VH or VL) sequence as provided herein according to one numbering scheme is not exclusive of an antibody comprising CDRs of the same variable domain as determined using a different numbering scheme. In certain embodiments, an antibody or antigen-binding fragment is provided that comprises CDRs in a VH sequence according to any one of SEQ
ID
NOs.: 22, 32, 42, 52, 62, 72, 82, 92, 102, 112, 122, 132, 142,152, 162, 172, 182 192, 202, 212, 222, 232, 242, 252, 262, 272, 282, 292, 302, 312, 322, 332, 342, 352, 362, 372, 382, 392, 402, 412, 422, and 432, and in a VL sequence according to any one of SEQ ID NOs.: 26, 36, 46, 56, 66, 76, 86, 96, 106, 116, 126, 136, 146, 156, 166, 176, 186, 196, 206, 216, 226, 236, 246, 256, 266, 276, 286, 296, 306, 316, 326, 336, 346, 356, 366, 376, 386, 396, 406, 416, 426, and 436, as determined using any known CDR
numbering method, including the Kabat, Chothia, EU, IMGT, Martin (Enhanced Chothia), Contact, and AHo numbering methods. In certain embodiments, CDRs are according to the IMGT numbering method. In certain embodiments, CDRs are according to the antibody numbering method developed by the Chemical Computing Group (CCG); e.g., using Molecular Operating Environment (MOE) software (www.chemcomp.com).
In certain embodiments, an antibody or an antigen-binding fragment is provided that comprises a heavy chain variable domain (VH) comprising a CDRH I , a CDRH2, and a CDRH3, and a light chain variable domain (VL) comprising a CDRL1, a CDRL2, and a CDRL3, wherein: (i) the CDRH1 comprises or consists of the amino acid sequence according to any one of SEQ ID NOs.: 23, 33, 43, 53, 63, 73, 83, 93, 103, 113, 123, 133, 143, 153, 163, 173, 183, 193, 203, 213, 223, 233, 243, 253, 263, 273, 283, 293, 303, 313, 323, 333, 343, 353, 363, 373, 383, 393, 403, 413, 423, or 433, or a sequence variant thereof comprising one, two, or three acid substitutions, one or more of which substitutions is optionally a conservative substitution and/or is a substitution to a germline-encoded amino acid; (ii) the CDRH2 comprises or consists of the amino acid sequence according to any one of SEQ ID NOs.: 24, 34, 44, 54, 64, 74, 84, 94, 104, 114, 124, 134, 144, 154, 164, 174, 184, 194, 204, 214, 224, 234, 244, 254, 264, 274, 284, 294, 304, 314, 324, 334, 344, 354, 364, 374, 384, 394, 404, 414, 424, or 434, or a sequence variant thereof comprising one, two, or three amino acid substitutions, one or more of which substitutions is optionally a conservative substitution and/or is a substitution to a germline-encoded amino acid; (iii) the CDRH3 comprises or consists of the amino acid sequence according to any one of SEQ ID NOs.. 25, 35, 45, 55, 65, 75, 85, 95, 105, 115, 125, 135, 145, 155, 165, 175, 185, 195, 205, 215, 225, 235, 245, 255, 265, 275, 285, 295, 305, 315, 325, 335, 345, 355, 365, 375, 385, 395, 405, 415, 425, or 435, or a sequence variant thereof comprising one, two, or three amino acid substitutions, one or more of which substitutions is optionally a conservative substitution and/or is a substitution to a germline-encoded amino acid, (iv) the CDRL1 comprises or consists of the amino acid sequence according to any one of SEQ
ID
NOs.: 27, 37, 47, 57, 67, 77, 87, 97, 107, 117, 127, 137, 147, 157, 167, 177, 187, 197, 207, 217, 227, 237, 247, 257, 267, 277, 287, 297, 307, 317, 327, 337, 347, 357, 367, 377, 387, 397, 407, 417, 427, or 437, or a sequence variant thereof comprising one, two, or three amino acid substitutions, one or more of which substitutions is optionally a conservative substitution and/or is a substitution to a germline-encoded amino acid; (v) the CDRL2 comprises or consists of the amino acid sequence according to any one of SEQ ID NOs 28, 38, 48, 58, 68, 78, 88, 98, 108, 118, 128, 138, 148, 158, 168, 178, 188, 198, 208, 218, 228, 238, 248, 258, 268, 278, 288, 298, 308, 318, 328, 338, 348, 358, 368, 378, 388, 398, 408, 418, 428, or 438, or a sequence variant thereof comprising one, two, or three amino acid substitutions, one or more of which substitutions is optionally a conservative substitution and/or is a substitution to a germline-encoded amino acid, and/or (vi) the CDRL3 comprises or consists of the amino acid sequence according to any one of SEQ ID NOs.: 29, 39, 49, 59, 69, 79, 89, 99, 109, 119, 129, 139, 149, 159, 169, 179, 189, 199, 209, 219, 229, 239, 249, 259, 269, 279, 289, 299, 309, 319, 329, 339, 349, 359, 369, 379, 389, 399, 409, 419, 429, or 439, or a sequence variant thereof comprising having one, two, or three amino acid substitutions, one or more of which substitutions is optionally a conservative substitution and/or is a substitution to a germline-encoded amino acid, wherein the antibody or antigen binding fragment is capable of binding to a surface glycoprotein of SARS-CoV-2. In some embodiments, the SARS-CoV-2 surface glycoprotein is expressed on a cell surface of a host cell and/or is present in a virion. In certain embodiments, the CDRs are according to the IMGT numbering method.
In any of the presently disclosed embodiments, the antibody or antigen-binding fragment is capable of preventing and/or neutralizing a SARS-CoV-2 infection in an in vitro model of infection and/or in an in vivo animal model of infection and/or in a human.
In any of the presently disclosed embodiments, the antibody or antigen-binding fragment comprises CDRH1, CDRH2, CDRH3, CDRL1, CDRL2, and CDRL3 amino acid sequences according to SEQ ID NOs.: (i) 23-25 and 27-29, respectively;
(ii) 33-35 and 37-39, respectively; (iii) 43-45 and 47-49, respectively; (iv) 53-55 and 57-59, respectively; (v) 63-65 and 67-69, respectively; (vi) 73-75 and 77-79, respectively; (vii) 83-85 and 87-89, respectively; (viii) 93-95 and 97-99, respectively; (ix) 103-105 and 107-109, respectively; (x) 113-115 and 117-119, respectively; (xi) 123-125 and 129, respectively; (xii) 133-135 and 137-139, respectively, (xiii) 143-145 and 147-149, respectively, (xiv) 153-155 and 157-159, respectively, (xv) 163-165 and 167-169, respectively; (xvi) 173-175 and 177-179, respectively; (xvii) 183-185 and 187-189, respectively; (xviii) 193-195 and 197-199, respectively; (xix) 203-205 and 207-209, respectively; (xx) 213-215 and 217-219, respectively; (xxi) 223-225 and 227-229, respectively; (xxii) 233-235 and 237-239, respectively; (xxiii) 243-245 and 247-249, respectively; (xxiv) 253-255 and 257-259, respectively; (xxv) 263-265 and 267-269, respectively; (xxvi) 273-275 and 277-279, respectively; (xxvii) 283-285 and 287-289, respectively; (xxviii) 293-295 and 297-299, respectively; (xxix) 303-305 and 307-309, respectively; (xxx) 313-315 and 317-319, respectively; (xxxi) 323-325 and 327-329, respectively; (xxxii) 333-335 and 337-339, respectively; (xxxiii) 343-345 and 347-349, respectively; (xxxiv) 353-355 and 357-359, respectively; (xxxv) 363-365 and 367-369, respectively; (xxxvi) 373-375 and 377-379, respectively; (xxxvii) 383-385 and 389, respectively; (xxxviii) 393-395 and 397-399, respectively; (xxxix) 403-405 and 407-409, respectively; (xxxx) 413-415 and 417-419, respectively; (xxxxi) 423-425 and 427-429, respectively; or (xxxxii) 433-435 and 437-439, respectively.

In some embodiments, an antibody or antigen-binding fragment is provided that comprises CDRH1, CDRH2, CDRH3, CDRL1, CDRL2, and CDRL3 amino acid sequences as set forth in SEQ ID NOs.:163-165 and 167-169, respectively. In certain embodiments, the antibody or antigen-binding fragment comprises VH and VL
amino acid sequences as set forth in SEQ ID NOs.:162 and 166, respectively.
In some embodiments, an antibody or antigen-binding fragment is provided that comprises CDRH1, CDRH2, CDRH3, CDRL1, CDRL2, and CDRL3 amino acid sequences as set forth in SEQ ID NOs.:103-105 and 107-109, respectively. In certain embodiments, the antibody or antigen-binding fragment comprises VH and VL
amino acid sequences as set forth in SEQ ID NOs.:102 and 106, respectively.
In some embodiments, an antibody or antigen-binding fragment is provided that comprises CDRH1, CDRH2, CDRH3, CDRL1, CDRL2, and CDRL3 amino acid sequences as set forth in SEQ ID NOs.:73-75 and 77-79, respectively. In certain embodiments, the antibody or antigen-binding fragment comprises VH and VL
amino acid sequences as set forth in SEQ ID NOs.:72 and 76, respectively.
In some embodiments, an antibody or antigen-binding fragment is provided that comprises CDRH1, CDRH2, CDRH3, CDRL1, CDRL2, and CDRL3 amino acid sequences as set forth in SEQ ID NOs.:63-65 and 67-69, respectively. In certain embodiments, the antibody or antigen-binding fragment comprises VH and VL
amino acid sequences as set forth in SEQ ID NOs.:62 and 66, respectively.
In some embodiments, an antibody or antigen-binding fragment is provided that comprises CDRH1, CDRH2, CDRH3, CDRL1, CDRL2, and CDRL3 amino acid sequences as set forth in SEQ ID NOs.:23-25 and 27-29, respectively. In certain embodiments, the antibody or antigen-binding fragment comprises VH and VL
amino acid sequences as set forth in SEQ ID NOs.:22 and 26, respectively.
In some embodiments, an antibody or antigen-binding fragment is provided that comprises CDRH1, CDRH2, CDRH3, CDRL1, CDRL2, and CDRL3 amino acid sequences as set forth in SEQ ID NOs .33-35 and 37-39, respectively In certain embodiments, the antibody or antigen-binding fragment comprises VH and VL
amino acid sequences as set forth in SEQ ID NOs.:32 and 36, respectively.

In some embodiments, an antibody or antigen-binding fragment is provided that comprises CDRH1, CDRH2, CDRH3, CDRL1, CDRL2, and CDRL3 amino acid sequences as set forth in SEQ ID NOs.:53-55 and 57-59, respectively. In certain embodiments, the antibody or antigen-binding fragment comprises VH and VL
amino acid sequences as set forth in SEQ ID NOs.:52 and 56, respectively.
In some embodiments, an antibody or antigen-binding fragment is provided that comprises CDRH1, CDRH2, CDRH3, CDRL1, CDRL2, and CDRL3 amino acid sequences as set forth in SEQ ID NOs.:363-365 and 367-369, respectively. In certain embodiments, the antibody or antigen-binding fragment comprises VH and VL
amino acid sequences as set forth in SEQ ID NOs.:362 and 366, respectively.
In certain embodiments, an antibody or an antigen-binding fragment of the present disclosure comprises a CDRH1, a CDRH2, a CDRH3, a CDRL1, a CDRL2, and a CDRL3, wherein each CDR is independently selected from a corresponding CDR
of Antibody 418_i, Antibody 4182, Antibody 4183, Antibody 4184, Antibody 4185, Antibody 4186, Antibody 4187, Antibody 4188, Antibody 418_9, Antibody 41810, Antibody 418 11, Antibody 41812, Antibody 41813, Antibody 41814, Antibody 41815, Antibody 41816, Antibody 41817, Antibody 41818, Antibody 41819, Antibody 41820, Antibody 41821, Antibody 41822, Antibody 41823, Antibody 41824, Antibody 41825, Antibody 41826, Antibody 41827, Antibody 41828, Antibody 418_29, Antibody 418_30, Antibody 41831, Antibody 418_33, Antibody 41834, Antibody 41835, Antibody 41837, Antibody 41838, Antibody 41839, Antibody 41840, Antibody 41841, Antibody 41842, Antibody 41843, or Antibody 4 18 44, as provided in Table I . That is, all combinations of CDRs from SARS-CoV-2 mAbs and the variant sequences thereof provided in Table 1 are contemplated.
Antibody 418 1 is also referred to herein as S2X28. Antibody 418 2 is also referred to herein as S2X303. Antibody 418 3 is also referred to herein as S2X320.
Antibody 418_4 is also referred to herein as S2X333 Antibody 418_5 is also referred to herein as S2M28. Antibody 418 6 is also referred to herein as S2M24 or S2M24v2.
Antibody 418 7 is also referred to herein as S2L7. Antibody 4 [8 8 is also referred to herein as S2L24. Antibody 418 9 is also referred to herein as S2L28. Antibody 418 10 is also referred to herein as S2X310. Antibody 418 11 is also referred to herein as S2X94. Antibody 418_12 is also referred to herein as S2X169. Antibody 418 13 is also referred to herein as S2L11. Antibody 418_14 is also referred to herein as S2L12.
Antibody 418 15 is also referred to herein as S2X186. Antibody 418 16 is also referred to herein as S2X175. Antibody 418 17 is also referred to herein as S2X170.
Antibody 418 18 is also referred to herein as S2X125. Antibody 418_19 is also referred to herein as S2X107. Antibody 418 20 is also referred to herein as S2X105.
Antibody 418 21 is also referred to herein as S2X102. Antibody 418_22 is also referred to herein as S2X15. Antibody 418 23 is also referred to herein as S2X49.
Antibody 418 24 is also referred to herein as S2X51. Antibody 418 25 is also referred to herein as S2X72. Antibody 418 26 is also referred to herein as S2X91.
Antibody 418 27 is also referred to herein as S2X98. Antibody 418 28 is also referred to herein as S2X124. Antibody 418 29 is also referred to herein as S2X158. Antibody 418 is also referred to herein as S2X161. Antibody 418_31 is also referred to herein as S2X165. Antibody 418_33 is also referred to herein as S2X173. Antibody 418_34 is also referred to herein as S2X176. Antibody 418 35 is also referred to herein as S2X316. Antibody 418 37 is also referred to herein as S2X90. Antibody 418 38 is also referred to herein as S2X93. Antibody 418_39 is also referred to herein as S2L14.
Antibody 418 40 is also referred to herein as S2L20 or S2L20v1. Antibody 418_41 is also referred to herein as S2L26. Antibody 418_42 is also referred to herein as S2L35.
Antibody 418 43 is also referred to herein as S2L38. Antibody 418 44 is also referred to herein as S2L50.
The term "CL" refers to an "immunoglobulin light chain constant region" or a "light chain constant region," i.e., a constant region from an antibody light chain. The term "CH" refers to an "immunoglobulin heavy chain constant region" or a "heavy chain constant region," which is further divisible, depending on the antibody isotype into CHL CH2, and CH3 (IgA, IgD, IgG), or CHL CH2, CH3, and CH4 domains (IgE, IgM) The Fc region of an antibody heavy chain is described further herein In any of the presently disclosed embodiments, an antibody or antigen-binding fragment of the present disclosure comprises any one or more of CL, a CHI, a CH2, and a CH3.
In certain embodiments, a CL comprises an amino acid sequence having 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to the amino acid sequence of SEQ ID NO. :8 or SEQ ID NO.: 9. In certain embodiments, a CH1-CH2-CH3 comprises an amino acid sequence having 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to the amino acid sequence of SEQ ID NO. :6 or SEQ
ID NO.:7.
It will be understood that, for example, production in a mammalian cell line can remove one or more C-terminal lysine of an antibody heavy chain (see, e.g., Liu et al.
mAbs 6(5):1145-1154 (2014)). Accordingly, an antibody or antigen-binding fragment of the present disclosure can comprise a heavy chain, a CH1-CH3, a CH3, or an Fc polypeptide wherein a C-terminal lysine residue is present or is absent; in other words, encompassed are embodiments where the C-terminal residue of a heavy chain, a CH3, or an Fc polypeptide is not a lysine (e.g., is a glycine), and embodiments where a lysine is the C-terminal residue. In certain embodiments, a composition comprises a plurality of an antibody and/or an antigen-binding fragment of the present disclosure, wherein one or more antibody or antigen-binding fragment does not comprise a lysine residue at the C-terminal end of the heavy chain, CH1-CH3, or Fc polypeptide, and wherein one or more antibody or antigen-binding fragment comprises a lysine residue at the C-terminal end of the heavy chain, CH1-CH3, or Fc polypeptide.
A "Fab" (fragment antigen binding) is the part of an antibody that binds to antigens and includes the variable region and CH1 of the heavy chain linked to the light chain via an inter-chain disulfide bond. Each Fab fragment is monovalent with respect to antigen binding, i.e., it has a single antigen-binding site. Pepsin treatment of an antibody yields a single large F(ab')2 fragment that roughly corresponds to two disulfide linked Fab fragments having divalent antigen-binding activity and is still capable of cross-linking antigen. Both the Fab and F(ab')2 are examples of "antigen-binding fragments." Fab' fragments differ from Fab fragments by having additional few residues at the carboxy terminus of the CH1 domain including one or more cysteines from the antibody hinge region Fab'-SH is the designation herein for Fab' in which the cysteine residue(s) of the constant domains bear a free thiol group. F(ab')2 antibody fragments originally were produced as pairs of Fab' fragments that have hinge cysteines between them. Other chemical couplings of antibody fragments are also known.

Fab fragments may be joined, e.g., by a peptide linker, to form a single chain Fab, also referred to herein as "scFab." In these embodiments, an inter-chain disulfide bond that is present in a native Fab may not be present, and the linker serves in full or in part to link or connect the Fab fragments in a single polypeptide chain. A
heavy chain-derived Fab fragment (e.g., comprising, consisting of, or consisting essentially of VH +
CH1, or "Fd") and a light chain-derived Fab fragment (e.g., comprising, consisting of, or consisting essentially of VL + CL) may be linked in any arrangement to form a scFab. For example, a scFab may be arranged, in N-terminal to C-terminal direction, according to (heavy chain Fab fragment ¨ linker ¨ light chain Fab fragment) or (light chain Fab fragment ¨ linker ¨ heavy chain Fab fragment). Peptide linkers and exemplary linker sequences for use in scFabs are discussed in further detail herein.
"Fv" is a small antibody fragment that contains a complete antigen-recognition and antigen-binding site. This fragment generally consists of a dimer of one heavy- and one light-chain variable region domain in tight, non-covalent association.
However, even a single variable domain (or half of an Fv comprising only three CDRs specific for an antigen) can have the ability to recognize and bind antigen, although typically at a lower affinity than the entire binding site.
"Single-chain Fv" also abbreviated as "sFv" or "scFv", are antibody fragments that comprise the VH and VL antibody domains connected into a single polypeptide chain. In some embodiments, the scFv polypeptide comprises a polypeptide linker disposed between and linking the Vu and VL domains that enables the scFv to retain or form the desired structure for antigen binding. Such a peptide linker can be incorporated into a fusion polypeptide using standard techniques well known in the art.
For a review of scFv, see Pluckthun in The Pharmacology of Monoclonal Antibodies, vol. 113, Rosenburg and Moore eds., Springer-Verlag, New York, pp. 269-315 (1994);
Borrebaeck 1995, infra. In certain embodiments, the antibody or antigen-binding fragment comprises a scFv comprising a VH domain, a VL domain, and a peptide linker linking the VH domain to the VL domain In particular embodiments, a scFv comprises a VH domain linked to a VL domain by a peptide linker, which can be in a VH-linker-VL orientation or in a VL-linker-VH orientation. Any scFv of the present disclosure may be engineered so that the C-terminal end of the VL domain is linked by a short peptide sequence to the N-terminal end of the VH domain, or vice versa (i.e., (N)VL(C)-linker-(N)VH(C) or (N)VH(C)-linker-(N)VL(C). Alternatively, in some embodiments, a linker may be linked to an N-terminal portion or end of the VH
domain, the VL domain, or both.
Peptide linker sequences may be chosen, for example, based on: (1) their ability to adopt a flexible extended conformation; (2) their inability or lack of ability to adopt a secondary structure that could interact with functional epitopes on the first and second polypeptides and/or on a target molecule; and/or (3) the lack or relative lack of hydrophobic or charged residues that might react with the polypeptides and/or target molecule. Other considerations regarding linker design (e.g., length) can include the conformation or range of conformations in which the VH and VL can form a functional antigen-binding site. In certain embodiments, peptide linker sequences contain, for example, Gly, Asn and Ser residues. Other near neutral amino acids, such as Thr and Ala, may also be included in a linker sequence. Other amino acid sequences which may be usefully employed as linker include those disclosed in Maratea et al., Gene 40:39 46 (1985); Murphy et al., Proc. Natl. Acad. Sci. USA 83:8258 8262 (1986); U.S.
Pat. No.
4,935,233, and U.S. Pat. No. 4,751,180. Other illustrative and non-limiting examples of linkers may include, for example, Glu-Gly-Lys-Ser-Ser-Gly-Ser-Gly-Ser-Glu-Ser-Lys-Val-Asp (SEQ ID NO: 19) (Chaudhary et al., Proc. Natl. Acad. Sci. USA 87:1066-1070 (1990)) and Lys-Glu-Ser-Gly-Ser-Val-Ser-Ser-Glu-Gln-Leu-Ala-Gln-Phe-Arg-Ser-Leu-Asp (SEQ ID NO: 20) (Bird et al., Science 242:423-426 (1988)) and the pentamer Gly-Gly-Gly-Gly-Ser (SEQ ID NO: 21) when present in a single iteration or repeated Ito 5 or more times, or more; see, e.g., SEQ ID NO: 17. Any suitable linker may be used, and in general can be about 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 15 23, 24, 25, 26, 27, 28, 29, 30, 40, 50, 60, 70, 80, 90, 100 amino acids in length, or less than about 200 amino acids in length, and will preferably comprise a flexible structure (can provide flexibility and room for conformational movement between two regions, domains, motifs, fragments, or modules connected by the linker), and will preferably be biologically inert and/or have a low risk of immunogenicity in a human. Exemplary linkers include those comprising or consisting of the amino acid sequence set forth in any one or more of SEQ ID NOs: 10-21.
In certain embodiments, the linker comprises or consists of an amino acid sequence having at least 75% (i.e., at least about 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or more) identity to the amino acid sequence set forth in any one of SEQ ID NOs: 10-21.
scFvs can be constructed using any combination of the VH and VL sequences or any combination of the CDRH1, CDRH2, CDRH3, CDRL1, CDRL2, and CDRL3 sequences disclosed herein.
In some embodiments, linker sequences are not required; for example, when the first and second polypeptides have non-essential N-terminal amino acid regions that can be used to separate the functional domains and prevent steric interference.
During antibody development, DNA in the germline variable (V), joining (J), and diversity (D) gene loci may be rearranged and insertions and/or deletions of nucleotides in the coding sequence may occur. Somatic mutations may be encoded by the resultant sequence, and can be identified by reference to a corresponding known germline sequence. In some contexts, somatic mutations that are not critical to a desired property of the antibody (e.g., binding to a SARS-CoV-2 antigen), or that confer an undesirable property upon the antibody (e.g., an increased risk of immunogenicity in a subject administered the antibody), or both, may be replaced by the corresponding germline-encoded amino acid, or by a different amino acid, so that a desirable property of the antibody is improved or maintained and the undesirable property of the antibody is reduced or abrogated. Thus, in some embodiments, the antibody or antigen-binding fragment of the present disclosure comprises at least one more germline-encoded amino acid in a variable region as compared to a parent antibody or antigen-binding fragment, provided that the parent antibody or antigen binding fragment comprises one or more somatic mutations. Variable region and CDR
amino acid sequences of exemplary anti-SARS-CoV-2 antibodies of the present disclosure are provided in Table 1 herein.
In certain embodiments, an antibody or antigen-binding fragment comprises an amino acid modification (e.g., a substitution mutation) to remove an undesired risk of oxidation, deamidation, and/or isomerization.

Also provided herein are variant antibodies that comprise one or more amino acid alterations in a variable region (e.g., VH, VL, framework or CDR) as compared to a presently disclosed ("parent") antibody, wherein the variant antibody is capable of binding to a SARS-CoV-2 antigen.
In certain embodiments, the VH comprises or consists of an amino acid sequence having at least 85% (i.e., 85%, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, or 100%) identity to the amino acid sequence according to any one of SEQ ID
NOs.: 22, 32, 42, 52, 62, 72, 82, 92, 102, 112, 122, 132, 142, 152, 162, 172, 182 192, 202, 212, 222, 232, 242, 252, 262, 272, 282, 292, 302, 312, 322, 332, 342, 352, 362, 372, 382, 392, 402, 412, 422, or 432, wherein the variation is optionally limited to one or more framework regions and/or the variation comprises one or more substitution to a germline-encoded amino acid, and/or (ii) the VL comprises or consists of an amino acid sequence having at least 85% (i.e., 85%, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, or 100%) identity to the amino acid sequence according to any one of SEQ ID
NOs.: 26, --------------------------------------------------------------------36, 46, 56, 66, 76, 86, 96, 106, 116, 126, 136, 146, 156, 166, 176, 186, 196, 206, 216, 226, 236, 246, 256, 266, 276, 286, 296, 306, 316, 326, 336, 346, 356, 366, 376, 386, 396, 406, 416, 426, or 436, wherein the variation is optionally limited to one or more framework regions and/or the variation comprises one or more substitution to a germline-encoded amino acid.
In certain embodiments, the VH comprises or consists of any VH amino acid sequence set forth in Table 1, and the VL comprises or consists of any VL
amino acid sequence set forth in Table 1. In particular embodiments, the VH and the VL
comprise amino acid sequences having at least have at least 85% (i.e., 85%, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, or 100%) identity to, or comprise or consist of, the amino acid sequences according to SEQ ID NOs.. (i) 22 and 26, respectively, (ii) 32 and 36, respectively; (iii) 42 and 46, respectively; (iv) 52 and 56, respectively; (v) 62 and 66, respectively; (vi) 72 and 76, respectively; (vii) 82 and 86, respectively; (viii) 92 and 96, respectively; (ix) 102 and 106, respectively; (x) 112 and 116, respectively; (xi) 122 and 126, respectively, (xii) 132 and 136, respectively, (xiii) 142 and 146, respectively; (xiv) 152 and 156, respectively; (xv) 162 and 166, respectively;
(xvi) 172 and 176, respectively; (xvii) 182 and 186, respectively; (xviii) 192 and 196, respectively; (xix) 202 and 206, respectively; (xx) 212 and 216, respectively;
(xxi) 222 and 226, respectively, (xxii) 232 and 236, respectively, (xxiii) 242 and 246, respectively; (xxiv) 252 and 256, respectively; (xxv) 262 and 266, respectively; (xxvi) 272 and 276, respectively, (xxvii) 282 and 286, respectively, (xxviii) 292 and 296, respectively; (xxix) 302 and 306, respectively; (xxx) 312 and 316, respectively; (xxxi) 322 and 326, respectively; (xxxii) 332 and 336, respectively; (xxxiii) 342 and 346, respectively, (xxxiv) 352 and 356, respectively, (xxxv) 362 and 366, respectively, (xxxvi) 372 and 376, respectively; (xxxvii) 382 and 386, respectively;
(xxxviii) 392 and 396, respectively; (xxxix) 402 and 406, respectively; (xxxx) 412 and 416, respectively;
(xxxxi) 422 and 426, respectively; or (xxxxii) 432 and 436, respectively.
In certain embodiments, an antibody or antigen-binding fragment of the present disclosure is monospecific (e.g., binds to a single epitope) or is multispecific (e.g., binds to multiple epitopes and/or target molecules). Antibodies and antigen binding fragments may be constructed in various formats. Exemplary antibody formats are disclosed in Spiess et al., Mol. Immunol. 67(2):95 (2015), and in Brinkmann and Kontermann, mAbs 9(2).182-212 (2017), which formats and methods of making the same are incorporated herein by reference and include, for example, Bispecific T cell Engagers (BiTEs), DARTs, Knobs-Into-Holes (KIH) assemblies, scFv-CH3-KIH
assemblies, KIH Common Light-Chain antibodies, TandAbs, Triple Bodies, TriBi Minibodies, Fab-scFv, scFv-CH-CL-scFv, F(ab')2-scFv2, tetravalent HCabs, Intrabodies, CrossMabs, Dual Action Fabs (DAFs) (two-in-one or four-in-one), DutaMabs, DT-IgG, Charge Pairs, Fab-arm Exchange, SEEDbodies, Triomabs, LUZ-Y
assemblies, Fcabs, la-bodies, orthogonal Fabs, DVD-Igs (e.g., US Patent No.
8,258,268, which formats are incorporated herein by reference in their entirety), IgG(H)-scFv, scFv-(H)IgG, IgG(L)-scFv, scFv-(L)IgG, IgG(L,H)-Fv, IgG(H)-V, V(H)-IgG, IgG(L)-V, V(L)-IgG, KIH IgG-scFab, 2scFv-IgG, IgG-2scFv, scFv4-Ig, Zybody, and DVI-IgG (four-in-one), as well as so-called FIT-Ig (e.g., PCT Publication No. WO
2015/103072, which formats are incorporated herein by reference in their entirety), so-called WuxiBody formats (e.g., PCT Publication No. WO 2019/057122, which formats are incorporated herein by reference in their entirety), and so-called In-Elbow-Insert Ig formats (IEI-Ig; e.g., PCT Publication Nos. WO 2019/024979 and WO 2019/025391, which formats are incorporated herein by reference in their entirety).
In certain embodiments, the antibody or antigen-binding fragment comprises two or more of VH domains, two or more VL domains, or both (i.e., two or more VH
domains and two or more VL domains). In particular embodiments, an antigen-binding fragment comprises the format (N-terminal to C-terminal direction) VH-linker-VL-linker-VH-linker-VL, wherein the two VH sequences can be the same or different and the two VL sequences can be the same or different. Such linked scFvs can include any combination of VH and VL domains arranged to bind to a given target, and in formats comprising two or more VH and/or two or more VL, one, two, or more different eptiopes or antigens may be bound. It will be appreciated that formats incorporating multiple antigen-binding domains may include VH and/or VL sequences in any combination or orientation. For example, the antigen-binding fragment can comprise the format VL-linker-VH-linker-VL-linker-VH, VH-linker-VL-linker-VL-linker-VH, or VL-linker-VH-linker-VH-linker-VL.
Monospecific or multispecific antibodies or antigen-binding fragments of the present disclosure constructed comprise any combination of the VH and VL
sequences and/or any combination of the CDRH1, CDRH2, CDRH3, CDRL1, CDRL2, and CDRL3 sequences disclosed herein. A bispecific or multispecific antibody or antigen-binding fragment may, in some embodiments, comprise one, two, or more antigen-binding domains (e.g., a VH and a VL) of the instant disclosure. Two or more binding domains may be present that bind to the same or a different SARS-CoV-2 epitope, and a bispecific or multi specific antibody or antigen-binding fragment as provided herein can, in some embodiments, comprise a further SARS-CoV-2 binding domain, and/or can comprise a binding domain that binds to a different antigen or pathogen altogether.
In any of the presently disclosed embodiments, the antibody or antigen-binding fragment can be multispecific; e.g., bispecific, trispecific, or the like.
In certain embodiments, the antibody or antigen-binding fragment comprises.
(i) a first VH and a first VL; and (ii) a second VH and a second VL, wherein the first VH
and the second VH are different and each independently comprise an amino acid sequence having at least 85% (i.e., 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) identity to the amino acid sequence set forth in any one of SEQ ID NOs.: 22, 32, 42, 52, 62, 72, 82, 92, 102, 112, 122, 132, 142, 152, 162, 172, 182 192, 202, 212, 222, 232, 242, 252, 262, 272, 282, 292, 302, 312, 322, 332, 342, 352, 362, 372, 382, 392, 402, 412, 422, or 432, and wherein the first VL and the second VL are different and each independently comprise an amino acid sequence having at least 85% (i.e., 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) identity to the amino acid sequence set forth in any one of SEQ ID NOs.: 26, 36, 46, 56, 66, 76, 86, 96, 106, 116, 126, 136, 146, 156, 166, 176, 186, 196, 206, 216, 226, 236, 246, 256, 266, 276, 286, 296, 306, 316, 326, 336, 346, 356, 366, 376, 386, 396, 406, 416, 426, or 436, and wherein the first VH and the first VL together form a first antigen-binding site, and wherein the second VH and the second VL together form a second antigen-binding site.
In certain embodiments, the antibody or antigen-binding fragment comprises:
(i) a first VH and a first VL; and (ii) a second VH and a second VL, wherein the first VH
comprises an amino acid sequence having at least 85% (i.e., 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) identity to the amino acid sequence set forth in SEQ ID NO: 52 and the first VL comprises an amino acid sequence haying at least 85% (i.e., 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) identity to the amino acid sequence set forth in SEQ ID NO: 56; and a) the second VH comprises an amino acid sequence having at least 85% (i.e., 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) identity to the amino acid sequence set forth in SEQ ID NO: 442 and the second VL comprises an amino acid sequence having at least 85% (i.e., 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) identity to the amino acid sequence set forth in SEQ ID NO:
446;
b) the second VH comprises an amino acid sequence having at least 85% (i.e., 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) identity to the amino acid sequence set forth in SEQ ID NO: 450 and the second VL comprises an amino acid sequence having at least 85% (i.e., 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) identity to the amino acid sequence set forth in SEQ ID NO: 454; or c) the second VH comprises an amino acid sequence having at least 85% (i.e., 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) identity to the amino acid sequence set forth in SEQ ID NO: 458 and the second VL comprises an amino acid sequence having at least 85% (i.e., 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) identity to the amino acid sequence set forth in SEQ ID NO: 462; and wherein the first VH and the first VL together form a first antigen-binding site, and wherein the second VH and the second VL
together form a second antigen-binding site.
In certain embodiments, the antibody or antigen-binding fragment comprises a Fc polypeptide, or a fragment thereof. The "Fc" fragment or Fc polypeptide comprises the carboxy-terminal portions (i.e., the CH2 and CH3 domains of IgG) of both antibody H chains held together by disulfides. Antibody "effector functions" refer to those biological activities attributable to the Fc region (a native sequence Fc region or amino acid sequence variant Fc region) of an antibody, and vary with the antibody isotype.
Examples of antibody effector functions include: Clq binding and complement dependent cytotoxicity; Fc receptor binding; antibody-dependent cell-mediated cytotoxicity (ADCC), phagocytosis, down regulation of cell surface receptors (e.g., B
cell receptor); and B cell activation. As discussed herein, modifications (e.g., amino acid substitutions) may be made to an Fc domain in order to modify (e.g., improve, reduce, or ablate) one or more functionality of an Fc-containing polypeptide (e.g., an antibody of the present disclosure). Such functions include, for example, Fc receptor (FcR) binding, antibody half-life modulation (e.g., by binding to FcRn), ADCC
function, protein A binding, protein G binding, and complement binding. Amino acid modifications that modify (e.g., improve, reduce, or ablate) Fc functionalities include, for example, the T250Q/M428L, M252Y/S254T/T256E, H433K/N434F, M428L/N434S, E233P/L234V/L235A/G236 + A327G/A330S/P331S, E333A, S239D/A330L/I332E, P2571/Q311, K326W/E333S, S239D/I332E/G236A, N297Q, K322A, S228P, L235E + E318A/K320A/K322A, L234A/L235A (also referred to herein as "LALA"), and L234A/L235A/P329G mutations, which mutations are summarized and annotated in "Engineered Fc Regions", published by InvivoGen (2011) and available online at invivogen.com/PDF/review/review-Engineered-Fc-Regions-invivogen.pdf?utm source=review&utm medium=pdf&utm campaign=review&utm content=Engineered-Fc-Regions, and are incorporated herein by reference. Unless the context indicates otherwise, Fc amino acid residues are numbered herein according to the EU numbering system.
For example, to activate the complement cascade, the Clq protein complex can bind to at least two molecules of IgG1 or one molecule of IgM when the immunoglobulin molecule(s) is attached to the antigenic target (Ward, E. S., and Ghetie, V., Ther. Immunol 2 (1995) 77-94). Burton, D. R., described (Mol.
Immtmol.
22 (1985) 161-206) that the heavy chain region comprising amino acid residues 318 to 337 is involved in complement fixation. Duncan, A. R., and Winter, G. (Nature (1988) 738-740), using site directed mutagenesis, reported that Glu318, Lys320 and Lys322 form the binding site to Clq. The role of Glu318, Lys320 and Lys 322 residues in the binding of Clq was confirmed by the ability of a short synthetic peptide containing these residues to inhibit complement mediated lysis.
For example, FcR binding can be mediated by the interaction of the Fc moiety (of an antibody) with Fc receptors (FcRs), which are specialized cell surface receptors on cells including hematopoietic cells. Fe receptors belong to the immunoglobulin superfamily, and shown to mediate both the removal of antibody-coated pathogens by phagocytosis of immune complexes, and the lysis of erythrocytes and various other cellular targets (e.g. tumor cells) coated with the corresponding antibody, via antibody dependent cell mediated cytotoxicity (ADCC; Van de Winkel, J. G., and Anderson, C.
L., J. Leukoc. Biol. 49 (1991) 511-524). FcRs are defined by their specificity for immunoglobulin classes; Fc receptors for IgG antibodies are referred to as FcyR, for IgE as Fcall, for IgA as FcaR and so on and neonatal Fc receptors are referred to as FcRn. Fc receptor binding is described for example in Ravetch, J. V., and Kinet, J. P., Annu. Rev. Immunol. 9 (1991) 457-492; Capel, P. J., et al., Immunomethods 4 (1994) 25-34; de Haas, M., et al., J Lab. Cl/n. Med. 126 (1995) 330-341; and Gessner, J. E., et al., Ann. Hematol. 76 (1998) 231-248 Cross-linking of receptors by the Fc domain of native IgG antibodies (FcyR) triggers a wide variety of effector functions including phagocytosis, antibody-dependent cellular cytotoxicity, and release of inflammatory mediators, as well as immune complex clearance and regulation of antibody production. Fc moieties providing cross-linking of receptors (e.g., FcyR) are contemplated herein. In humans, three classes of FcyR have been characterized to-date, which are: (i) FcyRI (CD64), which binds monomeric IgG with high affinity and is expressed on macrophages, monocytes, neutrophils and eosinophils; (ii) FcyRII (CD32), which binds complexed IgG
with medium to low affinity, is widely expressed, in particular on leukocytes, is believed to be a central player in antibody-mediated immunity, and which can be divided into FcyRITA, FcyRII,B and FeyRITC, which perform different functions in the immune system, but bind with similar low affinity to the IgG-Fe, and the ectodomains of these receptors are highly homologuous; and (iii) FcyRIII (CD16), which binds IgG
with medium to low affinity and has been found in two forms: FeyRITIA, which has been found on NK cells, macrophages, eosinophils, and some monocytes and T cells, and is believed to mediate ADCC; and FcyRIII,B, which is highly expressed on neutrophils.
FcyRITA is found on many cells involved in killing (e.g. macrophages, monocytes, neutrophils) and seems able to activate the killing process. FcyRIM
seems to play a role in inhibitory processes and is found on B-cells, macrophages and on mast cells and eosinophils. Importantly, it has been shown that 75% of all FcyRIIB
is found in the liver (Ganesan, L. P. et al., 2012: "FcyRIIb on liver sinusoidal endothelium clears small immune complexes," Journal of Immunology 189: 4981-4988). FeyMIB is abundantly expressed on Liver Sinusoidal Endothelium, called LSEC, and in Kupffer cells in the liver and LSEC are the major site of small immune complexes clearance (Ganesan, L. P. et al., 2012: FcyRIIb on liver sinusoidal endothelium clears small immune complexes. Journal of Immunology 189: 4981-4988).
In some embodiments, the antibodies disclosed herein and the antigen-binding fragments thereof comprise an Fc polypeptide or fragment thereof for binding to FeyRIIb, in particular an Fc region, such as, for example IgG-type antibodies.

Moreover, it is possible to engineer the Fc moiety to enhance FcyRIM binding by introducing the mutations S267E and L328F as described by Chu, S. Y. et al., 2008:
Inhibition of B cell receptor-mediated activation of primary human B cells by coengagement of CD 19 and FcgammaRII13 with Fc-engineered antibodies.
Molecular Immunology 45,3926-3933. Thereby, the clearance of immune complexes can be enhanced (Chu, S., et al., 2014: Accelerated Clearance of IgE In Chimpanzees Is Mediated By Xmab7195, An Fc-Engineered Antibody With Enhanced Affinity For Inhibitory Receptor FcyRIIb. Am J Respir Crit, American Thoracic Society International Conference Abstracts). In some embodiments, the antibodies of the present disclosure, or the antigen binding fragments thereof, comprise an engineered Fc moiety with the mutations S267E and L328F, in particular as described by Chu, S. Y. et al., 2008: Inhibition of B cell receptor-mediated activation of primary human B cells by coengagement of CD19 and FcgammaRIIb with Fc-engineered antibodies. Molecular Immunology 45, 3926-3933.
On B cells, FcyRIIB may function to suppress further immunoglobulin production and isotype switching to, for example, the IgE class. On macrophages, FcyRIIB is thought to inhibit phagocytosis as mediated through FcyRIIA. On eosinophils and mast cells, the B form may help to suppress activation of these cells through IgE binding to its separate receptor.
Regarding FcyRI binding, modification in native IgG of at least one of E233-G236, P238, D265, N297, A327 and P329 reduces binding to FcyRI. IgG2 residues at positions 233-236, substituted into corresponding positions IgG1 and IgG4, reduces binding of IgG1 and IgG4 to FcyRI by 103-fold and eliminated the human monocyte response to antibody-sensitized red blood cells (Armour, K. L., et al. Eur. J.
1111117111101.
29(1999) 2613-2624) Regarding FcyRII binding, reduced binding for FcyRIIA is found, e.g., for IgG
mutation of at least one of E233-G236, P238, D265, N297, A327, P329, D270, Q295, A327, R292 and K4I4.
Two allelic forms of human FcyRIIA are the "H131" variant, which binds to IgG1 Fc with high affinity, and the "R131" variant, which binds to IgG1 Fc with low affinity. See, e.g., Bruhns et al., Blood/13:3716-3725 (2009).
Regarding FcyRIII binding, reduced binding to FcyRIIIA is found, e.g., for mutation of at least one of E233-G236, P238, D265, N297, A327, P329, D270, Q295, A327, S239, E269, E293, Y296, V303, A327, K338 and D376. Mapping of the binding sites on human IgG 1 for Fc receptors, the above-mentioned mutation sites, and methods for measuring binding to FcyRI and FeyRIIA, are described in Shields, R. L., et al., J.
Biol. Chem. 276 (2001) 6591-6604.
Two allelic forms of human FcyRIIIA are the "F158" variant, which binds to IgG1 Fc with low affinity, and the "V158" variant, which binds to IgG1 Fc with high affinity. See, e.g., Bruhns et al., Blood/13:3716-3725 (2009).
Regarding binding to FcyRII, two regions of native IgG Fc appear to be involved in interactions between FcyRIIs and IgGs, namely (i) the lower hinge site of IgG Fc, in particular amino acid residues L, L, G, G (234 ¨ 237, EU
numbering), and (ii) the adjacent region of the CH2 domain of IgG Fc, in particular a loop and strands in the upper CH2 domain adjacent to the lower hinge region, e.g. in a region of (Wines, B.D., et al., J. Immunol. 2000; 164: 5313 ¨5318). Moreover, FcyR1 appears to bind to the same site on IgG Fc, whereas FcRn and Protein A bind to a different site on IgG Fc, which appears to be at the CH2-CH3 interface (Wines, B.D., et al., J.
Immunol.
2000; 164: 5313 ¨ 5318).
Also contemplated are mutations that increase binding affinity of an Fc polypeptide or fragment thereof of the present disclosure to a (i.e., one or more) Fcy receptor (e.g., as compared to a reference Fe polypeptide or fragment thereof or containing the same that does not comprise the mutation(s)). See, e.g., Delillo and Raveteh, Cell 161(5):1035-1045 (2015) and Ahmed et al., J. Struc. Biol.
194(1):78 (2016), the Fc mutations and techniques of which are incorporated herein by reference.
In any of the herein disclosed embodiments, an antibody or antigen-binding fragment can comprise a Fc polypeptide or fragment thereof comprising a mutation selected from G236A; S239D; A330L; and 1332E; or a combination comprising any two or more of the same; e.g., S239D/I332E; S239D/A330L/I332E;
G236A/S239D/I332E; G236A/A330L/I332E (also referred to herein as "GAALIE"), or G236A/S239D/A330L/I332E. In some embodiments, the Fc polypeptide or fragment thereof does not comprise S239D. In some embodiments, the Fc polypeptide or fragment thereof comprises S at position 239 (EU numbering) In certain embodiments, the Fc polypeptide or fragment thereof may comprise or consist of at least a portion of an Fc polypeptide or fragment thereof that is involved in binding to FcRn binding. In certain embodiments, the Fc polypeptide or fragment thereof comprises one or more amino acid modifications that improve binding affinity for (e.g., enhance binding to) FcRn (e.g., at a pH of about 6.0) and, in some embodiments, thereby extend in vivo half-life of a molecule comprising the Fc polypeptide or fragment thereof (e.g., as compared to a reference Fc polypeptide or fragment thereof or antibody that is otherwise the same but does not comprise the modification(s)). In certain embodiments, the Fc polypeptide or fragment thereof comprises or is derived from a IgG Fc and a half-life-extending mutation comprises any one or more of: M428L; N434S; N434H; N434A; N434S; M252Y; S254T; T256E;
T250Q; P257I Q31 11; D376V; T307A; E380A (EU numbering). In certain embodiments, a half-life-extending mutation comprises M428L/N434S (also referred to herein as "MLNS"). In certain embodiments, a half-life-extending mutation comprises M252Y/S254T/T256E. In certain embodiments, a half-life-extending mutation comprises T250Q/M428L. In certain embodiments, a half-life-extending mutation comprises P257I/Q3111. In certain embodiments, a half-life-extending mutation comprises P257I/N434H. In certain embodiments, a half-life-extending mutation comprises D376V/N434H. In certain embodiments, a half-life-extending mutation comprises T307A/E380A/N434A.
In some embodiments, an antibody or antigen-binding fragment includes a Fc moiety that comprises the substitution mtuations M428L/N434S. In some embodiments, an antibody or antigen-binding fragment includes a Fc polypeptide or fragment thereof that comprises the substitution mtuations G236A/A330L/I332E.
In certain embodiments, an antibody or antigen-binding fragment includes a (e.g., IgG) Fc moiety that comprises a G236A mutation, an A330L mutation, and a 1332E
mutation (GAALIE), and does not comprise a S239D mutation (e.g., comprises a native S
at position 239). In particular embodiments, an antibody or antigen-binding fragment includes an Fc polypeptide or fragment thereof that comprises the substitution mutations: M428L/N434S and G236A/A330L/1332E, and optionally does not comprise S239D (e.g., comprises S at 239) In certain embodiments, an antibody or antigen-binding fragment includes a Fc polypeptide or fragment thereof that comprises the substitution mutations: M428L/N434S and G236A/S239D/A330L/1332E.

In certain embodiments, the antibody or antigen-binding fragment comprises a mutation that alters glycosylation, wherein the mutation that alters glycosylation comprises N297A, N297Q, or N297G, and/or the antibody or antigen-binding fragment is partially or fully aglycosylated and/or is partially or fully afucosylated.
Host cell lines and methods of making partially or fully aglycosylated or partially or fully afucosylated antibodies and antigen-binding fragments are known (see, e.g., PCT
Publication No. WO 2016/181357; Suzuki et al. Clin. Cancer Res. 13(6):1875-82 (2007); Huang et al. MAbs 6:1-12 (2018)).
In certain embodiments, the antibody or antigen-binding fragment is capable of eliciting continued protection in vivo in a subject even once no detectable levels of the antibody or antigen-binding fragment can be found in the subject (i.e., when the antibody or antigen-binding fragment has been cleared from the subject following administration). Such protection is referred to herein as a vaccinal effect.
Without wishing to be bound by theory, it is believed that dendritic cells can internalize complexes of antibody and antigen and thereafter induce or contribute to an endogenous immune response against antigen. In certain embodiments, an antibody or antigen-binding fragment comprises one or more modifications, such as, for example, mutations in the Fc comprising G236A, A330L, and 1332E, that are capable of activating dendritic cells that may induce, e.g., T cell immunity to the antigen.
In any of the presently disclosed embodiments, the antibody or antigen-binding fragment comprises a Fc polypeptide or a fragment thereof, including a CH2 (or a fragment thereof, a CH3 (or a fragment thereof), or a CH2 and a CH3, wherein the CH2, the CH3, or both can be of any isotype and may contain amino acid substitutions or other modifications as compared to a corresponding wild-type CH2 or CH3, respectively. In certain embodiments, a Fc polypeptide of the present disclosure comprises two CH2-CH3 polypeptides that associate to form a dimer.
In any of the presently disclosed embodiments, the antibody or antigen-binding fragment can be monoclonal The term "monoclonal antibody" (mAb) as used herein refers to an antibody obtained from a population of substantially homogeneous antibodies, i.e., individual antibodies comprising the population are identical except for possible naturally occurring mutations that may be present, in some cases in minor amounts. Monoclonal antibodies are highly specific, being directed against a single antigenic site. Furthermore, in contrast to polyclonal antibody preparations that include different antibodies directed against different epitopes, each monoclonal antibody is directed against a single epitope of the antigen. In addition to their specificity, the monoclonal antibodies are advantageous in that they may be synthesized uncontaminated by other antibodies. The term "monoclonal" is not to be construed as requiring production of the antibody by any particular method. For example, monoclonal antibodies useful in the present invention may be prepared by the hybridoma methodology first described by Kohler et al., Nature 256:495 (1975), or may be made using recombinant DNA methods in bacterial, eukaryotic animal, or plant cells (see, e.g., U.S. Pat. No. 4,816,567). Monoclonal antibodies may also be isolated from phage antibody libraries using the techniques described in Clackson et al., Nature, 352:624-628 (1991) and Marks et Mol Biol., 222:581-597 (1991), for example.
Monoclonal antibodies may also be obtained using methods disclosed in PCT
Publication No. WO 2004/076677A2.
Antibodies and antigen-binding fragments of the present disclosure include "chimeric antibodies" in which a portion of the heavy and/or light chain is identical with or homologous to corresponding sequences in antibodies derived from a particular species or belonging to a particular antibody class or subclass, while the remainder of the chain(s) is identical with or homologous to corresponding sequences in antibodies derived from another species or belonging to another antibody class or subclass, as well as fragments of such antibodies, so long as they exhibit the desired biological activity (see, U.S. Pat. Nos. 4,816,567; 5,530,101 and 7,498,415; and Morrison et al., Proc.
Natl. Acad. Sci. USA, 81:6851-6855 (1984)). For example, chimeric antibodies may comprise human and non-human residues. Furthermore, chimeric antibodies may comprise residues that are not found in the recipient antibody or in the donor antibody.
These modifications are made to further refine antibody performance. For further details, see Jones et al., Nature 321:522-525 (1986); Riechmann etal., Nature 332:323-329 (1988); and Presta, C 117T . Op. StrucL Biol. 2:593-596 (1992). Chimeric antibodies also include primatized and humanized antibodies.

A "humanized antibody" is generally considered to be a human antibody that has one or more amino acid residues introduced into it from a source that is non-human.
These non-human amino acid residues are typically taken from a variable domain.
Humanization may be performed following the method of Winter and co-workers (Jones et at., Nature, 321:522-525 (1986); Reichmann et at., Nature, 332:323-(1988); Verhoeyen et al., Science, 239:1534-1536 (1988)), by substituting non-human variable sequences for the corresponding sequences of a human antibody.
Accordingly, such "humanized" antibodies are chimeric antibodies (U.S. Pat. Nos. 4,816,567;

5,530,101 and 7,498,415) wherein substantially less than an intact human variable domain has been substituted by the corresponding sequence from a non-human species.
In some instances, a -humanized" antibody is one which is produced by a non-human cell or animal and comprises human sequences, e.g., Hc domains.
A "human antibody" is an antibody containing only sequences that are present in an antibody that is produced by a human (i.e., sequences that are encoded by human antibody-encoding genes). However, as used herein, human antibodies may comprise residues or modifications not found in a naturally occurring human antibody (e.g., an antibody that is isolated from a human), including those modifications and variant sequences described herein. These are typically made to further refine or enhance antibody performance. In some instances, human antibodies are produced by transgenic animals. For example, see U.S. Pat. Nos. 5,770,429; 6,596,541 and 7,049,426.
In certain embodiments, an antibody or antigen-binding fragment of the present disclosure is chimeric, humanized, or human.
Polynucleotides, Vectors, and Host cells In another aspect, the present disclosure provides isolated polynucleotides that encode any of the presently disclosed antibodies or an antigen-binding fragment thereof, or a portion thereof (e.g., a CDR, a VH, a VL, a heavy chain, or a light chain).
In certain embodiments, the polynucleotide is codon-optimized for expression in a host cell. Once a coding sequence is known or identified, codon optimization can be performed using known techniques and tools, e.g., using the GenScript OptimiumGeneTM tool; see also Scholten et at., Cl/n. 1111171111101 119:135, 2006).

Codon-optimized sequences include sequences that are partially codon-optimized (i.e., one or more codon is optimized for expression in the host cell) and those that are fully codon-optimized.
It will also be appreciated that polynucleotides encoding antibodies and antigen-binding fragments of the present disclosure may possess different nucleotide sequences while still encoding a same antibody or antigen-binding fragment due to, for example, the degeneracy of the genetic code, splicing, and the like.
In certain embodiments, the polynucleotide comprises a polynucleotide having at least 50% (i.e., 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) identity to the polynucleotide sequence according to any one or more of SEQ ID NOs.:30, 31, 40, 41, 50, 51, 60, 61, 70, 71, 80, 81, 90, 91, 100, 101, 110, 111, 120, 121, 130, 131, 140, 141, 150, 151, 160, 161, 170, 171, 180, 181, 190, 191, 200, 201, 210, 211, 220, 221, 230, 231, 240, 241, 250, 251, 260, 261, 270, 271, 280, 281, 290, 291, 300, 301, 310, 311, 320, 321, 330, 331, 340, 341, 350, 351, 360, 361, 370, 371, 380, 381, 390, 391, 400, 401, 410, 411, 420, 421, 430, 431, 440, and 441, or any combination thereof (e.g., a polynucleotide comprises a polynucleotide having at least 50% identity to to SEQ ID NO. :30 and a polynucleotide having at least 50% identity to SEQ ID NO. :31).
It will be appreciated that in certain embodiments, a polynucleotide encoding an antibody or antigen-binding fragment is comprised in a polynucleotide that includes other sequences and/or features for, e.g-., expression of the antibody or antigen-binding fragment in a host cell. Exemplary features include a promoter sequence, a polyadenylation sequence, a sequence that encodes a signal peptide (e.g., located at the N-terminus of a expressed antibody heavy chain or light chain), or the like.
In any of the presently disclosed embodiments, the polynucleotide can comprise deoxyribonucleic acid (DNA) or ribonucleic acid (RNA). In some embodiments, the RNA comprises messenger RNA (mRNA).
Vectors are also provided, wherein the vectors comprise or contain a polynucleotide as disclosed herein (e.g., a polynucleotide that encodes an antibody or antigen-binding fragment that binds to SARS-CoV-2). A vector can comprise any one or more of the vectors disclosed herein. In particular embodiments, a vector is provided that comprises a DNA plasmid construct encoding the antibody or antigen-binding fragment, or a portion thereof (e.g., so-called "DMAb"; see, e.g., Muthumani et al., J
infect Dis. 214(3):369-378 (2016); Muthumani et al., Hum Vaccin Immunother 9:2253-2262 (2013)); Flingai et al., Sci Rep. 5:12616 (2015); and Elliott et al., NPJ
Vaccines 18 (2017), which antibody-coding DNA constructs and related methods of use, including administration of the same, are incorporated herein by reference).
In certain embodiments, a DNA plasmid construct comprises a single open reading frame encoding a heavy chain and a light chain (or a VH and a VL) of the antibody or antigen-binding fragment, wherein the sequence encoding the heavy chain and the sequence encoding the light chain are optionally separated by polynucleotide encoding a protease cleavage site and/or by a polynucleotide encoding a self-cleaving peptide. In some embodiments, the substituent components of the antibody or antigen-binding fragment are encoded by a polynucleotide comprised in a single plasmid. In other embodiments, the substituent components of the antibody or antigen-binding fragment are encoded by a polynucleotide comprised in two or more plasmids (e.g., a first plasmid comprises a polynucleotide encoding a heavy chain, VH, or VH+CH, and a second plasmid comprises a polynucleotide encoding the cognate light chain, VL, or VL+CL). In certain embodiments, a single plasmid comprises a polynucleotide encoding a heavy chain and/or a light chain from two or more antibodies or antigen-binding fragments of the present disclosure. An exemplary expression vector is pVaxl, available from Invitrogen . A DNA plasmid of the present disclosure can be delivered to a subject by, for example, electroporation (e.g., intramuscular electroporation), or with an appropriate formulation (e.g., hyaluronidase).
In a further aspect, the present disclosure also provides a host cell expressing an antibody or antigen-binding fragment according to the present disclosure; or comprising or containing a vector or polynucleotide according the present disclosure.
Examples of such cells include but are not limited to, eukaryotic cells, e.g., yeast cells, animal cells, insect cells, plant cells; and prokaryotic cells, including E. coli In some embodiments, the cells are mammalian cells. In certain such embodiments, the cells are a mammalian cell line such as CHO cells (e.g., DHFR- CHO cells (Urlaub et al., PNAS 77:4216 (1980)), human embryonic kidney cells (e.g., HEK293T cells), PER.C6 cells, YO cells, Sp2/0 cells. NSO cells, human liver cells, e.g. Hepa RG cells, myeloma cells or hybridoma cells. Other examples of mammalian host cell lines include mouse sertoli cells (e.g., TNI4 cells); monkey kidney CV1 line transformed by SV40 (COS-7); baby hamster kidney cells (BHK); African green monkey kidney cells (VERO-76); monkey kidney cells (CV1); human cervical carcinoma cells (BELA);
human lung cells (W138); human liver cells (Hep G2); canine kidney cells (MDCK;
buffalo rat liver cells (BRL 3A); mouse mammary tumor (MMT 060562); TRI
cells; MRC 5 cells; and FS4 cells. Mammalian host cell lines suitable for antibody production also include those described in, for example, Yazaki and Wu, Methods in Molecular Biology, Vol. 248 (B. K. C. Lo, ed., Humana Press, Totowa, N.J.), pp. 255-268 (2003).
In certain embodiments, a host cell is a prokaryotic cell, such as an E. coll.
The expression of peptides in prokaryotic cells such as E. coil is well established (see, e.g., Pluckthun, A. Bio/Technology 9:545-551 (1991). For example, antibodies may be produced in bacteria, in particular when glycosylation and Fc effector function are not needed. For expression of antibody fragments and polypeptides in bacteria, see, e.g., U.S. Pat. Nos. 5,648,237, 5,789,199, and 5,840,523.
In particular embodiments, the cell may be transfected with a vector according to the present description with an expression vector. The term "transfection"
refers to the introduction of nucleic acid molecules, such as DNA or RNA (e.g. mRNA) molecules, into cells, such as into eukaryotic cells. In the context of the present description, the term "transfection" encompasses any method known to the skilled person for introducing nucleic acid molecules into cells, such as into eukaryotic cells, including into mammalian cells. Such methods encompass, for example, electroporation, lipofection, e.g., based on cationic lipids and/or liposomes, calcium phosphate precipitation, nanoparticle based transfection, virus based transfection, or transfection based on cationic polymers, such as DEAE-dextran or polyethylenimine, etc In certain embodiments, the introduction is non-viral Moreover, host cells of the present disclosure may be transfected stably or transiently with a vector according to the present disclosure, e.g. for expressing an antibody, or an antigen-binding fragment thereof, according to the present disclosure.

In such embodiments, the cells may be stably transfected with the vector as described herein. Alternatively, cells may be transiently transfected with a vector according to the present disclosure encoding an antibody or antigen-binding fragment as disclosed herein. In any of the presently disclosed embodiments, a polynucleotide may be heterologous to the host cell.
Accordingly, the present disclosure also provides recombinant host cells that heterologously express an antibody or antigen-binding fragment of the present disclosure. For example, the cell may be of a species that is different to the species from which the antibody was fully or partially obtained (e.g., CHO cells expressing a human antibody or an engineered human antibody). In some embodiments, the cell type of the host cell does not express the antibody or antigen-binding fragment in nature Moreover, the host cell may impart a post-translational modification (PTM, e.g., glysocylation or fucosylation) on the antibody or antigen-binding fragment that is not present in a native state of the antibody or antigen-binding fragment (or in a native state of a parent antibody from which the antibody or antigen binding fragment was engineered or derived) Such a PTM may result in a functional difference (e.g., reduced immunogenicity). Accordingly, an antibody or antigen-binding fragment of the present disclosure that is produced by a host cell as disclosed herein may include one or more post-translational modification that is distinct from the antibody (or parent antibody) in its native state (e.g., a human antibody produced by a CHO cell can comprise a more post-translational modification that is distinct from the antibody when isolated from the human and/or produced by the native human B cell or plasma cell).
Insect cells useful expressing a binding protein of the present disclosure are known in the art and include, for example, ,S'podoptera frugipera Sf9 cells, Trichoplusia ni BTI-TN5B1-4 cells, and Spodoptera fruppera SfSWTO1 "Mimi CTM" cells. See, e.g., Palmberger et at., J. Biotechnot /53(3-4):160-166 (2011). Numerous baculoviral strains have been identified which may be used in conjunction with insect cells, particularly for transfecti on of S'podopterafrugiperda cells Eukaryotic microbes such as filamentous fungi or yeast are also suitable hosts for cloning or expressing protein-encoding vectors, and include fungi and yeast strains with "humanized" glycosylation pathways, resulting in the production of an antibody with a partially or fully human glycosylation pattern. See Gerngross, Nat.
Biotech. 22:1409-1414 (2004); Li et al., Nat. Biotech. 24:210-215 (2006).
Plant cells can also be utilized as hosts for expressing a binding protein of the present disclosure. For example, PLANTIBODIESTm technology (described in, for example, U.S. Pat. Nos. 5,959,177; 6,040,498; 6,420,548; 7,125,978; and 6,417,429) employs transgenic plants to produce antibodies.
In certain embodiments, the host cell comprises a mammalian cell. In particular embodiments, the host cell is a CHO cell, a HEK293 cell, a PER.C6 cell, a YO
cell, a Sp2/0 cell, a NSO cell, a human liver cell, a myeloma cell, or a hybridoma cell.
In a related aspect, the present disclosure provides methods for producing an antibody, or antigen-binding fragment, wherein the methods comprise culturing a host cell of the present disclosure under conditions and for a time sufficient to produce the antibody, or the antigen-binding fragment. Methods useful for isolating and purifying recombinantly produced antibodies, by way of example, may include obtaining supernatants from suitable host cell/vector systems that secrete the recombinant antibody into culture media and then concentrating the media using a commercially available filter. Following concentration, the concentrate may be applied to a single suitable purification matrix or to a series of suitable matrices, such as an affinity matrix or an ion exchange resin. One or more reverse phase HPLC steps may be employed to further purify a recombinant polypeptide. These purification methods may also be employed when isolating an immunogen from its natural environment. Methods for large scale production of one or more of the isolated/recombinant antibody described herein include batch cell culture, which is monitored and controlled to maintain appropriate culture conditions. Purification of soluble antibodies may be performed according to methods described herein and known in the art and that comport with laws and guidelines of domestic and foreign regulatory agencies.
Compositions Also provided herein are compositions that comprise any one or more of the presently disclosed antibodies, antigen-binding fragments, polynucleotides, vectors, or host cells, singly or in any combination, and can further comprise a pharmaceutically acceptable carrier, excipient, or diluent. Carriers, excipients, and diluents are discussed in further detail herein.
In certain embodiments, a composition comprises a plurality of an antibody and/or an antigen-binding fragment of the present disclosure, wherein one or more antibody or antigen-binding fragment does not comprise a lysine residue at the C-terminal end of the heavy chain, CH1-CH3, or Fc polypeptide, and wherein one or more antibody or antigen-binding fragment comprises a lysine residue at the C-terminal end of the heavy chain, CH1-CH3, or Fc polypeptide.
In certain embodiments, a composition comprises two or more different antibodies or antigen-binding fragments according to the present disclosure.
In certain embodiments, antibodies or antigen-binding fragments to be used in a combination each independently have one or more of the following characteristics neutralize naturally occurring SARS-CoV-2 variants; do not compete with one another for Spike protein binding; bind distinct Spike protein epitopes; have a reduced formation of resistance to SARS-CoV-2; when in a combination, have a reduced formation of resistance to SARS-CoV-2; potently neutralize live SARS-CoV-2 virus, exhibit additive or synergistic effects on neutralization of live SARS-CoV-2 virus when used in combination; exhibit effector functions; are protective in relevant animal model(s) of infection; are capable of being produced in sufficient quantities for large-scale production In certain embodiments, a composition comprises (a) antibody S2X333 (or an antigen-binding fragment thereof) or an antibody or antigen-binding fragment that competes with antibody S2X333 for SARS-CoV-2 S protein binding and (b) antibody S309 (or an antigen-binding fragment thereof) or an antibody or antigen-binding fragment that competes with antibody S309 for SARS-CoV-2 S protein binding.
In certain embodiments, a composition comprises (a) antibody S2X333 (or an antigen-binding fragment thereof) or an antibody or antiben-binding fragment that competes with antibody S2X333 for SARS-CoV-2 S protein binding and (11) antibody S2E12 (or an antigen-binding fragment thereof) or an antibody or antigen-binding fragment that competes with antibody S2E12 for SARS-CoV-2 S protein binding.

In certain embodiments, a composition comprises (a) antibody S2X333 (or an antigen-binding fragment thereof) or an antibody or antigen-binding fragment that competes with antibody S2X333 for SARS-CoV-2 S protein binding and (b) antibody S2M11 (or an antigen-binding fragment thereof) or an antibody or antigen-binding fragment that competes with antibody S2M11 for SARS-CoV-2 S protein binding.
Antibody S2X333 comprises the VH amino acid sequence of SEQ ID NO. :52 and the VL amino acid sequence of SEQ ID NO. :56.
Antibody S2E12 comprises the VH amino acid sequence of SEQ ID NO.450 and the VL amino acid sequence of SEQ ID NO.454.
Antibody S309 comprises the VH amino acid sequence of SEQ ID NO. :442 and the VL amino acid sequence of SEQ ID NO. :446. A variant VH of antibody S309 comprises the amino acid sequence of SEQ ID NO. 466.
Antibody S2M11 comprises the VH amino acid sequence of SEQ ID NO.458 and the VL amino acid sequence of SEQ ID NO. :462.
In certain embodiments, a composition comprises two or more different antibodies or antigen-binding fragments according to the present disclosure.
In certain embodiments, a composition comprises a first vector comprising a first plasmid, and a second vector comprising a second plasmid, wherein the first plasmid comprises a polynucleotide encoding a heavy chain, VH, or VH+CH, and a second plasmid comprises a polynucleotide encoding the cognate light chain, VL, or VL+CL of the antibody or antigen-binding fragment thereof. In certain embodiments, a composition comprises a polynucleotide (e.g., mRNA) coupled to a suitable delivery vehicle or carrier. Exemplary vehicles or carriers for administration to a human subject include a lipid or lipid-derived delivery vehicle, such as a liposome, solid lipid nanoparticle, oily suspension, submicron lipid emulsion, lipid microbubble, inverse lipid micelle, cochlear liposome, lipid microtubule, lipid microcylinder, or lipid nanoparticle (LNP) or a nanoscale platform (see, e.g., Li et at. Wilery Interdi.5cip Rev.
Nanomed Nanohiotechnol. 1/(2).e1530 (2019)). Principles, reagents, and techniques for designing appropriate mRNA and and formulating mRNA-LNP and delivering the same are described in, for example, Pardi et at. (I Control Release 2/7345-351 (2015));
Thess et al. (Mol Ther 23: 1456-1464 (2015)); Thran et al. (EMBO Mol Med 9(10):1434-1448 (2017); Kose et al. (Sc. Immunol. 4 eaaw6647 (2019); and Sabnis et at. (Mol. Ther. 26:1509-1519 (2018)), which techniques, include capping, codon optimization, nucleoside modification, purification of mRNA, incorporation of the mRNA into stable lipid nanoparticles (e.g., ionizable cationic lipid/phosphatidylcholine/cholesterol/PEG-lipid; ionizable lipid:distearoyl PC:cholesterol:polyethylene glycol lipid), and subcutaneous, intramuscular, intradermal, intravenous, intraperitoneal, and intratracheal administration of the same, are incorporated herein by reference.
Methods and Uses Also provided herein are methods for use of an antibody or antigen-binding fragment, nucleic acid, vector, cell, or composition of the present disclosure in the detection or diagnosis of SARS-CoV-2 infection (e.g., in a human subject, or in a sample obtained from a human subject).
Methods of diagnosis (e.g., in vitro, ex vivo) may include contacting an antibody or antibody fragment (e.g., antigen binding fragment) with a sample. Such a sample may be isolated from a subject, for example an isolated (e.g., fluid, tissue, or secretion) sample from a nasal passage, a sinus cavity, a salivary gland, a lung, a liver, a trachea, a bronchiole, a pancreas, a kidney, an ear, an eye, a placenta, an alimentary tract, a heart, an ovary, a pituitary gland, an adrenal, a thyroid gland, a brain, sera, plasma, skin, or blood. In some embodiments, the sample may comprise a nasal secretion, sputum, bronchial lavage, urine, stool, saliva, sweat, or any combination thereof.
Methods of diagnosis may also include the detection of an antigen/antibody complex, in particular following the contacting of an antibody or antibody fragment with a sample.
Such a detection step can be performed at the bench, i.e. without any contact to the human or animal body. Examples of detection methods are well-known to the person skilled in the art and include, e.g., ELISA (enzyme-linked immunosorbent assay), including direct, indirect, and sandwich ELISA.
Also provided herein are methods of treating a subject using an antibody or antigen-binding fragment of the present disclosure, or a composition comprising the same, wherein the subject has, is believed to have, or is at risk for having an infection by SARS-CoV-2. "Treat," "treatment," or "ameliorate" refers to medical management of a disease, disorder, or condition of a subject (e.g., a human or non-human mammal, such as a primate, horse, cat, dog, goat, mouse, or rat). In general, an appropriate dose or treatment regimen comprising an antibody or composition of the present disclosure is administered in an amount sufficient to elicit a therapeutic or prophylactic benefit.
Therapeutic or prophylactic/preventive benefit includes improved clinical outcome;
lessening or alleviation of symptoms associated with a disease; decreased occurrence of symptoms, improved quality of life, longer disease-free status, diminishment of extent of disease, stabilization of disease state; delay or prevention of disease progression;
remission; survival; prolonged survival; or any combination thereof. In certain embodiments, therapeutic or prophylactic/preventive benefit includes reduction or prevention of hospitalization for treatment of a SARS-CoV-2 infection (i.e., in a statistically significant manner) In certain embodiments, therapeutic or prophylactic/preventive benefit includes a reduced duration of hospitalization for treatment of a SARS-CoV-2 infection (i.e., in a statistically significant manner). In certain embodiments, therapeutic or prophylactic/preventive benefit includes a reduced or abrogated need for respiratory intervention, such as intubation and/or the use of a respirator device. In certain embodiments, therapeutic or prophylactic/preventive benefit includes reversing a late-stage disease pathology and/or reducing mortality.
A "therapeutically effective amount" or "effective amount" of an antibody, antigen-binding fragment, polynucleotide, vector, host cell, or composition of this disclosure refers to an amount of the composition or molecule sufficient to result in a therapeutic effect, including improved clinical outcome, lessening or alleviation of symptoms associated with a disease; decreased occurrence of symptoms; improved quality of life; longer disease-free status; diminishment of extent of disease, stabilization of disease state, delay of disease progression, remission, survival, or prolonged survival in a statistically significant manner. When referring to an individual active ingredient, administered alone, a therapeutically effective amount refers to the effects of that ingredient or cell expressing that ingredient alone When referring to a combination, a therapeutically effective amount refers to the combined amounts of active ingredients or combined adjunctive active ingredient with a cell expressing an active ingredient that results in a therapeutic effect, whether administered serially, sequentially, or simultaneously. A combination may comprise, for example, two different antibodies that specifically bind a SARS-CoV-2 antigen, which in certain embodiments, may be the same or different SARS-CoV-2 antigen, and/or can comprise the same or different epitopes.
Accordingly, in certain embodiments, methods are provided for treating a SARS-CoV-2 infection in a subject, wherein the methods comprise administering to the subject an effective amount of an antibody, antigen-binding fragment, polynucleotide, vector, host cell, or composition as disclosed herein.
Subjects that can be treated by the present disclosure are, in general, human and other primate subjects, such as monkeys and apes for veterinary medicine purposes.
Other model organisms, such as mice and rats, may also be treated according to the present disclosure. In any of the aforementioned embodiments, the subject may be a human subject. The subjects can be male or female and can be any suitable age, including infant, juvenile, adolescent, adult, and geriatric subjects.
A number of criteria are believed to contribute to high risk for severe symptoms or death associated with a SARS CoV-2 infection These include, but are not limited to, age, occupation, general health, pre-existing health conditions, and lifestyle habits. In some embodiments, a subject treated according to the present disclosure comprises one or more risk factors.
In certain embodiments, a human subject treated according to the present disclosure is an infant, a child, a young adult, an adult of middle age, or an elderly person. In certain embodiments, a human subject treated according to the present disclosure is less than 1 year old, or is 1 to 5 years old, or is between 5 and 125 years old (e.g., 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 105, 110, 115, or 125 years old, including any and all ages therein or therebetween). In certain embodiments, a human subject treated according to the present disclosure is 0-19 years old, 20-44 years old, 45-54 years old, 55-64 years old, 65-74 years old, 75-84 years old, or 85 years old, or older Persons of middle, and especially of elderly age are believed to be at particular risk. In particular embodiments, the human subject is 45-54 years old, 55-64 years old, 65-74 years old, 75-84 years old, or 85 years old, or older.

In some embodiments, the human subject is biologically male. In some embodiments, the human subject is biologically female.
In certain embodiments, a human subject treated according to the present disclosure is a resident of a nursing home or a long-term care facility, is a hospice care worker, is a healthcare provider or healthcare worker, is a first responder, is a family member or other close contact of a subject diagnosed with or suspected of having a SARS-CoV-2 infection, is overweight or clinically obese, is or has been a smoker, has or had chronic obstructive pulmonary disease (COPD), is asthmatic (e.g., having moderate to severe asthma), has an autoimmune disease or condition (e.g., diabetes), and/or has a compromised or depleted immune system (e.g., due to AIDS/HIV
infection, a cancer such as a blood cancer, a lymphodepleting therapy such as a chemotherapy, a bone marrow or organ transplantation, or a genetic immune condition), has chronic liver disease, has cardiovascular disease, has a pulmonary or heart defect, works or otherwise spends time in close proximity with others, such as in a factory, shipping center, hospital setting, or the like.
In certain embodiments, a subject treated according to the present disclosure has received a vaccine for SARS-CoV-2 and the vaccine is determined to be ineffective, e.g., by post-vaccine infection or symptoms in the subject, by clinical diagnosis or scientific or regulatory consensus.
In certain embodiments, treatment is administered as pen -exposure prophylaxis In certain embodiments, treatment is administered to a subject with mild-to-moderate disease, which may be in an outpatient setting. In certain embodiments, treatment is administered to a subject with moderate-to-severe disease, such as requiring hospitalization.
Typical routes of administering the presently disclosed compositions thus include, without limitation, oral, topical, transdermal, inhalation, parenteral, sublingual, buccal, rectal, vaginal, and intranasal. The term "parenteral", as used herein, includes subcutaneous injections, intravenous, intramuscular, intrasternal injection or infusion techniques. In certain embodiments, administering comprises administering by a route that is selected from oral, intravenous, parenteral, intragastric, intrapleural, intrapulmonary, intrarectal, intradermal, intraperitoneal, intratumoral, subcutaneous, topical, transdermal, intracisternal, intrathecal, intranasal, and intramuscular. In particular embodiments, a method comprises orally administering the antibody, antigen-binding fragment, polynucleotide, vector, host cell, or composition to the subject.
Pharmaceutical compositions according to certain embodiments of the present invention are formulated so as to allow the active ingredients contained therein to be bioavailable upon administration of the composition to a patient. Compositions that will be administered to a subject or patient may take the form of one or more dosage units, where for example, a tablet may be a single dosage unit, and a container of a herein described an antibody or antigen-binding in aerosol form may hold a plurality of dosage units. Actual methods of preparing such dosage forms are known, or will be apparent, to those skilled in this art; for example, see Remington: The Science and Practice of Pharmacy, 20th Edition (Philadelphia College of Pharmacy and Science, 2000). The composition to be administered will, in any event, contain an effective amount of an antibody or antigen-binding fragment, polynucleotide, vector, host cell_ or composition of the present disclosure, for treatment of a disease or condition of interest in accordance with teachings herein.
A composition may be in the form of a solid or liquid. In some embodiments, the carrier(s) are particulate, so that the compositions are, for example, in tablet or powder form. The carrier(s) may be liquid, with the compositions being, for example, an oral oil, injectable liquid or an aerosol, which is useful in, for example, inhalatory administration. When intended for oral administration, the pharmaceutical composition is preferably in either solid or liquid form, where semi solid, semi liquid, suspension and gel forms are included within the forms considered herein as either solid or liquid As a solid composition for oral administration, the pharmaceutical composition may be formulated into a powder, granule, compressed tablet, pill, capsule, chewing gum, wafer or the like. Such a solid composition will typically contain one or more inert diluents or edible carriers. In addition, one or more of the following may be present. binders such as carboxymethylcellulose, ethyl cellulose, microcrystalline cellulose, gum tragacanth or gelatin; excipients such as starch, lactose or dextrins, disintegrating agents such as alginic acid, sodium alginate, Primogel, corn starch and the like; lubricants such as magnesium stearate or Sterotex; glidants such as colloidal silicon dioxide; sweetening agents such as sucrose or saccharin; a flavoring agent such as peppermint, methyl salicylate or orange flavoring; and a coloring agent.
When the composition is in the form of a capsule, for example, a gelatin capsule, it may contain, in addition to materials of the above type, a liquid carrier such as polyethylene glycol or Oil.
The composition may be in the form of a liquid, for example, an elixir, syrup, solution, emulsion or suspension. The liquid may be for oral administration or for delivery by injection, as two examples. When intended for oral administration, preferred compositions contain, in addition to the present compounds, one or more of a sweetening agent, preservatives, dye/colorant and flavor enhancer. In a composition intended to be administered by injection, one or more of a surfactant, preservative, wetting agent, dispersing agent, suspending agent, buffer, stabilizer and isotonic agent may be included.
Liquid pharmaceutical compositions, whether they be solutions, suspensions or other like form, may include one or more of the following adjuvants: sterile diluents such as water for injection, saline solution, preferably physiological saline, Ringer's solution, isotonic sodium chloride, fixed oils such as synthetic mono or diglycerides which may serve as the solvent or suspending medium, polyethylene glycols, glycerin, propylene glycol or other solvents; antibacterial agents such as benzyl alcohol or methyl paraben; antioxidants such as ascorbic acid or sodium bi sulfite; chelating agents such as ethylenediaminetetraacetic acid; buffers such as acetates, citrates or phosphates and agents for the adjustment of tonicity such as sodium chloride or dextrose. The parenteral preparation can be enclosed in ampoules, disposable syringes or multiple dose vials made of glass or plastic. Physiological saline is a preferred adjuvant. An injectable pharmaceutical composition is preferably sterile.
A liquid composition intended for either parenteral or oral administration should contain an amount of an antibody or antigen-binding fragment as herein disclosed such that a suitable dosage will be obtained Typically, this amount is at least 001% of the antibody or antigen-binding fragment in the composition. When intended for oral administration, this amount may be varied to be between 0.1 and about 70% of the weight of the composition. Certain oral pharmaceutical compositions contain between about 4% and about 75% of the antibody or antigen-binding fragment. In certain embodiments, pharmaceutical compositions and preparations according to the present invention are prepared so that a parenteral dosage unit contains between 0.01 to 10% by weight of antibody or antigen-binding fragment prior to dilution.
The composition may be intended for topical administration, in which case the carrier may suitably comprise a solution, emulsion, ointment or gel base. The base, for example, may comprise one or more of the following: petrolatum, lanolin, polyethylene glycols, bee wax, mineral oil, diluents such as water and alcohol, and emulsifiers and stabilizers. Thickening agents may be present in a composition for topical administration. If intended for transdermal administration, the composition may include a transdermal patch or iontophoresis device. The pharmaceutical composition may be intended for rectal administration, in the form, for example, of a suppository, which will melt in the rectum and release the drug. The composition for rectal administration may contain an oleaginous base as a suitable nonirritating excipient.
Such bases include, without limitation, lanolin, cocoa butter and polyethylene glycol.
A composition may include various materials which modify the physical form of a solid or liquid dosage unit. For example, the composition may include materials that form a coating shell around the active ingredients. The materials that form the coating shell arc typically inert, and may be selected from, for example, sugar, shellac, and other enteric coating agents. Alternatively, the active ingredients may be encased in a gelatin capsule. The composition in solid or liquid form may include an agent that binds to the antibody or antigen-binding fragment of the disclosure and thereby assists in the delivery of the compound. Suitable agents that may act in this capacity include monoclonal or polyclonal antibodies, one or more proteins or a liposome. The composition may consist essentially of dosage units that can be administered as an aerosol. The term aerosol is used to denote a variety of systems ranging from those of colloidal nature to systems consisting of pressurized packages. Delivery may be by a liquefied or compressed gas or by a suitable pump system that dispenses the active ingredients. Aerosols may be delivered in single phase, bi phasic, or tri phasic systems in order to deliver the active ingredient(s). Delivery of the aerosol includes the necessary container, activators, valves, subcontainers, and the like, which together may form a kit. One of ordinary skill in the art, without undue experimentation, may determine preferred aerosols.
It will be understood that compositions of the present disclosure also encompass carrier molecules for polynucleotides, as described herein (e.g., lipid nanoparticles, nanoscale delivery platforms, and the like).
The pharmaceutical compositions may be prepared by methodology well known in the pharmaceutical art. For example, a composition intended to be administered by injection can be prepared by combining a composition that comprises an antibody, antigen-binding fragment thereof, or antibody conjugate as described herein and optionally, one or more of salts, buffers and/or stabilizers, with sterile, distilled water so as to form a solution. A surfactant may be added to facilitate the formation of a homogeneous solution or suspension. Surfactants are compounds that non-covalently interact with the peptide composition so as to facilitate dissolution or homogeneous suspension of the antibody or antigen-binding fragment thereof in the aqueous delivery system.
In general, an appropriate dose and treatment regimen provide the composition(s) in an amount sufficient to provide therapeutic and/or prophylactic benefit (such as described herein, including an improved clinical outcome (e.g., a decrease in frequency, duration, or severity of diarrhea or associated dehydration, or inflammation, or longer disease-free and/or overall survival, or a lessening of symptom severity). For prophylactic use, a dose should be sufficient to prevent, delay the onset of, or diminish the severity of a disease associated with disease or disorder.
Prophylactic benefit of the compositions administered according to the methods described herein can be determined by performing pre-clinical (including in vitro and in vivo animal studies) and clinical studies and analyzing data obtained therefrom by appropriate statistical, biological, and clinical methods and techniques, all of which can readily be practiced by a person skilled in the art.
Compositions are administered in an effective amount (e.g., to treat a Wuhan coronavirus infection), which will vary depending upon a variety of factors including the activity of the specific compound employed; the metabolic stability and length of action of the compound; the age, body weight, general health, sex, and diet of the subject; the mode and time of administration; the rate of excretion; the drug combination; the severity of the particular disorder or condition; and the subject undergoing therapy. In certain embodiments, tollowing administration of therapies according to the formulations and methods of this disclosure, test subjects will exhibit about a 10% up to about a 99% reduction in one or more symptoms associated with the disease or disorder being treated as compared to placebo-treated or other suitable control subjects.
Generally, a therapeutically effective daily dose of an antibody or antigen binding fragment is (for a 70 kg mammal) from about 0.001 mg/kg (i.e., 0.07 mg) to about 100 mg/kg (i.e., 7.0 g); preferably a therapeutically effective dose is (for a 70 kg mammal) from about 0.01 mg/kg (i.e., 0.7 mg) to about 50 mg/kg (i.e., 3.5 g);
more preferably a therapeutically effective dose is (for a 70 kg mammal) from about 1 mg/kg (i.e., 70 mg) to about 25 mg/kg (i.e., 1.75 g). For polynucleotides, vectors, host cells, and related compositions of the present disclosure, a therapeutically effective dose may be different than for an antibody or antigen-binding fragment.
In certain embodiments, a method comprises administering the antibody, antigen-binding fragment, polynucleotide, vector, host cell, or composition to the subject at 2, 3, 4, 5, 6, 7, 8, 9, 10 times, or more.
In certain embodiments, a method comprises administering the antibody, antigen-binding fragment, or composition to the subject a plurality of times, wherein a second or successive administration is performed at about 6, about 7, about 8, about 9, about 10, about 11, about 12, about 24, about 48, about 74, about 96 hours, or more, following a first or prior administration, respectively.
In certain embodiments, a method comprises administering the antibody, antigen-binding fragment, polynucleotide, vector, host cell, or composition at least one time prior to the subject being infected by SARS-CoV-2.
Compositions comprising an antibody, antigen-binding fragment, polynucleotide, vector, host cell, or composition of the present disclosure may also be administered simultaneously with, prior to, or after administration of one or more other therapeutic agents. Such combination therapy may include administration of a single pharmaceutical dosage formulation which contains a compound of the invention and one or more additional active agents, as well as administration of compositions comprising an antibody or antigen-binding fragment of the disclosure and each active agent in its own separate dosage formulation. For example, an antibody or antigen-binding fragment thereof as described herein and the other active agent can be administered to the patient together in a single oral dosage composition such as a tablet or capsule, or each agent administered in separate oral dosage formulations.
Similarly, an antibody or antigen-binding fragment as described herein and the other active agent can be administered to the subject together in a single parenteral dosage composition such as in a saline solution or other physiologically acceptable solution, or each agent administered in separate parenteral dosage formulations. Where separate dosage formulations are used, the compositions comprising an antibody or antigen-binding fragment and one or more additional active agents can be administered at essentially the same time, i.e., concurrently, or at separately staggered times, i.e., sequentially and in any order; combination therapy is understood to include all these regimens.
In certain embodiments, a combination therapy is provided that comprises one or more anti-SARS-CoV-2 antibody (or one or more nucleic acid, host cell, vector, or composition) of the present disclosure and one or more anti-inflammatory agent and/or one or more anti-viral agent. In particular embodiments, the one or more anti-inflammatory agent comprises a corticosteroid such as, for example, dexamethasone, predni sone, or the like. In some embodiments, the one or more anti-inflammatory agents comprise a cytokine antagonist such as, for example, an antibody that binds to IL6 (such as siltuximab), or to IL-6R (such as tocilizumab), or to IL-13, IL-7, IL-8, IL-9, IL- I 0, FGF, G-CSF, GM-CSF, IFN-y, IP- I 0, MCP-I, MIP- I A, MIP I -B, PDGR, TNF-a, or VEGF. In some embodiments, anti-inflammatory agents such as leronlimab, ruxolitinib and/or anakinra are used. In some embodiments, the one or more anti-viral agents comprise nucleotide analogs or nucelotide analog prodrugs such as, for example, remdesivir, sofosbuvir, acyclovir, and zidovudine. In particular embodiments, an anti-viral agent comprises lopinavir, ritonavir, favipiravir, or any combination thereof Other anti-inflammatory agents for use in a combination therapy of the present disclosure include non-steroidal anti-inflammatory drugs (NSAIDS). It will be appreciated that in such a combination therapy, the one or more antibody (or one or more nucleic acid, host cell, vector, or composition) and the one or more anti-inflammatory agent and/or one or the more antiviral agent can be administered in any order and any sequence, or together.
In some embodiments, an antibody (or one or more nucleic acid, host cell, vector, or composition) is administered to a subject who has previously received one or more anti-inflammatory agent and/or one or more antiviral agent. In some embodiments, one or more anti-inflammatory agent and/or one or more antiviral agent is administered to a subject who has previously received an antibody (or one or more nucleic acid, host cell, vector, or composition).
In certain embodiments, a combination therapy is provided that comprises two or more anti-SARS-CoV-2 antibodies of the present disclosure. A method can comprise administering a first antibody to a subject who has received a second antibody, or can comprise administering two or more antibodies together. For example, in particular embodiments, a method is provided that comprises administering to the subject (a) a first antibody or antigen-binding fragment, when the subject has received a second antibody or antigen-binding fragment; (b) the second antibody or antigen-binding fragment, when the subject has received the first antibody or antigen-binding fragment; or (c) the first antibody or antigen-binding fragment, and the second antibody or antigen-binding fragment.
In a related aspect, uses of the presently disclosed antibodies, antigen-binding fragments, vectors, host cells, and compositions are provided.
In certain embodiments, an antibody, antigen-binding fragment, polynucleotide, vector, host cell, or composition is provided for use in a method of treating a SARS-CoV-2 infection in a subject.
In certain embodiments, an antibody, antigen-binding fragment, or composition is provided for use in a method of manufacturing or preparing a medicament for treating a SARS-CoV-2 infection in a subject.
In certain embodiments, an antibody or antigen-binding fragment is provided for use in a method of detecting SARS-CoV-2 in a sample. In some embodiments, the method comprises contacting the sample with the antibody or antigen-binding fragment and detecting binding of the antibody or antigen-binding fragment to a SARS-CoV-2 protein or polypeptide in the sample. In some embodiments, binding to SARS-CoV-protein or polypeptide is detected by immunohistochemistry, ELISA, agglutination, immuno-dot, immuno-chromatography, and/or immuno-filtration.
In certain embodiments, an antibody or antigen-binding fragment is provided for use in a method of diagnosing a SARS-CoV-2 infection in a subject. In some embodiments, the method comprises testing a biological sample from the subject for the presence of a SARS-CoV-2 protein or polypeptide. In some embodiments, the testing comprises contacting the sample with the antibody or antigen-binding fragment and detecting binding of the antibody or antigen-binding fragment to the SARS-CoV-protein or polypeptide. In some embodiments, binding to SARS-CoV-2 protein or polypeptide is detected by immunohistochemistry, ELISA, agglutination, immuno-dot, immuno-chromatography, and/or immuno-filtration.
In some embodiments, a detection and/or diagnostic method as provided herein (such as using a disclosed antibody, antigen-binding fragment, composition, and/or kit) can provide a result within 1, 5, 10, 20, 30, 45, 60, 75, 90, or 120 minutes, or within one day, of beginning the method.
In another aspect, the present disclosure provides kits comprising materials useful for carrying out detection or diagnostic methods. In certain aspects, a kit comprising an antibody or antigen-binding fragment as described herein is provided. In some embodiments, the kit is used for detecting the presence of SARS-CoV-2 in a biological sample. In some embodiments, the kit is used for detecting the presence of a SARS-CoV-2 protein or polypeptide, for example, SARS-CoV-2 spike protein, in a biological sample. In some embodiments, the presence of a SARS-CoV-2 protein is detected by immunohistochemistry, immunoblot, ELISA, agglutination, immuno-dot, immuno-chromatography, and/or immuno-filtration. In some embodiments, the kit includes a secondary antibody detectably labeled with, for example, horseradish peroxidase (HRP), and/or instructions and/or other reagents for performing a detection method as provided herein In further aspects, a kit comprising a composition is provided, wherein the composition comprises an antibody or antigen-binding fragment as described herein and a carrier or excipient. In some embodiments, the kit is used for detecting the presence of SARS-CoV-2 in a biological sample. In some embodiments, the kit is used for detecting the presence of a SARS-CoV-2 protein or polypeptide, for example, SARS-CoV-2 spike protein, in a biological sample. In some embodiments, the presence of a SARS-CoV-2 protein is detected by immunohistochemistry, immunoblot, ELISA, agglutination, immuno-dot, immuno-chromatography, and/or immuno-filtration. In some embodiments, the kit includes a secondary antibody detectably labeled with, for example, horseradish peroxidase (I-1RP) and/or instructions and/or other reagents for performing a detection method as provided herein.
The methods for detecting the presence of a SARS-CoV-2 protein or polypeptide described herein may be performed by a diagnostic laboratory, an experimental laboratory, or a clinician, or they may be performed in-home by a caregiver or by a subject providing the sample. Provided herein are kits that can be used in one or more of these settings. Materials and reagents for characterizing biological samples and diagnosis a SARS-CoV-2 infection in a subject according to the methods herein by be assembled together as a kit. In some embodiments, a kit comprises an antibody or antigen-binding fragment according to the present disclosure and instructions for using the kit.
Kits comprising an antibody or antigen-binding fragment as described herein may futher comprise one or more substrates to anchor the antigen binding molecules, including membranes, beads, plastic tubes, or other surfaces, secondary antibodies, sample buffer, labeling buffer or reagents, wash buffers or reagents, immunodetection buffer or reagents, and detection means. In some embodiments, the kit comprises a substrate to which antibodies or antigen-binding fragments are anchored.
Protocols for using these buffers and reagents for performing different steps of the procedure may be included in the kit. The reagents may be supplied in a solid (e.g., lyophilized) or liquid form. Kits of the present disclosure may optionally comprise different containers (e.g., vial, ampoule, test tube, flask or bottle) for each individual buffer or reagent. Each component will generally be suitable as aliquoted in its respective container or provided in a concentrated form. Other containers suitable for conducting certain steps of the disclosed methods may also be provided. The individual containers of the kit a preferably maintained in close confinement for commercial sale.

In some embodiments, kits of the present disclosure further include control samples, reference samples, or any combination thereof. Instructions for using the kit, according to one or more methods of this disclosure, may comprise instructions for processing the biological sample obtained from a subject, performing the test, interpreting the results, or any combination thereof. Kits of the present disclosure may further include a notice in the form prescribed by a governmental agency (e.g., FDA) regulating the manufacture, use, or sale of pharmaceuticals or biological products.
In any of the presently disclosed embodiments, an antibody or antigen-binding fragment for use in a detection and/or diagnostic method can comprise a detectable agent. Exemplary detectable agents include enzymes (e.g., a chromogenic reporter enzyme, such as horseradish peroxidase (HRP) or an alkaline phosphatase (AP)), dyes, (e.g., cyanin dye, coumarin, rhodamine, xanthene, fluorescein or a sulfonated derivative thereof, and fluorescent proteins, including those described by Shaner et al., Nature Methods (2005)), fluorescent labels or moieties (e.g., PE, Pacific blue, Alexa fluor, APC, and FITC) DNA barcodes (e.g., ranging from five up to 75 nucleotides long), and peptide tags (e.g., Strep tag, Myc tag, His tag, Flag tag, Xpress tag, Avi tag, Calmodulin tag, Polyglutamate tag, HA tag, Nus tag, S tag, X tag, SBP tag, Softag, V5 tag, CBP, GST, MBP, GFP, Thioredoxin tag).
The present disclosure also provides the following non-limiting Embodiments.
Embodiment 1. An antibody, or antigen-binding fragment thereof, comprising a heavy chain variable domain (VH) comprising a CDRH I , a CDRH2, and a CDRH3, and a light chain variable domain (VL) comprising a CDRL1, a CDRL2, and a CDRL3, wherein:
(i) the CDRH1 comprises or consists of the amino acid sequence according to any one of SEQ ID NOs.: 53, 23, 33, 43, 63, 73, 83, 93, 103, 113, 123, 133, 143, 153, 163, 173, 183, 193, 203, 213, 223, 233, 243, 253, 263, 273, 283, 293, 303, 313, 323, 333, 343, 353, 363, 373, 383, 393, 403, 413, 423, or 433, or a sequence variant thereof comprising one, two, or three acid substitutions, one or more of which substitutions is optionally a conservative substitution and/or is a substitution to a germline-encoded amino acid, (ii) the CDRH2 comprises or consists of the amino acid sequence according to any one of SEQ ID NOs.. 54, 24, 34, 44, 64, 74, 84, 94, 104, 114, 124, 134, 144, 154, 164, 174, 184, 194, 204, 214, 224, 234, 244, 254, 264, 274, 284, 294, 304, 314, 324, 334, 344, 354, 364, 374, 384, 394, 404, 414, 424, or 434, or a sequence variant thereof comprising one, two, or three amino acid substitutions, one or more of which substitutions is optionally a conservative substitution and/or is a substitution to a germline-encoded amino acid;
(iii) the CDRH3 comprises or consists of the amino acid sequence according to any one of SEQ ID NOs.: 55, 25, 35, 45, 65, 75, 85, 95, 105, 115, 125, 135, 145, 155, 165, 175, 185, 195, 205, 215, 225, 235, 245, 255, 265, 275, 285, 295, 305, 315, 325, 335, 345, 355, 365, 375, 385, 395, 405, 415, 425, or 435, or a sequence variant thereof comprising one, two, or three amino acid substitutions, one or more of which substitutions is optionally a conservative substitution and/or is a substitution to a germline-encoded amino acid;
(iv) the CDRL1 comprises or consists of the amino acid sequence according to any one of SEQ ID NOs.: 57, 27, 37, 47, 67, 77, 87, 97, 107, 117, 127, 137, 147, 157, 167, 177, 187, 197, 207, 217, 227, 237, 247, 257, 267, 277, 287, 297, 307, 317, 327, 337, 347, 357, 367, 377, 387, 397, 407, 417, 427, or 437, or a sequence variant thereof comprising one, two, or three amino acid substitutions, one or more of which substitutions is optionally a conservative substitution and/or is a substitution to a germline-encoded amino acid;
(v) the CDRL2 comprises or consists of the amino acid sequence according to any one of SEQ ID NOs.. 58, 28, 38, 48, 68, 78, 88, 98, 108, 118, 128, 138, 148, 158, 168, 178, 188, 198, 208, 218, 228, 238, 248, 258, 268, 278, 288, 298, 308, 318, 328, 338, 348, 358, 368, 378, 388, 398, 408, 418, 428, or 438, or a sequence variant thereof comprising one, two, or three amino acid substitutions, one or more of which substitutions is optionally a conservative substitution and/or is a substitution to a germline-encoded amino acid; and/or (vi) the CDRL3 comprises or consists of the amino acid sequence according to any one of SEQ ID NOs.: 59, 29, 39, 49, 69, 79, 89, 99, 109, 119, 129, 139, 149, 159, 169, 179, 189, 199, 209, 219, 229, 239, 249, 259, 269, 279, 289, 299, 309, 319, 329, 339, 349, 359, 369, 379, 389, 399, 409, 419, 429, or 439, or a sequence variant thereof comprising having one, two, or three amino acid substitutions, one or more of which substitutions is optionally a conservative substitution and/or is a substitution to a germline-encoded amino acid, wherein the antibody or antigen binding fragment is capable of binding to a surface glycoprotein of a SARS-CoV-2, optionally when the surface glycoprotein is expressed on a cell surface of a host cell and/or on a virion.
Embodiment 2. The antibody or antigen-binding fragment of Embodiment 1, which is capable of neutralizing a SARS-CoV-2 infection in an in vitro model of infection and/or in an in vivo animal model of infection and/or in a human.
Embodiment 3. The antibody or antigen-binding fragment of any one of Embodiments 1-2, comprising CDRH1, CDRH2, CDRH3, CDRL1, CDRL2, and CDRL3 amino acid sequences according to SEQ ID NOs.:
(i) 53-55 and 57-59, respectively, (ii) 33-35 and 37-39, respectively;
(iii) 43-45 and 47-49, respectively;
(iv) 23-25 and 27-29, respectively;
(v) 63-65 and 67-69, respectively;
(vi) 73-75 and 77-79, respectively;
(vii) 83-85 and 87-89, respectively;
(viii) 93-95 and 97-99, respectively;
(ix) 103-105 and 107-109, respectively (x) 113-115 and 117-119, respectively;
(xi) 123-125 and 127-129, respectively;
(xii) 133-135 and 137-139, respectively;
(xiii) 143-145 and 147-149, respectively, (xiv) 153-155 and 157-159, respectively;
(xv) 163-165 and 167-169, respectively;

(xvi) 173-175 and 177-179, respectively;
(xvii) 183-185 and 187-189, respectively, (xviii) 193-195 and 197-199, respectively;
(xix) 203-205 and 207-209, respectively;
(xx) 213-215 and 217-219, respectively;
(xxi) 223-225 and 227-229, respectively;
(xxii) 233-235 and 237-239, respectively, (xxiii) 243-245 and 247-249, respectively;
(xxiv) 253-255 and 257-259, respectively;
(xxv) 263-265 and 267-269, respectively;
(xxvi) 273-275 and 277-279, respectively;
(xxvii) 283-285 and 287-289, respectively, (xxviii) 293-295 and 297-299, respectively;
(xxix) 303-305 and 307-309, respectively;
(xxx) 313-315 and 317-319, respectively;
(xxxi) 323-325 and 327-329, respectively;
(xxxii) 333-335 and 337-339, respectively, (xxxiii) 343-345 and 347-349, respectively;
(xxxiv) 353-355 and 357-359, respectively;
(xxxv) 363-365 and 367-369, respectively;
(xxxvi) 373-375 and 377-379, respectively;
(xxxvii) 383-385 and 387-389, respectively, (xxxviii) 393-395 and 397-399, respectively;
(xxxix) 403-405 and 407-409, respectively;
(xxxx) 413-415 and 417-419, respectively, (xxxxi) 423-425 and 427-429, respectively; or (xxxxii) 433-435 and 437-439, respectively.
Embodiment 4 The antibody or antigen-binding fragment of any one of Embodiments 1-3, wherein.
(i) the VH comprises or consists of an amino acid sequence having at least 85% identity to the amino acid sequence according to any one of SEQ ID NOs.:
52, 22, 32, 42, 62, 72, 82, 92, 102, 112, 122, 132, 142, 152, 162, 172, 182 192, 202, 212, 222, 232, 242, 252, 262, 272, 282, 292, 302, 312, 322, 332, 342, 352, 362, 372, 382, 392, 402, 412, 422, and 432, wherein the variation is optionally limited to one or more framework regions and/or the variation comprises one or more substitution to a germline-encoded amino acid; and/or (ii) the VL comprises or consists of an amino acid sequence having at least 85% identity to the amino acid sequence according to any one of SEQ ID NOs..
56, 26, 36, 46, 66, 76, 86, 96, 106, 116, 126, 136, 146, 156, 166, 176, 186, 196, 206, 216, 226, 236, 246, 256, 266, 276, 286, 296, 306, 316, 326, 336, 346, 356, 366, 376, 386, 396, 406, 416, 426, and 436, wherein the variation is optionally limited to one or more framework regions and/or the variation comprises one or more substitution to a germline-encoded amino acid Embodiment 5.
The antibody or antigen-binding fragment of any one of Embodiments 1-4, wherein the VH and the VL comprise or consist of the amino acid sequences according to SEQ ID NOs.:
(i) 52 and 56, respectively;
(ii) 32 and 36, respectively, (iii) 42 and 46, respectively;
(iv) 22 and 26, respectively;
(v) 62 and 66, respectively;
(vi) 72 and 76, respectively;
(vii) 82 and 86, respectively, (viii) 92 and 96, respectively;
(ix) 102 and 106, respectively;
(x) 112 and 116, respectively, (xi) 122 and 126, respectively;
(xii) 132 and 136, respectively;
(xiii) 142 and 146, respectively;
(xiv) 152 and 156, respectively, (xv) 162 and 166, respectively;
(xvi) 172 and 176, respectively;

(xvii) 182 and 186, respectively;
(xviii) 192 and 196, respectively;
(xix) 202 and 206, respectively;
(xx) 212 and 216, respectively;
(xxi) 222 and 226, respectively;
(xxii) 232 and 236, respectively;
(xxiii) 242 and 246, respectively;
(xxiv) 252 and 256, respectively;
(xxv) 262 and 266, respectively;
(xxvi) 272 and 276, respectively;
(xxvii) 282 and 286, respectively;
(xxviii) 292 and 296, respectively;
(xxix) 302 and 306, respectively;
(xxx) 312 and 316, respectively;
(xxxi) 322 and 326, respectively;
(xxxii) 332 and 336, respectively;
(xxxiii) 342 and 346, respectively, (xxxiv) 352 and 356, respectively;
(xxxv) 362 and 366, respectively;
(xxxvi) 372 and 376, respectively;
(xxxvii) 382 and 386, respectively;
(xxxviii) 392 and 396, respectively;
(xxxix) 402 and 406, respectively;
(xxxx) 412 and 416, respectively;
(xxxxi) 422 and 426, respectively; or (xxxxii) 432 and 436, respectively.
Embodiment 6.
The antibody or antigen-binding fragment of any one of Embodiments 1-5, which. (i) recognizes an epitope in a Domain A of SARS-CoV-2;
(ii) is capable of neutralizing a SARS CoV-2 infection; (iii) is capable of eliciting at least one immune effector function against SARS CoV-2; (iv) is capable of preventing shedding, from a cell infected with SARS CoV-2, of Si protein; or (v) any combination of (i)-(iv).
Embodiment 7. The antibody or antigen-binding fragment of any one of Embodiments 1-6, which is a IgG, IgA, IgM, IgE, or IgD isotype.
Embodiment 8. The antibody or antigen-binding fragment of any one of Embodiments 1-7, which is an IgG isotype selected from IgGl, IgG2, IgG3, and IgG4.
Embodiment 9. The antibody or antigen-binding fragment of any one of Embodiments 1-8, which is human, humanized, or chimeric.
Embodiment 10. The antibody or antigen-binding fragment of any one of Embodiments 1-9, wherein the antibody, or the antigen-binding fragment, comprises a human antibody, a monoclonal antibody, a purified antibody, a single chain antibody, a Fab, a Fab', a F(ab')2, a Fv, a scFv, or a scFab.
Embodiment 11. The antibody or antigen-binding fragment of Embodiment

10, wherein the scFv comprises more than one VH domain and more than one VL
domain.
Embodiment 12. The antibody or antigen-binding fragment of any one of Embodiments 1-11, wherein the antibody or antigen-binding fragment is a multi-specific antibody or antigen binding fragment.
Embodiment 13. The antibody or antigen-binding fragment of Embodiment 12, wherein the antibody or antigen binding fragment is a bi specific antibody or antigen-binding fragment.
Embodiment 14. The antibody or antigen-binding fragment of Embodiment 12 or 13, comprising:
(i) a first VH and a first VL; and (ii) a second VH and a second VL, wherein the first VH and the second VH are different and each independently comprise an amino acid sequence having at least 85% identity to the amino acid sequence set forth in any one of SEQ ID NOs 52, 22, 32, 42, 62, 72, 82, 92, 102, 112, 122, 132, 142, 152, 162, 172, 182 192, 202, 212, 222, 232, 242, 252, 262, 272, 282, 292, 302, 312, 322, 332, 342, 352, 362, 372, 382, 392, 402, 412, 422, and 432, wherein the first VL and the second VL are different and each independently comprise an amino acid sequence having at least 85% identity to the amino acid sequence set forth in any one of SEQ ID NOs.: 56, 26, 36, 46, 66, 76, 86, 96, 106, 116, 126, 136, 146, 156, 166, 176, 186, 196, 206, 216, 226, 236, 246, 256, 266, 276, 286, 296, 306, 316, 326, 336, 346, 356, 366, 376, 386, 396, 406, 416, 426, and 436, and wherein the first VH and the first VL together form a first antigen-binding site, and wherein the second VH and the second VL together form a second antigen-binding site.
Embodiment 15. The antibody or antigen-binding fragment of any one of Embodiments 1-14, wherein the antibody or antigen-binding fragment further comprises a Fc polypeptide or a fragment thereof Embodiment 16. The antibody or antigen-binding fragment of Embodiment 15, wherein the Fc polypeptide or fragment thereof comprises:
(i) a mutation that enhances binding to a FcRn as compared to a reference Fc polypeptide that does not comprise the mutation; and/or (ii) a mutation that enhances binding to a FcyR as compared to a reference Fe polypeptide that does not comprise the mutation.
Embodiment 17. The antibody or antigen-binding fragment of Embodiment 16, wherein the mutation that enhances binding to a FcRn comprises: M428L;
N434S;
N434H; N434A; N434S; M252Y; S254T; T256E; T250Q; P2571; Q311 I; D376V;
T307A; or E380A; or any combination thereof.
Embodiment 18. The antibody or antigen-binding fragment of Embodiment 16 or 17, wherein the mutation that enhances binding to FcRn comprises:
(i) M428L/N434S;
(ii) M252Y/S254T/T256E;
(iii) T250Q/M428L;
(iv) P257I/Q3111;
(v) P2571/N434H;
(vi) D376V/N434H;
(vii) T307A/E380A/N434A; or (viii) any combination of (i)-(vii).

Embodiment 19. The antibody or antigen-binding fragment of any one of Embodiments 16-18, wherein the mutation that enhances binding to FcRn comprises M428L/N434S.
Embodiment 20. The antibody or antigen-binding fragment of any one of Embodiments 16-19, wherein the mutation that enhances binding to a FcyR
comprises S239D; 1332E; A330L; G236A; or any combination thereof Embodiment 21. The antibody or antigen-binding fragment of any one of Embodiments 16-20, wherein the mutation that enhances binding to a FcyR
comprises:
(i) S239D/I332E;
(ii) S239D/A330L/1332E;
(iii) G236A/S239D/I332E; or (iv) G236A/A330L/1332E.
Embodiment 22. The antibody or antigen-binding fragment of any one of Embodiments 16-21, wherein the Fc polypeptide comprises a L234A mutation and a L235A mutation.
Embodiment 23. The antibody or antigen-binding fragment of any one of Embodiments 1-22, which comprises a mutation that alters glycosylation, wherein the mutation that alters glycosylation comprises N297A, N297Q, or N297G, and/or which is aglycosylated and/or afucosylated.
Embodiment 24. An isolated polynucleotide encoding the antibody or antigen-binding fragment of any one of Embodiments 1-23, or encoding a VH, a heavy chain, a VL, and/or a light chain of the antibody or the antigen-binding fragment.
Embodiment 25. The polynucleotide of Embodiment 24, wherein the polynucleotide comprises deoxyribonucleic acid (DNA) or ribonucleic acid (RNA), wherein the RNA optionally comprises messenger RNA (mRNA).
Embodiment 26. The polynucleotide of Embodiment 24 or 25, which is codon-optimized for expression in a host cell.
Embodiment 27 The polynucleotide of any one of Embodiments 24-26, comprising a polynucleotide having at least 50% identity to the polynucleotide sequence according to any one or more of SEQ ID NOs.: 60, 61, 30, 31, 40, 41, 50, 51, 70, 71, 80, 81, 90, 91, 100, 101, 110, 111, 120, 121, 130, 131, 140, 141, 150, 151, 160, 161, 170, 171, 180, 181, 190, 191, 200, 201, 210, 211, 220, 221, 230, 231, 240, 241, 250, 251, 260, 261, 270, 271, 280, 281, 290, 291, 300, 301, 310, 311, 320, 321, 330, 331, 340, 341, 350, 351, 360, 361, 370, 371, 380, 381, 390, 391, 400, 401, 410, 411, 420, 421, 430, 431, 440, and 441, or any combination thereof.
Embodiment 28. A recombinant vector comprising the polynucleotide of any one of Embodiments 24-27.
Embodiment 29. A host cell comprising the polynucleotide of any one of Embodiments 24-27 and/or the vector of Embodiment 28, wherein the polynucleotide is heterologous to the host cell.
Embodiment 30. A human B cell comprising the polynucleotide of any one of Embodiments 24-28, wherein polynucleotide is heterologous to the human B
cell and/or wherein the human B cell is immortalized.
Embodiment 31. A composition or combination comprising:
(i) the antibody or antigen-binding fragment of any one of Embodiments 1-23;
(ii) the polynucleotide of any one of Embodiments 24-27;
(iii) the recombinant vector of Embodiment 28, (iv) the host cell of Embodiment 29; and/or (v) the human B cell of Embodiment 30, and an optional pharmaceutically acceptable excipient, carrier, or diluent.
Embodiment 32. The composition or combination of Embodiment 31, comprising two or more antibodies or antigen-binding fragments of any one of Embodiments 1-23, and/or comprising one or more antibody according to any one of Embodiments 1-23 and an antibody or antigen-binding fragment that binds to a SARS
CoV-2 surface glycoprotein RBD.
Embodiment 33. A composition comprising the polynucleotide of any one of Embodiments 24-27 encapsulated in a carrier molecule, wherein the carrier molecule optionally comprises a lipid, a lipid-derived delivery vehicle, such as a liposome, a solid lipid nanoparticle, an oily suspension, a submicron lipid emulsion, a lipid microbubble, an inverse lipid micelle, a cochlear liposome, a lipid microtubule, a lipid microcylinder, lipid nanoparticle (LNP), or a nanoscale platform.

Embodiment 34. A method of treating a SARS-CoV-2 infection in a subject, the method comprising administering to the subject an effective amount of (i) the antibody or antigen-binding fragment of any one of Embodiments 1-23;
(ii) the polynucleotide of any one of Embodiments 24-27;
(iii) the recombinant vector of Embodiment 28;
(iv) the host cell of Embodiment 29;
(v) the human B cell of Embodiment 30; and/or (vi) the composition or combination of any one of Embodiments 31-33.
Embodiment 35. The antibody or antigen-binding fragment of any one of Embodiments 1-23, the polynucleotide of any one of Embodiments 24-27, the recombinant vector of Embodiment 28, the host cell of Embodiment 29, the human B
cell of Embodiment 30, and/or the composition or combination of any one of Embodiments 31-33 for use in a method of treating a SARS-CoV-2 infection in a subject.
Embodiment 36. The antibody or antigen-binding fragment of any one of Embodiments 1-23, the polynucleotide of any one of Embodiments 24-27, the recombinant vector of Embodiment 28, the host cell of Embodiment 29, the human B
cell of Embodiment 30, and/or the composition or combination of any one of Embodiments 31-33 for use in the preparation of a medicament for the treatment of a SARS-CoV-2 infection in a subject.
Embodiment 37. A method for in vitro or ex vivo diagnosis of a SARS-CoV-2 infection, the method comprising:
(i) contacting a sample from a subject with an antibody or antigen-binding fragment of any one of Embodiments 1-23; and (ii) detecting a complex comprising an antigen and the antibody, or comprising an antigen and the antigen binding fragment.
Embodiment 38 The method of Embodiment 37, wherein the sample comprises blood isolated from the subject.

Embodiment 39. An antibody, or an antigen-binding fragment thereof, that competes for binding to a SARS-CoV-2 surface glycoprotein with the antibody or antigen-binding fragment of any one of Embodiments 1-23.
Embodiment 40. A method of preventing or treating or neutralizing a coronavirus infection in a subject, the method comprising administering to a subject an effective amount of (i) an antibody or antigen-binding fragment of any one of Embodiments 1-23 or 39 and (ii) an antibody or antigen-binding fragment that is capable of specifically binding to a SARS CoV-2 S protein RBD.
Embodiment 41. A method of detecting a SARS-CoV-2 protein or polypeptide in a sample, comprising contacting the sample with the antibody or antigen-binding fragment of any one of Embodiments 1-23 or 39 and detecting binding of the antibody or antigen-binding fragment to the SARS-CoV-2 protein or polypeptide Embodiment 42. The method of Embodiment 41, wherein detecting binding of the antibody or antigen-binding fragment to the SARS-CoV-2 protein or polypeptide comprises immunohistochemistry, ELISA, agglutination, immuno-dot, immuno-chromatography, and/or immuno-filtration Embodiment 43. The antibody or antigen-binding fragment thereof of any one of Embodiments 1-23 for use in a method of detecting a SARS-CoV-2 protein or polypeptide in a sample, the method comprising contacting the sample with the antibody or antigen-binding fragment and detecting binding of the antibody or antigen-binding fragment to the SARS-CoV-2 protein or polypeptide, wherein, optionally, detecting binding of the antibody or antigen-binding fragment to the SARS-CoV-protein or polypepti de comprises immunohistochemistry, ELISA, agglutination, immuno-dot, immuno-chromatography, and/or immuno-filtration.
Embodiment 44. A method of diagnosing a SARS-CoV-2 infection in a subject, comprising testing a biological sample from the subject for the presence of a SARS-CoV-2 protein or polypeptide, wherein the testing comprises contacting the sample with the antibody or antigen-binding fragment of any one of Embodiments and detecting binding of the antibody or antigen-binding fragment to the SARS-CoV-2 protein or polypeptide, wherein, optionally, detecting binding of the antibody or antigen-binding fragment to the SARS-CoV-2 protein or polypeptide comprises immunohistochemistry, ELISA, agglutination, immuno-dot, immuno-chromatography, and/or immuno-filtration.
Embodiment 45. The method of Embodiment 44, wherein the SARS-CoV-2 protein or polypeptide is detected by immunohistochemistry.
Embodiment 46. The method of any one of Embodiments 41-45, wherein the sample comprises a nasal secretion, sputum, a bronchial lavage, urine, stool, saliva, sweat, or any combination thereof.
Embodiment 47. An antibody or antigen-binding fragment thereof for use in a method of diagnosing a SARS-CoV-2 infection in a subject, the method comprising testing a biological sample from the subject for the presence of a SARS-CoV-2 protein or polypeptide, wherein the testing comprises contacting the sample with the antibody or antigen-binding fragment and detecting binding of the antibody or antigen-binding fragment to the SARS-CoV-2 protein or polypeptide, wherein, optionally, detecting binding of the antibody or antigen-binding fragment to the SARS-CoV-2 protein or polypeptide comprises immunohistochemistry, ELISA, agglutination, immuno-dot, immuno-chromatography, and/or immuno-filtration, wherein, optionally, the antibody or antigen-binding fragment is the antibody or antigen-binding fragment thereof of any one of Embodiments 1-23.
Embodiment 48. The antibody or antigen-binding fragment of any one of Embodiments 1-23 or the antibody or antigen-binding fragment for use of Embodiment 43 or 47, or the method of any one of Embodiments 41, 42, or 44-46, wherein the antibody or antigen-binding fragment comprises a detectable agent.
Embodiment 49. A kit comprising the antibody or antigen-binding fragment thereof of any one of Embodiments 1-23, and optional instructions for using the antibody or antigen-binding fragment to detect the presence of a SARS-CoV-protein or polypeptide in a biological sample.
Embodiment 50. The kit according to Embodiment 49 for use in a method of detecting the presence of a SARS-CoV-2 protein or polypeptide in a biological sample.
Embodiment 5L The kit of for use of Embodiment 50, wherein the method comprises detecting the presence of a SARS-CoV-2 protein or polypeptide by immunohistochemistry, ELISA, agglutination, immuno-dot, immuno-chromatography, and/or immuno-filtration.
Embodiment 52. The kit of Embodiment 49 or the kit for use of any one of Embodiments 50 or 51, further comprising a detectably labeled secondary antibody.
Embodiment 53. The kit of Embodiment 49 or the kit for use of any one of Embodiments 50-52, further comprising one or more of a sample buffer, a wash buffer, an immunodetection buffer, a substrate, detection means, a control sample, a reference sample, and instructions for use.
Embodiment 54. The kit of Embodiment 49 or the kit for use of any one of Embodiments 50-53, wherein the sample comprises a nasal secretion, sputum, bronchial lavage, urine, stool, saliva, and/or sweat.
Embodiment 55. The composition or combination of Embodiment 32, comprising (a) antibody S2X333 (or an antigen-binding fragment thereof) or an antibody or antigen-binding fragment thereof that competes with antibody S2X333 for SARS-CoV-2 S protein binding and (b) antibody S309 (or an antigen-binding fragment thereof) or an antibody or antigen-binding fragment thereof that competes with antibody S309 for SARS-CoV-2 S protein binding.
Embodiment 56. The composition of Embodiment 32, comprising a) antibody S2X333 (or an antigen-binding fragment thereof) or an antibody or an antigen-binding fragment thereof that competes with antibody S2X333 for SARS-CoV-2 S
protein binding and b) antibody S2E12 (or an antigen-binding fragment thereof) or an antibody or an antigen-binding fragment thereof that competes with antibody S2E12 for SARS-CoV-2 S protein binding.
Embodiment 57. The composition of Embodiment 32, comprising (a) antibody S2X333 (or an antigen-binding fragment thereof) or an antibody or an antigen-binding fragment thereof that competes with antibody S2X333 for SARS-CoV-2 S
protein binding and (b) antibody S2M11 (or an antigen-binding fragment thereof) or an antibody or an antigen-binding fragment thereof that competes with antibody for SARS-CoV-2 S protein binding.
Embodiment 58. The antibody or antigen-binding fragment of Embodiment 12 or 13, comprising (i) a first VH and a first VL; and (ii) a second VH and a second VL, wherein the first VH comprises an amino acid sequence having at least 85%
(i.e., 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) identity to the amino acid sequence set forth in SEQ ID NO: 52 and the first VL comprises an amino acid sequence having at least 85% (i.e., 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) identity to the amino acid sequence set forth in SEQ ID NO: 56; and a) the second VH comprises an amino acid sequence having at least 85% (i.e., 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) identity to the amino acid sequence set forth in SEQ ID NO: 442 and the second VL comprises an amino acid sequence having at least 85% (i.e., 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) identity to the amino acid sequence set forth in SEQ ID NO: 446;
b) the second VH comprises an amino acid sequence having at least 85% (i.e., 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) identity to the amino acid sequence set forth in SEQ ID NO: 450 and the second VL comprises an amino acid sequence having at least 85% (i.e., 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) identity to the amino acid sequence set forth in SEQ ID NO: 454; or c) the second VH comprises an amino acid sequence having at least 85% (i.e., 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) identity to the amino acid sequence set forth in SEQ ID NO: 458 and the second VL comprises an amino acid sequence having at least 85% (i.e., 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) identity to the amino acid sequence set forth in SEQ ID NO: 462; and wherein the first VH and the first VL together form a first antigen-binding site, and wherein the second VH and the second VL together form a second antigen-binding site.
Embodiment 59. A method of treating or preventing SARS-CoV-2 infection comprising administering a composition or combination of any one of Embodiments 55-57 or the antibody or antigen-binding fragment of Embodiment 58.

Embodiment 60. The composition or combination of any one of Embodiments 55-57, wherein, optionally the antibody or antigen-binding fragment of a) and/or b) comprises (i) a Fc polypeptide comprising a mutation that enhances binding to a FcRn as compared to a reference Fc polypeptide that does not comprise the mutation;
and/or (ii) a Fc polypeptide comprising a mutation that enhances binding to a FcyR as compared to a reference Fc polypeptide that does not comprise the mutation.
Embodiment 61. The antibody or antigen-binding fragment of Embodiment 58, or the method of Embodiment 59, wherein, optionally, the antibody or antigen-binding fragment comprises (i) a Fc polypeptide comprising a mutation that enhances binding to a FcRn as compared to a reference Fc polypeptide that does not comprise the mutation; and/or (ii) a Fc polypeptide comprising a mutation that enhances binding to a FcyR as compared to a reference Fc polypeptide that does not comprise the mutation.
Table 1. Sequences SEQ
Sequence ID Sequence Description NO.
1 attaaaggtt tataccttcc caggtaacaa accaaccaac tttcgatctc ttgtagatct 61 gttctctaaa cgaactttaa aatctgtgtg gctgtcactc ggctgcatgc ttagtgcact 121 cacgcagtat aattaataac taattactgt cgttgacagg acacgagtaa ctcgtctatc 181 ttctgcaggc tgcttacggt ttcgtccgtg ttgcagccga tcatcagcac atctaggttt 241 cgtccgggtg tgaccgaaag gtaagatgga SARS-CoV-2 gagccttgtc cctgghtca acgagaaaac 301 acacgtccaa Wuhan seafood ctcagthgc ctghttaca ggttcgcgac gtgctcgtac gtggctttgg market pneumonia 361 agactccgtg gaggaggtct tatcagaggc cgtcaacat virus isolate cttaaagatg gcacttgtgg 421 cttagtagaa gttgaaaaag Wuhan-Hu-1 1 gcgttttgcc tcaacttgaa cagccctatg tgttcatcaa 481 genomic sequence acgttcggat gctcgaactg cacctcatgg tcatgttatg gttgagctgg (GenBank: tagcagaact 541 cgaaggcatt cagtacggtc gtagtggtga MN908947.3;Janu gacacttggt gtccttgtcc ctcatgtggg 601 cgaaatacca ary 23, 2020) gtggcttacc gcaaggttct tcttcgtaag aacggtaata aaggagctgg 661 tggccatagt tacggcgccg atctaaagtc atttgactta ggcgacgagc ttggcactga 721 tccttatgaa gattlicaag aaaactggaa cactaaacat agcagtggtg ttacccgtga 781 actcatgcgt gag cttaacg gaggggcata cactcgctat gtcgataaca acttctgtgg 841 ccctgatggc taccctcttg agtgcattaa agaccttcta gcacgtgctg gtaaagcttc 901 atgcactttg SEQ
Sequence ID Sequence Description NO.
tccgaacaac tggactttat tgacactaag aggggtgtat actgctgccg 961 tgaacatgag catgqnattg cttggtacac ggaacgttct gaaaagagct atgaattgca 1021 gacacc Lilt gaaattaaat tggcaaagaa atttgacacc ttcaatgggg aatgtccaaa 1081 ttttgtattt cccttaaatt ccataatcaa gactattcaa ccaagggttg aaaagaaaaa 1141 gcttgatggc tttatgggta gaattcgatc tgtctatcca gttgcgtcac caaatgaatg 1201 caaccaaatg tgcctttcaa ctctcatgaa gtgtgatcat tgtggtgaaa cttcatggca 1261 gacgggcgat tttgttaaag ccacttgcga attilgtggc actgagaatt tgactaaaga 1321 aggtgccact acttgtggtt acttacccca aaatgctgtt gttaaaattt attgtccagc 1381 atgtcacaat tcagaagtag gacctgagca tagtcttgcc gaataccata atgaatctgg 1441 cttgaaaacc attcttcgta agggtggtcg cactattgcc tttggaggct gtgtgttctc 1501 ttatgttggt tgccataaca agtgtgccta ttgggttcca cgtgctagcg ctaacatagg 1561 ttgtaaccat acaggtgttg ttggagaagg ttccgaaggt cttaatgaca accttcttga 1621 aatactccaa aaagagaaag tcaacatcaa tattgttggt gactttaaac ttaatgaaga 1681 gatcgccatt attliggcat clittictgc ttccacaagt gcttttgtgg aaactgtgaa 1741 aggtttggat tataaagcat tcaaacaaat tgttgaatcc tgtggtaatt ttaaagttac 1801 aaaaggaaaa gctaannaag gtgcctggaa tattggtgaa cagaaatcaa tactgagtcc 1861 tctttatgca tttgcatcag aggctgctcg tgttgtacga tcaattttct cccgcactct 1921 tgaaactgct caaaattctg tgcgtgtttt acagaaggcc gctataacaa tactagatgg 1981 aatttcacag tattcactga gactcattga tgctatgatg ttcacatctg atttggctac 2041 taacaatcta gttgtaatgg cctacattac aggtggtgtt gttcagttga cttcgcagtg 2101 gctaactaac atctttggca ctgtttatga annactcaaa cccgtccttg attggcttga 2161 agagaagttt aaggaaggtg tagagtact tagagacggt tgggaaattg ttaaatttat 2221 ctcaacctgt gcttgtgaaa ttgtcggtgg acaaattgtc acctgtgcaa aggaaattaa 2281 ggagagtgtt cagacattct ttaagcttgt aaataaattt ttggctttgt gtgctgactc 2341 tatcattatt ggtggagcta aacttaaagc cttgaattta ggtgaaacat ttgtcacgca 2401 ctcaaaggga ttgtacagaa agtgtgttaa atccagagaa gaaactggcc tactcatgcc 2461 tctaaaagcc ccaaaagaaa ttatcttctt agagggagaa acacttccca cagaagtgtt 2521 aacagaggaa gttgtcttga aaactggtga tttacaacca ttagaacaac ctactagtga 2581 agctgttgaa gctccattgg ttggtacacc agtttgtatt aacgggctta tgttgctcga 2641 aatcaaagac acagaaaagt actgtgccct tgcacctaat atgatggtaa caaacaatac 2701 cttcacactc aaaggcggtg caccaacaaa ggttactitt ggtgatgaca ctgtgataga 2761 agtgcaaggt tacaagagtg SEQ
Sequence ID Sequence Description NO.
tgaatatcac ttttgaactt gatgaaagga ttgataaagt 2821 acttaatgag aagtgctctg cctatacagt tgaactcggt acagaagtaa atgagttcgc 2881 ctgtgttgtg gcagatgctg tcataaaaac tttgcaacca gtatctgaat tacttacacc 2941 actgggcatt gatttagatg agtggagtat ggctacatac tacttatttg atgagtctgg 3001 tgagtttaaa ttggcttcac atatgtattg ttctttctac cctccagatg aggatgaaga 3061 agaaggtgat tgtgaagaag aagagifiga gccatcaact caatatgagt atggtactga 3121 agatgattac caaggtaaac ctttggaatt tggtgccact tctgctgctc ttcaacctga 3181 agaagagcaa gaagaagatt ggttagatga tgatagtcaa caaactgttg gtcaacaaga 3241 cggcagtgag gacaatcaga caactactat tcaaacaatt gttgaggttc aacctcaatt 3301 agagatggaa cttacaccag ttgttcagac tattgaagtg aatagtitta gtggttatrt 3361 aaaacttact gacaatgtat acattaaaaa tgcagacatt gtggaagaag ctaaaaaggt 3421 aaaaccaaca gtggttgtta atgcagccaa tgtttacctt aaacatggag gaggtgttgc 3481 aggagcctta aataaggcta ctaacaatgc catgcaagtt gaatctgatg attacatagc 3541 tactaatgga ccacttaaag tgggtggtag ttgtgtttta agcggacaca atcttgctaa 3601 acactgtctt catgttgtcg gcccaaatgt taacaaaggt gaagacattc aacttcttaa 3661 gagtgcttat ganaatttta atcagcacga agttctactt gcaccattat tatcagctgg 3721 tatilliggt gctgacccta tacattcttt aagagtttgt gtagatactg ttcgcacaaa 3781 tgtctactta gctgtattg ataaaaatct ctatgacaaa cttgtttcaa gcitiligga 3841 aatgaagagt gaaaagcaag ttgaacaaaa gatcgctgag attcctaaag aggaagttaa 3901 gccatttata actgaaagta aaccttcagt tgaacagaga aaacaagatg ataagaaaat 3961 caaagcttgt gttgaagaag ttacaacaac tctggaagaa actaagttcc tcacagaaaa 4021 cttgttactt tatattgaca ttaatggcaa tcttcatcca gattctgcca ctcttgttag 4081 tgacattgac atcactttct taaagaaaga tgctccatat atagtgggtg atgttgttca 4141 agagggtgtt ttaactgctg tggttatacc tactaaaaag gctggtggca ctactgaaat 4201 gctagcgaaa gctttgagaa aagtgccaac agacaattat ataaccactt acccgggtca 4261 gggtttaaat ggttacactg tagaggaggc aaagacagtg cttaaaaagt gtaaaagtgc 4321 caltacatt ctaccatcta ttatctctaa tgagaagcaa gaaattcttg gaactgtttc 4381 ttggaatttg cgagaaatgc ttgcacatgc agaagaaaca cgcaaattaa tgcctgtctg 4441 tgtggaaact aaagccatag tttcaactat acagcgtaaa tataagggta ttaaaataca 4501 agagggtgtg gttgattatg gtgetagatt ttaattlac accagtaaaa caactgtagc 4561 gtcacttatc aacacactta acgatctaaa tgaaactctt gttacaatgc SEQ
Sequence ID Sequence Description NO.
cacttggcta 4621 tgtaacacat ggcttaaatt tggaagaagc tgctcggtat atgagatctc tcaaagtgcc 4681 agctacagtt tctgtttctt cacctgatgc tgttacagcg tataatggtt atcttacttc 4741 acttctaaa acacctgaag aacattttat tgaaaccatc tcacttgctg gttcctataa 4801 agattggtcc tattctggac aatctacaca actaggtata gaatttctta agagaggtga 4861 taanagtgta tattacacta gtaatcctac cacattccac ctagatggtg aagttatcac 4921 ctttgacaat cttaagacac ttctttcttt gagagaagtg aggactatta aggtgtttac 4981 aacagtagac aacattaacc tccacacgca agttgtggac atgtcaatga catatggaca 5041 acagt-ttggt ccaacttatt tggatggagc tgatgttact aaaataaaac ctcataattc 5101 acatgaaggt aaaacatill atglittacc taatgatgac actctacgtg ttgaggcttt 5161 tgagtactac cacacaactg atcctagttt tctgggtagg tacatgtcag cattaaatca 5221 cactaaaaag tggaaatacc cacaagt-taa tggtttaact tctattinnt gggcagataa 5281 caactgttat cttgccactg cattgttaac actccaacaa atagagttga agtttaatcc 5341 acctgctcta caagatgctt attacagagc aagggctggt gaagctgcta actIllgtgc 5401 acttatctta gcctactgta ataagacagt aggtgagtta ggtgatgtta gaganacaat 5461 gagttacttg tttcaacatg ccaatttaga ttcttgcaaa agagtcttga acg-tggtgtg 5521 taaaacttgt ggacaacagc agacaaccct taagggtgta gaagctgtta tgtacatggg 5581 cacacifict tatgaacaat ttaagaaagg tgttcagata ccttgtacgt gtggtaaaca 5641 agctacaaaa tatctagtac aacaggagtc accttttgtt atgatgtcag caccacctgc 5701 tcagtatgaa cttaagcatg gtacatttac ttgtgctagt gagtacactg gtaattacca 5761 gtgtggtcac tataaacata taacttctaa agaaactttg tattgcatag acggtgcttt 5821 acttacaaag tcctcagaat acaaaggtcc tattacggat g ttlictaca aagaaaacag 5881 ttacacaaca accataaaac cagttactta taaattggat ggtgttgttt gtacagaaat 5941 tgaccctaag ttggacaatt attataagaa agacaattct tatttcacag agcaaccaat 6001 tgatcttgta ccaaaccaac catatccaaa cgcaagcttc gataatitta agtttgtatg 6061 tgataatatc aaatttgctg atgatttaaa ccagttaact ggttataaga aacctgcttc 6121 aagagagctt aaagttacat ttttccctga cttaaatggt gatgtggtgg ctattgatta 6181 taaacactac acaccctctt ttaagaaagg agctaaattg ttacataaac ctattgtttg 6241 gcatgttaac aatgcaacta ataaagccac gtataaacca aatacctggt gtatacgttg 6301 tctttggagc acaaaaccag ttganacatc aaattcgttt gatgtactga agtcagagga 6361 cgcgcaggga atggataatc ttgcctgcga agatctaaaa ccagtctctg aagaagtagt 6421 gganaatcct accatacaga aagacglict tgagtgtaat SEQ
Sequence ID Sequence Description NO.
gtgaaaacta ccgaagttgt 6481 aggagacatt atacttaaac cagcaaataa tagtttaaaa attacagaag aggttggcca 6541 cacagatcta atggctgctt atgtagacaa ttctagtctt actattaaga aacctaatga 6601 attatctaga gtattaggtt tgaaaaccct tgctactcat ggtttagctg ctgttaatag 6661 tgtcccttgg gatactatag ctaattatgc taagcctttt cttaacaaag ttgttagtac 6721 aactactaac atagttacac ggtgtttaaa ccgtgtttgt actaattata tgccttattt 6781 ctttacttta ttgctacaat tgtgtacttt tactagaagt acaaattcta gaattaaagc 6841 atctatgccg actactatag caaagaatac tgttaagagt gtcggtaaat ttigtctaga 6901 ggcttcattt aattatttga agtcacctaa lltttctaaa ctgataaata ttataatttg 6961 gttlttacta ttaagtgttt gcctaggttc tttaatctac tcaaccgctg ctttaggtgt 7021 tttaatgtct aatttaggca tgccttctta ctgtactggt tacagagaag gctatttgaa 7081 ctctactaat gtcactattg caacctactg tactggttct ataccttgta gtgtttgtct 7141 tagtggttta gattctttag acacctatcc ttctttagaa actatacaaa ttaccatttc 7201 atcttttaaa tgggatttaa ctgcttttgg cttagttgca gagtggtttt tggcatatat 7261 tcttttcact aggittlict atgtacttgg attggctgca atcatgcaat tglitticag 7321 ctattligca gtacatttta ttagtaattc ttggcttatg tggttaataa ttaatcttgt 7381 acaaatggcc ccgatttcag ctatggttag aatgtacatc ttctttg cat cattttatta 7441 tgtatggaaa agttatgtgc atgttgtaga cggttgtaat tcatcaactt gtatgatgtg 7501 ttacaaacgt aatagagcaa caagagtcga atgtacaact attgttaatg gtgttagaag 7561 gtccttttat gtctatgcta atggaggtaa aggcttttgc aaactacaca attggaattg 7621 tgttaattgt gatacattct gtgctggtag tacatttatt agtgatgaag ttgcgagaga 7681 cttgtcacta cagtttaaaa gaccaataaa tcctactgac cagtcttctt acatcgttga 7741 tagtgttaca gtgaagaatg gttccatcca tctttacttt gataaagctg gtcaaaagac 7801 ttatgaaaga cattctctct ctcattttgt taacttagac aacctgagag ctaataacac 7861 taaagglica ttgcctatta atgttatagt ttitgatggt aaatcaaaat gtgaagaatc 7921 atctgcaaaa tcagcgtctg tttactacag tcagcttatg tgtcaaccta tactgttact 7981 agatcaggca ttagtgtctg atgttggtga tagtgcggaa gttgcagtta aaatgtttga 8041 tgcttacgtt aatacg LIII catcaacttt taacgtacca atggaa,aaac tcaaaacact 8101 agttgcaact gcagaagctg aacttgcaaa gaatgtgtcc ttagacaatg tcttatctac 8161 ttttatttca gcagctcggc aagggtttgt tgattcagat gtagaaacta aagatgttgt 8221 tgaatgtctt aaattgtcac atcaatctga catagaagtt actggcgata gttgtaataa 8281 ctatatgctc acctataaca aagttgaaaa catgacaccc cgtgaccttg SEQ
Sequence ID Sequence Description NO.
gtgcttgtat 8341 tgactgtagt gcgcgtcata ttaatgcgca ggtagcqnaa agtcacaaca ttgctttgat 8401 atggaacgtt aaagatttca tgtcattgtc tgaacaacta cgaaaacaaa tacgtagtgc 8461 tgctaaaaag aataacttac cttttaagtt gacatgtgca actactagac aagttgttaa 8521 tgttgtaaca acaaagatag cacttaaggg tggtaaaatt gttaataatt ggttgaagca 8581 gttaattaaa gttacacttg tgttcctttt tgttgctgct attttctatt taataacacc 8641 tgttcatgtc atgtctaaac atactgactt ttcaagtgaa atcataggat acaaggctat 8701 tgatggtggt gtcactcgtg acatagcatc tacagatact tgttttgcta acaaacatgc 8761 tgaLLILgac acatggttta gccagcgtgg tggtagttat actaatgaca aagcttgccc 8821 attgattgct gcagtcataa caagagaagt gggittlgtc gtgcctggtt tgcctggcac 8881 gatattacgc acaactaatg gtgacttlit gcatttctta cctagagttt ttagtgcagt 8941 tggtaacatc tgttacacac catcaaaact tatagagtac actgactttg caacatcagc 9001 ttgtgttttg gctgctgaat gtacaatill taaagatgct tctggtaagc cagtaccata 9061 ttgttatgat accaatgtac tagaaggttc tgttgcttat gaaagtttac gccctgacac 9121 acgttatgtg ctcatggatg gctctattat tcaatttcct aacacctacc ttgaaggttc 9181 tgttagagtg gtaacaactt ttgattctga gtactgtagg cacggcactt gtgaaagatc 9241 agaagctggt gtttgtgtat ctactagtgg tag atgggta cttaacaatg attattacag 9301 atctttacca ggaglitict gtggtgtaga tgctgtaaat ttacttacta atatgtttac 9361 accactaatt caacctattg gtgclligga catatcagca tctatagtag ctggtggtat 9421 tgtagctatc gtagtaacat gccttgccta ctattttatg aggtttagaa gagclitAgg 9481 tgaatacagt catgtagttg cctttaatac tttactattc cttatgtcat tcactgtact 9541 ctgtttaaca ccagtttact cattcttacc tggtgtttat tctgttattt acttgtactt 9601 gacattttat cttactaatg atgtttcttt tttagcacat attcagtgga tggttatgtt 9661 cacaccttta gtacctttct ggataacaat tgcttatatc atttgtattt ccacaaagca 9721 tttctattgg ttctttagta attacctaaa gagacgtgta gtctttaatg gtgtttcctt 9781 tagtactill gaagaagctg cgctgtgcac cittligtta aataaagaaa tgtatctaaa 9841 gttgcgtagt gatgtgctat tacctcttac gcaatataat agatacttag ctctttataa 9901 taagtacaag tattttagtg gagcaatgga tacaactagc tacagagaag ctgcttgttg 9961 tcatctcgca aaggctctca atgacttcag taactcaggt tctgatgttc tttaccaacc 10021 accacaaacc tctatcacct cagctgtttt gcagagtggt tttagaaaaa tggcattccc 10081 atctggtana gttgagggtt gtatggtaca agtaacttgt ggtacaacta cacttaacgg 10141 tctttggctt gatgacgtag tttactgtcc aagacatgtg atctgcacct ctgaagacat 10201 gcttaaccct aattatgaag SEQ
Sequence ID Sequence Description NO.
atttactcat tcgtaagtct aatcataatt tcttggtaca 10261 ggctggtaat gttcaactca gggttattgg acattctatg caaaattgtg tacttaagct 10321 taaggttgat acagccaatc ctaagacacc taagtataag tttgttcgca ttcaaccagg 10381 acagactttt tcagtgttag cttgttacaa tggttcacca tctggtgttt accaatgtgc 10441 tatgaggccc aatttcacta ttaagggttc attccttaat ggttcatgtg gtagtgttgg 10501 ttttaacata gattatgact gtgtctcttt ttgttacatg caccatatgg aattaccaac 10561 tggagttcat gctggcacag acttagaagg taacttttat ggaccillig ttgacaggca 10621 aacagcacaa gcagctggta cggacacaac tattacagtt aatglittag cttggttgta 10681 cgctgctgtt ataaatggag acaggtggtt tctcaatcga tttaccacaa ctcttaatga 10741 ctttaacctt gtggctatga agtacaatta tgaacctcta acacaagacc atgttgacat 10801 actaggacct ctttctgctc aaactggaat tgccglltla gatatgtgtg cttcattaaa 10861 agaattactg caaaatggta tgaatggacg taccatattg ggtagtgctt tattagaaga 10921 tgaatttaca ccttttgatg ttgttagaca atgctcaggt gttactttcc aaagtgcagt 10981 gagaagaaca atcaagggta cacaccactg gttgttactc acaalittga cttcactitt 11041 aglillagtc cagagtactc aatggtcttt gttc111111 ttgtatgaaa atgccILLIL 11101 acctillgct atgggtatta ttgctatgtc tgcttttgca atgatgtttg tcaaacataa 11161 gcatgcattt ctctgtttgt ttttgttacc ttctcttgcc actgtagctt attttaatat 11221 ggtctatatg cctgctagtt gggtgatgcg tattatgaca tggttggata tggttgatac 11281 tagtttgtct ggttttaagc taaaagactg tgttatgtat gcatcagctg tagtgttact 11341 aatccttatg acagcaagaa ctgtgtatga tgatggtgct aggagagtgt ggacacttat 11401 gaatgtcttg acactcgttt ataaagttta ttatggtaat gctttagatc aagccatttc 11461 catgtgggct cttataatct ctgttacttc taactactca ggtgtagtta caactgtcat 11521 gtttliggcc agaggtattg ittilatgtg tgttgagtat tgccctattt tcttcataac 11581 tggtaataca cttcagtgta taatgctagt ttattgtttc ttaggctatt tttgtacttg 11641 ttactttggc ctcttttgtt tactcaaccg ctactttaga ctgactcttg gtgtttatga 11701 ttacttagtt tctacacagg agtttagata tatgaattca cagggactac tcccacccaa 11761 gaatagcata gatgccttca aactcaacat taaattgttg ggtgttggtg gcanaccttg 11821 tatcaaagta gccactgtac agtctaaaat gtcagatgta aagtgcacat cagtagtctt 11881 actctcagtt ttgcaacaac tcagagtaga atcatcatct aaattgtggg ctcaatgtgt 11941 ccagttacac aatgacattc tcttagctaa agatactact gaagcctttg aaaaaatggt 12001 ttcactactt tctgtLLLgc tttccatgca gggtgctgta gacataaaca agctttgtga 12061 agaaatgctg gacaacaggg caaccttaca agctatagcc tcagagttta SEQ
Sequence ID Sequence Description NO.
gttcccttcc 12121 atcatatgca gcttttgcta ctgctcaaga agcttatgag caggctgttg ctaatggtga 12181 ttctgaagtt gttcttaaaa agttgaagaa gtctttgaat gtggctaaat ctgaatttga 12241 ccgtgatgca gccatgcaac gtaagttgga aaagatggct gatcaagcta tgacccaaat 12301 gtataaacag gctagatctg aggacaagag ggcaaaagtt actagtgcta tgcagacaat 12361 gcttttcact atgcttagaa agttggataa tgatgcactc aacaacatta tcaacaatgc 12421 aagagatggt tgtgttccct tgaacataat acctcttaca acagcagcca aactaatggt 12481 tgtcatacca gactataaca catataaaaa tacgtgtgat ggtacaacat ttacttatgc 12541 atcagcattg tgggaaatcc aacaggttgt agatgcagat agtaaaattg ttcaacttag 12601 tgaaattagt atggacaatt cacctaattt agcatggcct cttattgtaa cagctttaag 12661 ggccaattct gctgtcaaat tacagaataa tgagcttagt cctgttgcac tacgacagat 12721 gtcttgtgct gccggtacta cacaaactgc ttgcactgat gacaatgcgt tagcttacta 12781 caacacaaca aagggaggta ggtttgtact tgcactgtta tccgatttac aggatttgaa 12841 atgggctaga ttccctaaga gtgatggaac tggtactatc tatacagaac tggaaccacc 12901 ttgtaggttt gttacagaca cacctaaagg tcctaaagtg aagtatttat actttattaa 12961 aggattaaac aacctaaata gaggtatggt acttggtagt ttagctgcca cagtacgtct 13021 acaagctggt aatgcaacag aagtgcctgc caattcaact gtattatctt tctg(gcttt 13081 tgctgtagat gctgctaaag cttacaaaga ttatctagct agtgggggac naccaatcac 13141 taattgtgtt aagatgttgt gtacacacac tggtactggt caggcaataa cagttacacc 13201 ggaagccaat atggatcaag aatcctttgg tggtgcatcg tgttgtctgt actgccgttg 13261 ccacatagat catccaaatc ctaaaggatt ttgtgactta aaaggtaagt atgtacnnat 13321 acctacaact tgtgctaatg accctgtggg tillacactt aaaaacacag tctgtaccgt 13381 ctgcggtatg tggaaaggtt atggctgtag ttgtgatcaa ctccgcgaac ccatgcttca 13441 gtcagctgat gcacaatcgt ttttnnacgg gtttgcggtg taagtgcagc ccgtcttaca 13501 ccgtgcggca caggcactag tactgatgtc gtatacaggg cttttgacat ctacaatgat 13561 aaagtagctg glittgctaa attcctaaaa actaattgtt gtcgcttcca agnnaaggac 13621 gaagatgaca atttaattga ttcttacttt gtagttaaga gacacacttt ctctaactac 13681 caacatgaag aaacaattta taatttactt aaggattgtc cagctgttgc taaacatgac 13741 ttctttaagt ttagaataga cggtgacatg gtaccacata tatcacgtca acgtcttact 13801 aaatacacaa tggcagacct cgtctatgct ttaaggcatt ttgatgaagg taattgtgac 13861 acattaaaag nnatacttgt cacatacaat tgttgtgatg atgattattt caatannaag SEQ
Sequence ID Sequence Description NO.
13921 gactggtatg attligtaga aaacccagat atattacgcg tatacgccaa cttaggtgaa 13981 cgtgtacgcc aagctttgtt aaaaacagta caattctgtg atgccatgcg aaatgctggt 14041 attgttggtg tactgacatt agataatcaa gatctcaatg gtaactggta tgatttcggt 14101 gatttcatac aaaccacgcc aggtagtgga gttcctgttg tagattctta ttattcattg 14161 ttaatgccta tattaacctt gaccagggct ttaactgcag agtcacatgt tgacactgac 14221 ttaacaaagc cttacattaa gtgggatttg ttaaaatatg acttcacgga agagaggtta 14281 aaactattg accgttattt taaatattgg gatcagacat accacccaaa ttgtgttaac 14341 tgtt-tggatg acagatgcat tctgcattgt gcaaactt-ta atgttl-tatt ctctacagtg 14401 ttcccaccta caagttttgg accactagtg agaaaaatat ttgttgatgg tgttccattt 14461 gtagtttcaa ctggatacca cttcagagag ctaggtgttg tacataatca ggatgtaaac 14521 ttacatagct ctagacttag altaaggaa ttacttgtgt atgctgctga ccctgctatg 14581 cacgctgctt ctggtaatct attactagat aaacgcacta cgtgcttttc agtagctgca 14641 cttactaaca atgttgctl-t tcaaactgtc aaacccggta att-ttaacaa agacttctat 14701 gactttgctg tgtctaaggg tttctttaag gaaggaagtt ctgttgaatt aaaacacttc 14761 ttctttgctc aggatggtaa tgctgctatc agcgattatg actactatcg ttataatcta 14821 ccaacaatgt gtgatatcag acaactacta tttgtagttg aagttgttga taagtacttt 14881 gattgttacg atggtggctg tattaatgct aaccaagtca tcgtcaacaa cctagacaaa 14941 tcagctggtt ttccatt-taa tagatggggt aaggctagac tttattatga ttcaatgagt 15001 tatgaggatc aagatgcact tttcgcatat acaanacgta atgtcatccc tactataact 15061 caaatgaatc ttaagtatgc cattagtgca aagaatagag ctcgcaccgt agctggtgtc 15121 tctatctgta gtactatgac caatagacag tttcatcaaa aattattgaa atcaatagcc 15181 gccactagag gagctactgt agtaattgga acaagcaaat tctatggtgg ttggcacaac 15241 atgttaaaaa ctgtttatag tgatgtagaa aaccctcacc ttatgggttg ggattatcct 15301 aaatgtgata gagccatgcc taacatgctt agaattatgg cctcacttgt tcttgctcgc 15361 aaacatacaa cgtgttgtag cttgtcacac cgtttctata gattagctaa tgagtgtgct 15421 caagtattga gtgaaatggt catgtgtggc ggttcactat atgttaaacc aggtggaacc 15481 tcatcaggag atgccacaac tgcttatgct aatagtgt[t ttaacatttg tcaagctgtc 15541 acggccaatg ttaatgcact tttatctact gatggtaaca aaattgccga taagtatgtc 15601 cgcaatttac aacacagact ttatgagtgt ctctatagaa atagagatgt tgacacagac 15661 tttgtgaatg agattacgc atatrtgcgt aaacatact caatgatgat actctctgac 15721 gatgctgttg tgtgtttcaa tagcacttat gcatctcaag gtctagtggc tagcataaag 15781 SEQ
Sequence ID Sequence Description NO.
aactttaagt cagttcttta ttatcaaaac aatgattla tgtctgaagc qaaatgttgg 15841 actgagactg accttactaa aggacctcat gaatttlgct ctcaacatac aatgctagtt 15901 aaacagggtg atgattatgt gtaccttcct tacccagatc catcaagaat cctaggggcc 15961 ggctgittig tagatgatat cgtaaaaaca gatggtacac ttatgattga acggttcgtg 16021 tctttagcta tagatgctta cccacttact aaacatccta atcaggagta tgctgatgtc 16081 tttcatttgt acttacaata cataagaaag ctacatgatg agttaacagg acacatgtta 16141 gacatgtatt ctgttatgct tactaatgat aacacttcaa ggtattggga acctgagttt 16201 tatgaggcta tgtacacacc gcatacagtc ttacaggctg ttggggcttg tgttctttgc 16261 aattcacaga cttcattaag atgtggtgct tgcatacgta gaccattctt atgttgtaaa 16321 tgctgttacg accatgtcat atcaacatca cataaattag tcttgtctgt taatccgtat 16381 gtttgcaatg ctccaggttg tgatgtcaca gatgtgactc aactttactt aggaggtatg 16441 agctattatt gtaaatcaca taaaccaccc attaglittc cattgtgtgc taatggacaa 16501 glitaggtt tatataqaaa tacatgtgtt ggtagcgata atgttactga ctttaatgca 16561 attgcaacat gtgactggac aaatgctggt gattacattt tag ctaacac ctgtactgaa 16621 agactcaagc Ettagcagc agaaacgctc aaagctactg aggagacatt taaactgtct 16681 tatggtattg ctactgtacg tgaagtgctg tctgacagag aattacatct ttcatgggaa 16741 gttggtaaac ctagaccacc acttaaccga aattatgtct ttactggtta tcgtgtaact 16801 aaaaacagta aagtacaqat aggagagtac acctttgaaa aaggtgacta tggtgatgct 16861 gttgtttacc gaggtacaac aacttacaaa ttaaatgttg gtgattattt tgtgctgaca 16921 tcacatacag taatgccatt aagtgcacct acactagtgc cacaagagca ctatgttaga 16981 attactggct tatacccaac actcaatatc tcagatgagt tttctagcaa tgttgcaaat 17041 tatcqaaagg ttggtatgca aaagtattct acactccagg gaccacctgg tactggtaag 17101 agtcatittg ctattggcct agctctctac tacccttctg ctcgcatagt gtatacagct 17161 tgctctcatg ccgctgttga tgcactatgt gagaaggcat taaaatattt gcctatagat 17221 aaatgtagta gaattatacc tgcacgtgct cgtgtagagt gttttgataa attcaaagtg 17281 aattcaacat tagaacagta tgtctittgt actgtaaatg cattgcctga gacgacagca 17341 gatatagttg tctttgatga aatttcaatg gccacaaatt atgatttgag tgttgtcaat 17401 gccagattac gtgctaagca ctatgtgtac attggcgacc ctgctcaatt acctgcacca 17461 cgcacattgc taactaaggg cacactagaa ccagaatatt tcaattcagt gtgtagactt 17521 atgaaaacta taggtccaga catgttcctc ggaacttgtc ggcgttgtcc tgctgaaatt 17581 gttgacactg tgagtgcttt ggtttatgat SEQ
Sequence ID Sequence Description NO.
aataagctta aagcacataa agacaaatca 17641 gctcaatgct ttaaaatgtt ttataagggt gttatcacgc atgatgtttc atctgcaatt 17701 aacaggccac anataggcgt ggtaagagaa ttccttacac gtaaccctgc ttggagaaaa 17761 gctgtcttta tttcacctta taattcacag aatgctgtag cctcaaagat tttgggacta 17821 ccaactcaaa ctgttgattc atcacagggc tcagaatatg actatgtcat attcactcaa 17881 accactgaaa cagctcactc ttgtaatgta aacagattta atgttgctat taccagagca 17941 aaagtaggca tactttgcat aatgtctgat agagaccttt atgacapEtt gcaatttaca 18001 agtcttgaaa ttccacgtag gaatgtggca actttacaag ctgaaaatgt aacaggactc 18061 tttaaagatt gtagtaaggt aatcactggg ttacatccta cacaggcacc tacacacctc 18121 agtgttgaca ctaaattcaa aactgaaggt Uatgtgttg acatacctgg catacctaag 18181 gacatgacct atagaagact catctctatg atgggtttta aaatgaatta tcaagttaat 18241 ggttacccta acatgtttat cacccgcgaa gaagctataa gacatgtacg tgcatggatt 18301 ggcttcgatg tcgaggggtg tcatgctact agagaagctg ttggtaccaa tttaccttta 18361 cagctaggtt tttctacagg tgttaaccta gttgctgtac ctacaggtta tgttgataca 18421 cctaataata cagattlitc cagagttagt gctaaaccac cgcctggaga tcaatttaaa 18481 cacctcatac cacttatgta caaaggactt ccttggaatg tagtgcgtat aaagattgta 18541 caaatgttaa gtgacacact taaaaatctc tctgacagag tcgtatttgt cttatgggca 18601 catggctttg agttgacatc tatgaagtat tftgtgaaaa taggacctga gcgcacctgt 18661 tgtctatgtg atagacgtgc cacatgcttt tccactgctt cagacactta tgcctgttgg 18721 catcattcta ttggatttga ttacgtctat aatccgttta tgattgatgt tcaacaatgg 18781 ggttttacag gtaacctaca aagcaac cat gatctgtatt gtcaagtcca tggtaatgca 18841 catgtagcta gttgtgatgc aatcatgact aggtgtctag ctgtccacga gtgctttgtt 18901 aagcgtgttg actggactat tgaatatcct ataattggtg atgaactgaa gattaatgcg 18961 gcttgtagaa aggttcaaca catggttgtt aaagctgcat tattagcaga caaattccca 19021 gttcttcacg acattggtaa ccctaaagct attaagtgtg tacctcaagc tgatgtagaa 19081 tggaagttct atgatgcaca gccttgtagt gacaaagctt ataaaataga agaattattc 19141 tattcttatg ccacacattc tgacaaattc acagatggtg tatgcctatt ttggaattgc 19201 aatgtcgata gatatcctgc taattccatt gtttgtagat ttgacactag agtgctatct 19261 aaccttaact tgcctggttg tgatggtggc agtttgtatg taaataaaca tgcattccac 19321 acaccagctt ttgataaang tgcttttgtt aatttaaaac aattaccatt tttctattac 19381 tctgacagtc catgtgagtc tcatggaaaa caagtagtgt cagatataga ttatgtacca 19441 ctaaagtctg ctacgtgtat SEQ
Sequence ID Sequence Description NO.
aacacgttgc aatttaggtg gtgctgtctg tagacatcat 19501 gctaatgagt acagattgta tctcgatgct tataacatga tgatctcagc tggctttagc 19561 ttgtgggttt acaaacaatt tgatacttat aacctctgga acacttttac aagacttcag 19621 agtttagaaa atgtggcttt taatgttgta aataagggac actttgatgg acaacagggt 19681 gaagtaccag tttctatcat taataacact gtttacacaa aagttgatgg tgttgatgta 19741 gaattgifig aaaataaaac aacattacct gttaatgtag catttgagct ttgggctaag 19801 cgcaacatta aaccagtacc agaggtgaaa atactcaata atttgggtgt ggacattgct 19861 gctaatactg tgatctggga ctacaaaaga gatgctccag cacatatatc tactattggt 19921 gtagttcta tgactgacat agccaagaaa ccaactgaaa cgatttgtgc accactcact 19981 gtctlattg atggtagagt tgatggtcaa gtagacttat ttagaaatgc ccgtaatggt 20041 gttcttatta cagaaggtag tgttonaggt ttacaaccat ctgtaggtcc caaacaagct 20101 agtcttaatg gagtcacatt aattggagaa gccgtaaaaa cacagttcaa ttattataag 20161 aaagttgatg gtgttgtcca acaattacct gaaacttact ttactcagag tagaaattta 20221 caagaattta aacccaggag tcanatggaa attgatttct tagaattagc tatggatgaa 20281 ttcattgaac ggtataaatt agaaggctat gccttcgaac atatcgttta tggagatttt 20341 agtcatagtc agttaggtgg tttacatcta ctgattggac tagctaaacg ilitaaggaa 20401 tcaccittig aattagaaga ititattcct atggacagta cagttaaaaa ctatttcata 20461 acagatgcgc aaacaggttc atctaagtgt gtgtgttctg ttattgattt attacttgat 20521 galitigttg anataatana atcccaagat ttatctgtag tttctaaggt tgtcaaagtg 20581 actattgact atacagaaat ttcatttatg attggtgta aagatggcca tgtagaaaca 20641 ttttacccaa aattacaatc tagtcaagcg tggcaaccgg gtgttgctat gcctaatctt 20701 tacaaaatgc aaagaatgct attagaaaag tgtgaccttc aaaattatgg tgatagtgca 20761 acattaccta aaggcataat gatgaatgtc gcaaaatata ctcaactgtg tcaatattta 20821 aacacattaa cattagctgt accctataat atgagagtta tacattligg tgctggttct 20881 gataaaggag ttgcaccagg tacagctgtt ttaagacagt ggttgcctac gggtacgctg 20941 cttgtcgatt cagatcttaa tgactttgtc tctgatgcag attcaacttt gattggtgat 21001 tgtgcaactg tacatacagc taataaatgg gatctcatta ttagtgatat gtacgaccct 21061 aagactaa a atgttacaaa agaaaatgac tctaaagagg gttttttcac ttacatttgt 21121 gggtttatac aacaaaagct agctcttgga ggttccgtgg ctataaagat aacagaacat 21181 tcttggaatg ctgatcttta taagctcatg ggacacttcg catggtggac agcctrtgtt 21241 actaatgtga atgcgtcatc atctgaagca tlittaattg gatgtaatta tcttggcaaa 21301 ccacgcgaac aaatagatgg SEQ
Sequence ID Sequence Description NO.
ttatgtcatg catgcaaatt acatattttg gaggaataca 21361 aatccaattc agttgtcttc ctattcttta tttgacatga gtaaatttcc ccttaaatta 21421 aggggtactg ctgttatgtc tttaaaagaa ggtcaaatca atgatatgat tttatctctt 21481 cttagtaaag gtagacttat aattagagaa aacaacagag ttgttatttc tagtgatgtt 21541 cttgttaaca actaaacgaa caatgtttgt ttttcttgtt ttattgccac tagtctctag 21601 tcagtgtgtt aatcttacaa ccagaactca attaccccct gcatacacta attctttcac 21661 acgtggtgtt tattaccctg acaaag tilt cagatcctca galtacatt caactcagga 21721 cttgactta cctLicalL ccaatgttac ttggttccat gctatacatg tctctgggac 21781 caatggtact aagaggtttg ataaccctgt cctaccattt aatgatggtg Matittgc 21841 ttccactgag aagtctaaca taataagagg ctggattat ggtactactt tagattcgaa 21901 gacccagtcc ctacttattg ttaataacgc tactaatgtt gttattaaag tctgtgaatt 21961 tcaattttgt aatgatccat ttttgggtgt ttattaccac aaanacaaca aaagttggat 22021 gganagtgag ttcagagttt attctagtgc gaataattgc acttttgaat atgtctctca 22081 gcctIttctt atggaccttg aaggaaaaca gggtaatttc aaaaatctta gggaatttgt 22141 gtttaagaat attgatggtt aLittaaaat atattctaag cacacgccta ttaatttagt 22201 gcgtgatctc cctcagggtt tttcggcttt agaaccattg gtagatttgc caataggtat 22261 taacatcact aggtttcaaa ctttacttgc tttacataga agttatttga ctcctggtga 22321 ttcttcttca ggttggacag ctggtgctgc agcttattat gtgggttatc ttcaacctag 22381 gactlacta ttaaaatata atgaaaatgg aaccattaca gatgctgtag actgtgcact 22441 tgaccctctc tcagaaacaa agtgtacgtt gaaatccttc actgtagaaa aaggaatcta 22501 tcaaacttct aactttagag tccaaccaac agaatctatt gttagatttc ctaatattac 22561 aaacttgtgc cctaggtg aagtLittaa cgccaccaga tttgcatctg tttatgcttg 22621 gaacaggaag agaatcagca actgtgttgc tgattattct gtcctatata attccgcatc 22681 attttccact tttaagtgtt atggagtgtc tcctactaaa ttaaatgatc tctgctttac 22741 taatgtctat gcagattcat ttgtaattag aggtgatgaa gtcagacaaa tcgctccagg 22801 gcaaactgga aagattgctg attataatta taaattacca gatgatttta caggctgcgt 22861 tatagcttgg aattctaaca atcttgattc taaggttggt ggtaattata attacctgta 22921 tagattgttt aggaagtcta atctcaaacc Ltttgagaga gatatttcaa ctgaaatcta 22981 tcaggccggt agcacacctt gtaatggtgt tgaagglitt aattgttact ttcctttaca 23041 atcatatggt ttccaaccca ctaatggtgt tggttaccaa ccatacagag tagtagtact 23101 ttcttttgaa cttctacatg caccagcaac tgtttgtgga SEQ
Sequence ID Sequence Description NO.
cctaaaaagt ctactaattt 23161 ggttaaaaac aaatgtgtca atttcaactt caatggttta acaggcacag gtgttcttac 23221 tgagtctaac aaanagtttc tgcctttcca acaatttggc agagacattg ctgacactac 23281 tgatgctgtc cgtgatccac agacacttga gattcttgac attacaccat gttcttttgg 23341 tggtgtcagt gttataacac caggaacaaa tacttctaac caggttgctg ttctttatca 23401 ggatgttaac tgcacagaag tccctgttgc tattcatgca gatcaactta ctcctacttg 23461 gcgtgtttat tctacaggtt ctaatgtttt tcaaacacgt gcaggctgtt taataggggc 23521 tgaacatgtc aacaactcat atgagtgtga catacccatt ggtgcaggta tatgcgctag 23581 ttatcagact cagactaatt ctcctcggcg ggcacgtagt gtagctagtc aatccatcat 23641 tgcctacact atgtcacttg gtgcagaaaa ttcagttgct tactctaata actctattgc 23701 catacccaca aattttacta ttagtgttac cacagaaatt ctaccagtgt ctatgaccaa 23761 gacatcagta gattgtacaa tgtacatttg tggtgattca actgaatgca gcaatctttt 23821 gttgcaatat ggcaglittt gtacacaatt anaccgtgct ttaactggaa tagctgttga 23881 acaagacaaa aacacccaag aagitlitgc acaagtcaaa caaatttaca aaacaccacc 23941 aattaaagat tttggtggtt ttaattlitc acaaatatta ccagatccat caaaaccaag 24001 caagaggtca tttattgaag atctac Litt caacaaagtg acacttgcag atgctggctt 24061 catcaaacaa tatggtgatt gccttggtga tattgctgct agagacctca tttgtgcaca 24121 anagtttaac ggccttactg ttttgccacc tttgctcaca gatgaaatga ttgctcaata 24181 cacttctgca ctgttagcgg gtacaatcac ttctggttgg acctttggtg caggtgctgc 24241 attacaaata ccatttgcta tgcaaatggc ttataggttt aatggtattg gagttacaca 24301 gaatgttctc tatgagaacc aaaaattgat tgccaaccaa tttaatagtg ctattggcaa 24361 aattcaagac tcactttctt ccacagcaag tgcacttgga aaacttcaag atgtggtcaa 24421 ccarmatgca caagctttaa acacgcttgt tanacaactt agctccaatt ttggtgcaat 24481 ttcaagtgtt ttaaatgata tcctttcacg tcttgacaaa gttgaggctg aagtgcaaat 24541 tgataggttg atcacaggca gacttcaaag tttgcagaca tatgtgactc aacaattaat 24601 tagagctgca gaaatcagag cttctgctaa tcttgctgct actaaaatgt cagagtgtgt 24661 acttggacaa tcaaanagag ttgailittg tggaaagggc tatcatctta tgtccttccc 24721 tcagtcagca cctcatggtg tagtcttctt gcatgtgact tatgtccctg cacaagaaaa 24781 gaacttcaca actgctcctg ccatttgtca tgatggaaaa gcacactttc ctcgtgaagg 24841 tgtctttgtt tcaaatggca cacactggtt tgtaacacaa aggaaltitt atgaaccaca 24901 aatcattact acagacaaca catttgtgtc tggtaactgt gatgttgtaa taggaattgt 24961 caacaacaca gtttatgatc ctttgca,9cc tgaattagac tcattcaagg SEQ
Sequence ID Sequence Description NO.
aggagttaga 25021 taaatattlt aagaatcata catcaccaga tgttgattta ggtgacatct ctggcattaa 25081 tgcttcagtt gtaaacattc aaaaagaaat tgaccgcctc aatgaggttg ccaagaattt 25141 aaatgaatct ctcatcgatc tccaagaact tggaaagtat gagcagtata taaaatggcc 25201 atggtacatt tggctaggtt ttatagctgg cttgattgcc atagtaatgg tgacaattat 25261 gctttgctgt atgaccagtt gctgtagttg tctcaagggc tgttgttctt gtggatcctg 25321 ctgcaaattt gatgaagacg actctgagcc agtgctcaaa ggagtcaaat tacattacac 25381 ataaacgaac ttatggattt gtttatgaga atcttcacaa ttggaactgt aactttgaag 25441 caaggtgaaa tcaaggatgc tactccttca gattligttc gcgctactgc aacgataccg 25501 atacaagcct cactccatt cggatggctt attgttggcg ttgcacttct tgctg UM 25561 cagagcgctt ccaaaatcat aaccctcaaa aagagatggc aactagcact ctccaagggt 25621 gttcact-ttg tttgcaactt gctgttgttg tttgtaacag tttactcaca ccttttgctc 25681 gttgctgctg gccttgaagc ccctlitctc tatctttatg ctttagtcta cttcttgcag 25741 agtataaqct ttgtaagaat aataatgagg ct-ttggcttt gctggapatg ccgttccaaa 25801 aacccattac tttatgatgc caactatitt ctttgctggc atactaattg ttacgactat 25861 tgtatacctt acaatagtgt aacttcttca attgtcatta cttcaggtga tggcacaaca 25921 agtcctattt ctgaacatga ctaccagatt ggtggttata ctgaaaaatg ggaatctgga 25981 gtaaaagact gtgttgtatt acacagttac ttcacttcag actattacca gctgtactca 26041 actcaattga gtacagacac tggtgttgaa catgttacct tcttcatcta caataaaatt 26101 gttgatgagc ctgaagaaca tgtccaaatt cacacaatcg acggttcatc cggagttgtt 26161 aatccagtaa tggaaccaat ttatgatgaa ccgacgacga ctactagcgt gcctttgtaa 26221 gcacaagctg atgagtacga acttatgtac tcattcgttt cggaagagac aggtacgtta 26281 atagttaata gcgtacttct ttlicttgct ttcgtggtat tcttgctagt tacactagcc 26341 atccttactg cgcttcgatt gtgtgcgtac tgctgcaata ttgttaacgt gagtcttgta 26401 aaaccttctt tttacgttta ctctcgtgtt aaaaatctga attcttctag agttcctgat 26461 cttctggtct aaacgaacta aatattatat tag ittitct gtttggaact ttaatittag 26521 ccatggcaga ttccaacggt actattaccg ttgaagagct taaanagctc cttgaacaat 26581 ggaacctagt aataggtttc ctattcctta catggatttg tcttctacaa tttgcctatg 26641 ccaacaggaa taggittlig tatataatta agttaatttt ectctggctg ttatggccag 26701 taactttagc ttglitigtg cttgctgctg tttacagaat anattggatc accggtggaa 26761 ttgctatcgc aatggcttgt cttgtaggct tgatgtggct cagctacttc attgcttctt 26821 tcagactgtt tgcgcgtacg cgttccatgt ggtcattcaa tccagaaact aacattcttc 26881 SEQ
Sequence ID Sequence Description NO.
tcaacgtgcc actccatggc actattctga ccagaccgct tctagaaagt gaactcgtaa 26941 tcggagctgt gatccttcgt ggacatcttc gtattgctgg acaccatcta ggacgctgtg 27001 acatcaagga cctgcctaaa gaaatcactg ttgctacatc acgaacgctt tcttattaca 27061 aattgggagc ttcgcagcgt gtagcaggtg actcaggttt tgctgcatac agtcgctaca 27121 ggattggcaa ctataaatta aacacagacc attccagtag cagtgacaat attgctttgc 27181 ttgtacagta agtgacaaca gatgtttcat ctcgttgact ttcaggttac tatagcagag 27241 atattactaa ttattatgag gactittaaa gtttccattt ggaatcttga ttacatcata 27301 aacctcataa ttaaaaatit atctaagtca ctaactgaga ataaatattc tcaattagat 27361 gaagagcaac caatggagat tgattaaacg aacatgaaaa ttattctitt cttggcactg 27421 ataacactcg ctacttgtga gctttatcac taccaagagt gtgttagagg tacaacagta 27481 clataaaag aaccttgctc ttctggaaca tacgagggca attcaccatt tcatcctcta 27541 gctgataaca aatttgcact gacttgcttt agcactcaat ttgalligc ttgtcctgac 27601 ggcgtaaaac acgtctatca gttacgtgcc agatcagttt cacctaact gttcatcaga 27661 caagaggaag ttcaagaact ttactctcca alltlictta ttgttgcggc aatagtgttt 27721 ataacacttt gcttcacact caaaagaaag acagaatgat tgaactttca ttaattgact 27781 tctatttgtg ctttttagcc tttctgctat tecttgitti aattatgctt attatctttt 27841 ggttctcact tgaactgcaa gatcataatg aaacttgtca cgcctaaacg aacatgaaat 27901 ttcttglitt cttaggaatc atcacaactg tagctgcatt tcaccaagaa tgtagtttac 27961 agtcatgtac tcaacatcaa ccatatgtag ttgatgaccc gtgtcctatt cacttctatt 28021 ctaaatggta tattagagta ggagctagaa aatcagcacc tttaattgaa ttgtgcgtgg 28081 atgaggctgg ttctaaatca cccattcagt acatcgatat cggtaattat acagtttcct 28141 gtttaccttt tacaattaat tgccaggaac ctaaattggg tagtcttgta gtgcgttgtt 28201 cgttctatga agactatta gagtatcatg acgttcgtgt tgttttagat ttcatctaaa 28261 cgaacaaact aaaatgtctg ataatggacc ccaaaatcag cgaaatgcac cccgcattac 28321 gtttggtgga ccctcagatt caactggcag taaccagaat ggagaacgca g-tggggcgcg 28381 atcaaaacaa cgtcggcccc aaggtttacc caataatact gcgtcttggt tcaccgctct 28441 cactcaacat ggcaaggaag accttanatt ccctcgagga caaggcgttc caattaacac 28501 caatagcagt ccagatgacc aaattggcta ctaccgaaga gctaccagac gaattcgtgg 28561 tggtgacggt aaaatgaaag atctcagtcc aagatggtat ttctactacc taggaactgg 28621 gccagaagct ggacttccct atggtgctaa caaagacggc atcatatggg ttgcaactga 28681 gggagccttg aatacaccaa aagatcacat tggcacccgc aatcctgcta acaatgctgc 28741 aatcgtgcta caacttcctc aaggaacaac SEQ
Sequence ID Sequence Description NO.
attgccaaaa ggcttctacg cagaagggag 28801 cagaggcggc agtcaagcct cttctcgttc ctcatcacgt agtcgcaaca gttcaagaaa 28861 ttcaactcca ggcagcagta ggggaacttc tcctgctaga atggctggca atggcggtga 28921 tgctgctctt gctttgctgc tgcttgacag attgaaccag cttgagagca aaatgtctgg 28981 taaaggccaa caacaacaag gccaaactgt cactaagaaa tctgctgctg aggcttctaa 29041 gaagcctcgg caaaaacgta ctgccactaa agcatacaat gtaacacaag ctttcggcag 29101 acgtggtcca gaacaaaccc aaggaaattt tggggaccag gaactaatca gacaaggaac 29161 tgattacaaa cattggccgc aaattgcaca atttgccccc agcgcttcag cgttcttcgg 29221 aatgtcgcgc attggcatgg aagtcacacc ttcgggaacg tggttgacct acacaggtgc 29281 catcaaattg gatgacaaag atccaaattt caaagatcaa gtcaltilgc tgaataagca 29341 tattgacgca tacaaaacat tcccaccaac agagcctaaa aaggacaaaa agaagaaggc 29401 tgatgaaact caagccttac cgcagagaca gaagaaacag caaactgtga ctcttcttcc 29461 tgctgcagat ttggatgatt tctccaaaca attgcaacaa tccatgagca gtgctgactc 29521 aactcaggcc taaactcatg cagaccacac aaggcagatg ggctatataa acgattcgc 29581 ttttccgttt acgatatata gtctactctt gtgcagaatg aattctcgta actacatagc 29641 acaagtagat gtagttaact ttaatctcac atagcaatct ttaatcagtg tgtaacatta 29701 gggaggactt gaaagagcca ccacattttc accgaggcca cgcggagtac gatcgagtgt 29761 acagtgaaca atgctaggga gagctgccta tatggaagag ccctaatgtg taaaattaat 29821 tttagtagtg ctatccccat gtgattltaa tagcttctta ggagaatgac aaaaaaaaaa 29881 aaaaaaaaaa aaaaaaaaaa aaa ME S I ,VPGFNEKTHVQI , ST ;PVT ,QVR DVI ,VR GEGDS
VEEVLSEARQHLKDGTCGLVEVEKGVLPQLEQPY
VFIKRSDARTAPHGHVMVELVAELEGIQYGRSGE

TLGVLVPHVGEIPVAYRKVLLRKNGNKGAGGHS
Wuhan seafood YGADLKSFDLGDELGTDPYEDFQEN
market pneumonia WNTKHSSGVTRELMRELNGGAYTRYVDNNFCGP
virus isolate DGYPLECIKDLLARAGKASCTLSEQLDFIDTKRGV
Wuhan-Hu-1 YCCREHEHEIAWYTERSEKSYELQTPFEIKLAKKF
genomic sequence 2 DTENGECPNFVFPLNSIIKTIQPRVEKKKLDGFMG
(GenBank:
RIRSVYPVASPNECNQMCLSTLMKCDHCGETSWQ
MN908947.3janu TGDFVKATCEFCGTENLTKEGATTCGYLPQNAVV
ary 23, 2020) ¨
KIY CPACHN S E V GPEHSLAEYHN ESGLKTILRKGG
amino acid RTIAFGGCVESYVGCHNKCAYWVPRASANIGCNH
translation TGVVGEGSEGLNDNL
LEILQKEKVNINIVGDFKLNEEIAIILASFSASTSAF
VETVKGLDYKAFKQIVESCGNEKVTKGKAKKGA

SEQ
Sequence ID Sequence Description NO.
WNIGEQKSILSPLYAFASEAARVVRSIFSRTLETAQ
N SVRVLQKAAITILD GIS QYSLRLIDAMMFTSDLA
TNNLVVMAYITGGVVQLTSQWLTNIFGTVYEKLK
PVLDWLEEKFKEGVEFLRDGWEIVKFISTCACEIV
GGQIVTCAKEIKE S V QTFFKLVNKFLALCAD SIIIG
GAKLKALNLGETFVTHSKGLYRKCVKSREETGLL
MPLKAPKEIIFLEGETLPTEVLTEEVVLKTGDLQPL
EQPTSEAVEAPLVGTPVCINGLMLLEIKDTEKYCA
LAPNMMVTNNTFTLKGGAPTKVTFGDDTVIEVQ
GYKSVNITFELDERIDKVLNEKC SAYTVELGTEVN
EFACVVADAVIKTL QPV S ELLTPLGIDLDEW S MA
TYYLFDESGEFKLASHMY C SFYPPDEDEEEGDCE
EEEFEPSTQYEYGTEDDYQGKPLEFGATSAALQPE
EEQEEDWLDDDS QQTVGQQDGSEDNQTTTIQTIV
EV QPQLEMELTP V V QTIEVN SFSGYLKLTDN VY IK
NADIVEEAKKVKP'TVVVNAANVYLKHGGGVAG
A LNK A TNNAMQVESDDYIATNGPLKVGGSCVLS
GHNLAKHCLHVVGPN VNKGEDIQLLKSAYENFN
QHEVLLAPLLSAGIFGADPIHSLRVCVDTVRTNVY
LAVFDKNLYDKLVSSFLEMKSEKQVEQKIAEIPKE
EVKPFITESKPSVEQRKQDDKKIKACVEEVTTTLE
ETKFLTENLLLYIDINGNLHPD S A TLV SDIDITFLK
KDAPYIVGDVVQEGVLTAVVIPTKKAGGTTEMLA
KALRKVPTDNYITTYPGQGLNGYTVEEAKTVLKK
CK S A FYILP SIISNEKQEILG'TVSWNLREMLAHAEE
TRKLMPVCVETKAIVSTIQRKYKGIKIQEGVVDYG
ARFYFYTSKTTVASLINTLNDLNETLVTMPLGYVT
HGLNLEEAARYMRSLKVPATVSVS SPDAVTAYN
GYLTSS SKTPEEHFIETISLAGSYKDWSYSGQ STQL
GIEFLKRGDKS VYYTSN PTTFHLDGEVITFDNLKT
LLSLREVRTIKVFT'TVDN1NLHTQVVDMSMTYGQ
QFGPTYLDGADVTKIKPHNSHEGKTFYVLPNDDT
LRVEAFEYYHTTDPSFLGRYMSALNHTKKWKYP
QVNGLTSIKWADNNCYLATALLTLQQIELKFNPP
AL QDAYYRARAGEAANFCALILAYCNKTVGELG
DVRETMSYLFQHANLDSCKRVLNVVCKTCGQQQ
TTLKGVEAVMYMGTLSYEQFKKGVQIPCTCGKQ
A TKYLVQ QE S PFVMM S A PP A QYELKHGTFTC A SE
YTGNYQ CGHYKHITSKETLYCIDGALLTKSSEYK
GPITDVFYKENSYTTTIKPVTYKLDGVVCTEIDPK
LDNYYKKDNSYFTEQPIDLVPNQPYPNASFDNFK
FVCDNIKFADDLNQLTGYKKPASRELKVTFFPDL
NGDVVAIDYKHYTPSFKKGAKLLHKPI
ATNKATYKPNTWCIRCLWSTKPVETSNSFDVLKS
EDAQGMDNLACEDLKPVSEEVVENPTIQKDVLEC

SEQ
Sequence ID Sequence Description NO.
NVKTTEVVGDIILKPANNSLKITEEVGHTDLMAA
YVDNSSLTIKKPNELSRVLGLKTLATHGLAAVNS
VPWDTIANYAKPFLNKVV STTTNIVTRCLNRV CT
NYMPYFFTLLLQLCTFTRSTNSRIKASMPTTIAKN
TVKSVGKFCLEASFNYLKSPNFSKLINIIIWFLLLS

NSTNVTIATYCTGSIPCSVCLSGLDSLDTYPSLETI
QITIS SFKWDLTAFGLVAEWFLAYILFTRFFYVLG
LAAIMQLFFSYFAVHFISNSWLMWLIINLVQMAPI
SAMVRMYIFFASFYYVWKSYVHVVDGCNSSTCM
MCYKRNRATRVECTTIVNGVRRSFYVYANGGKG

KRPINPTDQSSYIVDSVTVKNGSIHLYFDKAGQKT
YERHSLSHFVNLDNLRANNTKGSLPINVIVFDGKS
KCEES SAKSASVYY SQLMCQPILLLDQALVSDVG
DSAEVAVKMFDAYVNTF SSTFNVPMEKLKTLVA
TAEAELAKNVSLDNVLSTFISA A RQGFVDSDVET

NMTPRDLGACIDC SARHINAQVAKSHNIALIWNV
KDFMSLSEQLRKQIRSAAKKNNLPFKLTCATTRQ
VVNVVTTKIALKGGKIVNNWLKQLIKVTLVFLFV
A A IFYLITPVHVMSKHTDF S SEIIGYK A IDGGVTRD
IA STDTCFANKHADFDTWF S QRGGSYTNDKACPL
IAAVITREVGFVVPGLPGTILRTTNGDFLHFLPRVF
SAVGNICY'TPSKLIEYTDFATSACVLAAECTIFKD
A SGKPVPYCYDTNVLEGSVAYE SLRPDTRYVLM
DGSIIQFPNTYLEGSVRVVTTFDSEYCRHGTCERS
EAGVCVSTSGRWVLNNDYYRSLPGVFCGVDAVN
LLTNMFTPLIQPIGALDISA SIVAGGIVAIVVTC LA
YYFMRFRRAFGEY SHVVAFNTLLFLMSFTVLCLT
PVYSFLPGVYSVIYLYLTFYLTNDVSFLAHIQWM
VMFTPLVPFWITIAYIICISTKHFYWFFSNYLKRRV
VFNGV SF STFEEAALCTFLLNKEMYLKLRSDVLLP
LTQYNRYLALYNKYKYF SGAMDTTSYREAAC CH
LAKALNDFSNSGSDVLYQPPQTSITSAVLQSGFRK
MAFPSGKVEGCMVQVTCGTTTLNGLWLDDVVY
CP RHVICT SEDMLNPNYEDLLIRKSNHNFLVQAG
NVQLRVIGHSMQNCVLKLKVDTANPKTPKYKFV
RIQ PGQTF SVLACYNGSP SGVYQ CA MRPNFTIKGS
FLNGS CGSVGFNIDYDCV SF CYMITHMELPTGVHA
GTDLEGNFYGPFVDRQTAQAAGTDTTITVNVLA
WLYAAVINGDRWFLNRFTTTLNDFNLVAMKYNY
EPLTQDHVDILGPLSAQTGIAVLDMCASLKELLQN
GMNGRTILGSALLEDEFTPFDVVRQC SGVTFQ SA
VKRTIKGTHHWLLLTILTSLLVLVQSTQWSLFFFL

SEQ
Sequence ID Sequence Description NO.
YENAFLPFAMGIIAMSAFAMMFVKHKHAFLCLFL
LP S LATVAYFNMVYMPA SWVMRIMTWLDMVDT
SL S GFKLKD CVMYA SAVVLLILMTARTVYDD GA
RRVWTLMNVLTLVYKVYYGNALDQAISMWALII
SVTSNYSGVVTTVMFLARGIVFMCVEYCPIFFITG
NTLQCIMLVYCFLGYFCTCYFGLFCLLNRYFRLTL
GVYDYLVSTQEFRYMNSQGLLPPKNSIDAFKLNI
KLLGVGGKPCIKVATVQSKMSDVKCTSVVLL SVL
QQLRVES SSKLWAQCVQLHNDILLAKDTTEAFEK
MVSLLSVLLSMQGAVDINKLCEEMLDNRATLQAI
A SEF S S LP SYAAFATAQEAYEQAVANGD SEVVLK
KLKKSLN VAKSEFDRDAAMQRKLEKMADQAMT
QMYKQARSEDKRAKVTSAMQTMLFTMLRKLDN
DALNNIINNARDGCVPLNIIPLTTAAKLMVVIPDY
NTYKN TCDGTTFTYASALWEIQQVVDADSKIVQL
SEISMDNSPNLAWPLIVTALRANSAVKLQNNEL SP
VALRQMS CA AGTTQTACTDDNALAYYNTTKGGR
FVLALLSDLQDLKWARFPKSDGTGTIYTELEPPCR
FVTDTPKGPKVKYLYFIKGLNNLNRGMVLGSLAA
TVRLQAGNATEVPAN S TVL SF CAFAVDAAKAYK
DYLASGGQPITNCVKMLCTHTGTGQAITVTPEAN
MD QESFGGA SC CLYCRCHIDHPNPKGFCDLKGKY
VQIPTTCANDPVGFTLKN'TVCTVCGMWKGYGC S
CD QLREPMLQ SADA Q SFLNRVCGVSAARLTPCGT
GTSTDVVYRAFDIYNDKVA GFAKFLK'TNCCRFQE
KDEDDNLID SYFVVKRHTFSNYQHEETIYNLLKD
CPAVAKHDFFKFRIDGDMVPHISRQRLTKYTMAD
LVYALRHFDEGNCDTLKEILVTYNCCDDDYFNKK
DWYDFVENPDILRVYANLGERVRQALLKTVQFC
DAMRNAGIVGVLILDN QDLNGN W YDFGDFIQ TT
PG SGVPVV
D SYY S LLMPILTLTR A LTA E SHVDTDLTKPYIKWD
LLKYDFTEERLKLFDRYFKYWDQTYHPNCVNCL
DDRCILHCANFNVLFSTVFPPTSFGPLVRKIFVDG
VPFVVSTGYHFRELGVVHNQDVNLHSSRLSFKEL
LVYAADPAMHAASGNLLLDKRTTCFSVAALTNN
VAFQTVKPGNFNKDFYDFAVSKGFFKEGS SVELK
HFFFAQDGNA A IS DYDYYRYNLPTMC DIRQ LLFV
VEVVDKYFDCYDGGCINANQVIVNNLDKSAGFPF
NKWGKARLYYDSMSYEDQDALFAYTKRNVIPTIT
QMNLKYAISAKNRARTVAG V SIC STMTNRQFHQ
KLLKSIAATRGATVVIGTSKFYGGWHNMLKTVYS
DVENPHLMGWDYPKCDRAMPNMLRIMASLVLA
RKHTTCCSL SHRFYRLANECAQVLSEMVMCGGS
LYVKPGGTSSGDATTAYANSVFNICQAVTANVNA

SEQ
Sequence ID Sequence Description NO.
LLSTDGNKIADKYVRNLQHRLYECLYRNRDVDT
DFVNEFYAYLRKHFSMMILSDDAVVCFNSTYASQ
GLVASIKNFKSVLYYQNNVFMSEAKCWTETDLT
KGPHEFCSQHTMLVKQGDDYVYLPYPDPSRILGA
GCFVDDIVKTDGTLMIERFVSLAIDAYPLTKHPNQ
EYADVFHLYLQYIRKLHDELTGHMLDMY S V MLT
NDNTSRYWEPEFYEAMYTPHTVLQAVGACVLCN
SQTSLRCGACIRRPFLCCKCCYDHVISTSHKLVLS
VNPYVCNAPGCDVTDVTQLYLGGMSYYCKSHKP
PISFPLCANGQVFGLYKNTCVGSDNVTDFNAIATC
DWTNAGDYILANTCTERLKLFAAETLKATEETFK
L SY GIATVREVL SDRELHL SW EV GKPRPPLN RN Y
VFTGYRVTKNSKVQIGEYTFEKGDYGDAVVYRG
TTTYKLNVGDYFVLTSHTVMPLSAPTLVPQEHYV
RITGLYPTLNISDEFSSN VAN Y QKVGMQKY STLQ
GPPGTGKSHFAIGLALYYPSARIVYTACSHAAVD
ALCEKALKYLPIDKC SRIIPARARVECFDKFKVNS
TLEQYVFCTVNALPETTADIVVFDEISMATNYDLS
VVNARLRAKHYVYIGDPAQLPAPRTLLTKGTLEP
EYFNSVCRLMKTIGPDMFLGTCRRCPAEIVDTV SA
LVYDNKLKAHKDKSAQCFKMFYKGVITHDVS SAT
NRPQIGVVREFLTRNPAWRK AVFISPYNSQNAVA
SKILGLPTQTVDSSQGSEYDYVIFTQTTETAHSCN
VNRFNVAI I RAKVGILCIMSDRDLYDKLQFTSLEI
PRRNVA TLQ A ENVTGLFKDC SKVITGLHPTQ A PT
HLSVDTKFKTEGLCVDIPGIPKDMTYRRLISMMGF
KMNYQVNGYPNMFITREEAIRHVRAWIGFDVEG
CHATREAVGTNLPLQLGFSTGVNLVAVPTGYVDT
PNNTDFSRVSAKPPPGDQFKHLIPLMYKGLPWNV
VRIKIVQMLSDILKNLSDRVVFVLWAHGFELTSM
KYFVKIGPERTCCLCDRRATCF STA SDTYACWHEI
SIGFDYVYNPFMIDVQQWGFTGNLQSNHDLYCQ
VHGNAHVASCDAIMTRCLAVHECFVKRVDWTIE
YPIIGDELKINAACRKVQHMVVKAALLADKFPVL
HDIGNPKAIKCVPQADVEWKFYDAQPCSDKAYKI
EELFYSYATHSDKFTDGVCLFWNCNVDRYPAN ST
VCRFDTRVLSNLNLPGCDGGSLYVNKHAFHTPAF
DK SAFVNLKQLPFFYYSDSPCESHGKQVVSDIDY
VPLKSATCITRCNLGGAVCRHHANEYRLYLDAYN
MMISAGFSLWVYKQFDTYNLWNTFTRLQSLENV
AFNVVNKGHFDGQQGEVPVSIINNTVYTKVDGV
DVELFENKTTLPVNVAFELWAKRNIKPVPEVKILN
NLGVDIAANTVIWDYKRDAPAHISTIGVCSMTDIA
KKPTETICAPLTVFFDGRVDGQVDLFRNARNGVLI
TEGSVKGLQPSVGPKQASLNGVTLIGEAVKTQFN

SEQ
Sequence ID Sequence Description NO.
YYKKVDGVVQQLPETYFTQSRNLQEFKPRSQMEI
DFLELAMDEFIERYKLEGYAFEHIVYGDFSHSQLG
GLHLLIGLAKRFKESPFELEDFIPMDSTVKNYFITD
AQTGSSKCVCSVIDLLLDDFVEIIKSQDLSVVSKV
VKVTIDYTEISFMLWCKDGHVETFYPKLQSSQAW
QPGVAMPNLYKMQRMLLEKCDLQNYGDSATLP
KGIMMNVAKYTQLCQYLNTLTLAVPYNMRVIHF
GAGSDKGVAPGTAVLRQWLPTGTLLVDSDLNDF
VSDADSTLIGDCATVHTANKWDLIISDMYDPKTK
NVTKENDSKEGFFTYICGFIQQKLALGGSVAIKITE
HSWNADLYKLMGHFAWWTAFVTNVNASSSEAF
LIGCNYLGKPREQIDGYVMHANYIFWRNTNPIQLS
SYSLFDMSKFPLKLRGTAVMSLKEGQINDMILSLL
SKGRLIIRENNRVVISSDVLVNN
mfvflvllpl vssqcvnitt rtqlppaytn sftrgvyypd kvfrssvlhs tqdlflpffs 61 nvtwfhaihv sgtngtkrfd npvlpfndgv yfasteksni irgwifgttl dsktqslliv 121 nnatnvvikv cefqfcndpf lgvyyhknnk swmesefrvy ssannctfey vsqpflmdle181 gkqgnfknlr efvflmidgy fkiyskhtpi nlvrdlpqgf saleplvdlp iginitrfqt 241 llallirsylt pgdsssgwta gaaayyvgyl qprtfllkyn engtitdavd caldplsetk 301 ctlksftvck giyqtsnfry qptcsivrfp nitnlcpfgc vfilatrfasv yawnrkrisn 361 cvadysvlyn sasfstfkcy gvsptklndl cftnvyadsf virgdevrqi apgqtgkiad 421 ynyklpddft gcviawnsnn ldskvggnyn ylvrlfrksn lkpferdist surface eiyqagstpc 481 ngvegfitcyf plqsygfqpt ngvgyqpyry glycoprotein vvlsfellha patvcgpkks tnlvknkcvn 541 frifngligtg 1SARS-CoV-2 vltesnkkfl pfqqfgrdia dttdavrdpq tleilditpc sfggvsvitp Wuhan seafood 601 gtntsnqvav lyqdvnctev pvaihadqlt ptwrvystgs '3 market pneumonia - nvfqtragcl igaehvnnsy 661 eedipigagi casyqtqlns virus]; GenBank: prrarsvasq siiaytmslg aensvaysnn siaiptnfti 721 QHD43416 .1 ; svt-teilpvs mtktsvdctm yicgdstecs nlllqygsfc tqlnraltgi January 23, 2020 aveqdkntqe 781 vfaqvkqiyk tppikdfggf nfsqilpdps kpskrsfied llfnkvtlad agfikqygdc 841 lgdiaardli caqkfngltv 1pplltdemi aqytsallag titsgw-tfga gaalqipfam 901 qmayrfngig vtqnvlyenq klianqfnsa igkiqdslss tasalgklqd vvnqnaqaln 961 tivkqlssnf gaissvindi lsrldkveae vqidrlitgr lqslqtyvtq qliraaeira 1021 sanlaatkms ecvlgqskry dfcgkgyhlm sfpqsaphgv vflhvtyvpa qeknfttapa 1081 ichdgkahfp regvfvsngt hwfvtqmfy epqiittdnt fvsgncdvvi givnntvydp 1141 lqpeldsfice eldkyficnht spdvdlgdis ginasvvniq keidfineva knlneslidl 1201 qelgkyeqyi kwpwyiwlgf iagliaivmv timlccmtsc csclkgccsc gscckfdedd 1261 sepvlkgvkl hyt SEQ
Sequence ID Sequence Description NO.
surface glycoprotein RBD
nitni cpfgevfn atrfasvyawn rkri sncvadysvlynsasfstfkcygvs [SARS-CoV-2 Wuhan seafood ptklndleftnvyadsfvirgdevrqiapgqtgkiadynyklpddftgcvia wnsnnldskvggnynylyrlfrksnlkpferdisteiyqagstpcngvegfn market pneumonia cyfpl qsygfqptngvgyqpyrvvvl sfellh apatvcgpkk stnlvknkc virus]; GenBank:
vnfnfngltgtg QHD43416.1;
January 23, 2020 Receptor Binding Motif (RBM) in surface glycoprotein RBD
[SARS-CoV-2 Nsnnldskvggnynylyrlfrksnlkpferdisteiyqag stpcngvegfnc Wuhan seafood yfplqsygfqgtngvgyqpy market pneumonia virus]; GenBank:
QHD43416.1;
January 23, 2020 ASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPE
PVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVV
TVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSC
DKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRT
SARS-CoV-2 PEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKT

Glm17 IgHG1*01 6VSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDEL
(aa) TKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYK
TTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCS
VLHEALHSHYTQKSLSLSPGK
ASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPE
PVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVV
TVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSC
DKTHTCPPCPAPELLAGPSVFLFPPKPKDTLMISRT
SARS-CoV-2 PEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKT
CH1-CH3 LS, 7 KPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCK
ALE Glm17 VSNKALPLPEEKTISKAKGQPREPQVYTLPPSRDE
IgHG1*01 (aa) LTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNY
KTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSC
SVLHEALHSHYTQKSLSLSPGK

SEQ
Sequence ID Sequence Description NO.
GQPKAAPSVTLFPPS SEELQANKATLVCLISDFYP
GAVTVAWKADSSPVKAGVETTTPSKQSNNKYAA
SARS-CoV-2 CL SSYLSLTPEQWKSHRSYSCQVTHEGSTVEKTVAP
IgLC*01 (aa) 8 TECS
RTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPR
EAKVQWKVDNALQSGNSQESVTEQDSKD STYSL
SARS-CoV-2 CL SSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSF
(CK) klm3 9 NRGEC
IgKC*01 (aa) Linker (aa) 10 GSTSGSGKPGSGEGSTKG
Linker (aa) 11 GSGKPGSGEG
Linker (aa) 12 GKPGSGEG
Linker (aa) 13 SGKPGSGE
Linker (aa) 14 BPXXXZ, wherein each X is independently a glycine (G) or serine (S). B is a positively charged amino acid and Z is glycine (G) or a negatively charged amino acid Linker (aa) 15 (GxS)y, wherein xis 1-10 and y is 1-Linker (aa) 16 GGGGSGGGGSGGGGS
Linker (aa) 17 GGGGSGGGGSGGGGSGGGGSGGGGS
GGGGSGGGGSGGGGSGGGGSGGGGS
Linker (aa) 18 GSTSGGGSGGGSGGGGSS
Linker (aa) 19 EGKS SG SG SESKVD
Linker (aa) 20 KESGSVSSEQLAQFRSLD
Linker (aa) 21 GGGGS
EVQLVESGGGVVQPGRSLRLSCAASGFTFSTYAM
Antibody 418 1 HWVRQAPGKGLEWVAVILSDGSNKYYADSVKG

VH (aa) RFTISRDNSKNTLYLQMNSLRAEDTAVYYCARDR
SPLVGFGDNYGMDVWGQGTTVTVSS
Antibody 418 1 23 GFTFSTYA
CDRH1 (aa) Antibody 418_i 24 ILSDGSNK
CDRH2 (aa) SEQ
Sequence ID Sequence Description NO.
Antibody 418 1 25 ARDRSPLVGFGDNYGMDV
CDRH3 (aa) SYELTQPPSVSVSPGQTARITCSGDALPKKYAYW
Antibody 418 1 YQQKSGQAPVLVIYEDSKRPSGIPERFSGSSSGTM
VL (aa) 26 ATLTISGAQVEDEADYYCSSTDSSGNQGVFGGGT
KLTVL
Antibody 418_i 27 ALPKKY
CDRL1 (aa) Antibody 418 1 CDRL2 (aa) Antibody 418 1 29 SSTDSSGNQGV
CDRL3 (aa) GAGGTGCAGCTGGTGGAGTCTGGGGGAGGCGT
GGTCCAGCCTGGGAGGTCCCTGAGACTCTCCTG
TGCAGCCTCTGGATTCACCTTCAGTACCTATG
CTATGCACTGGGTCCGCCAGGCTCCAGGCAAG
GGGCTAGAGTGGGTGGCAGTTATATTATCTGA
Antibody 418 1 TGGAAGTAATAAATATTACGCAGACTCTGTGA
30 H (nt AGGGC CGATTCAC CATC TC CAGAGACAATTC CA

) V
AGAACACGCTGTATCTGCAAATGAACAGCCTGA
GAGCTGAGGACACGGCTGTGTATTACTGTGCG
AGAGATCGAAGTCCCCTCGTGCCATTCGCGC
ACAACTATGGTATGGACGTCTGGGGCCAAGG
GACCACGGTCACCGTCTCCTCA
TCCTATGAGCTGACACAGCCACCCTCGGTGTCA
GTGTCCCCAGGACAAACGGCCAGGATCACCTGC
TCTGGAGATGCATTGCCAAAAAAATATGCTTA
TTGGTACCAGCAGAAGTCAGGCCAGGCCCCTGT
Antibody 418 1 GCTGGTCATCTATGAGGACAGCAAACGACCCT
VL (nt) 31 CCGGGATC CC TGAGAGATTC TCTGGC TCCAGC T
CAGGGACAATGGCCACCTTGACTATCAGTGGGG
CCCAGGTGGAGGATGAAGCTGACTACTACTGTT
CCTCAACAGACACCACTGGTAATCAAGGGGT
ATTCGGCGGAGGGACCAAGCTGACCGTCCTAG
QITLKESGPTLVKPTQTLTLTCKLSGFSVNTGGVG
Antibody 418 2 VGWIRQPPGKALEWLALIYWNDDKLYSP SLKSRL
VH (aa) 32 TVTKDTSKNQVVLTMTNMDPVDTATYYCAHVL
VWFGEVLPDAFDVWGQGTMVTVSS
Antibody 418_2 33 GFSVNTGGVG
CDRH1 (aa) SEQ
Sequence ID Sequence Description NO.
Antibody 418 2 34 IYWNDDK
CDRH2 (aa) Antibody 418 2 CDRH3 (aa) SYELTQPPSVSVSPGQTASITCSGDKLGETYASW
Antibody 418 2 YQQKPGQSPILVIYQDNKRPSGIPERFSGSNSENTA
VL (aa) 36TLTISGTQTMDEADYYCQAWDKTIAGFGGGTKL
TVL
Antibody 418_2 37 KLGETY
CDRL1 (aa) Antibody 418 2 38 QDN
CDRL2 (aa) Antibody 418_2 39 QAWDKTIAG
CDRL2 (aa) CAGATCACCTTGAAGGAGTCTGGTCCTACGCTG
GTGAAACCCACACAGACCCTCACGCTGACCTGC
AAATTATCTGGGTTTTCAGTCAACACTCGTGC
AGTGGGTGTGGGCTGGATCCGTCAGCCCCCAG
GAAAGGCCCTGGAGTGGCTTGCACTCATTTATT
Antibody 418 2 GGAATGATGATAAGTTGTACAGCCCATCTCTG
VH (nt) 40AAGAGCAGGCTCACCGTCACCAAGGACACATC
CAAAAACCAGGTGGTCCTTACAATGACCAACAT
GGACCCTGTGGACACAGCCACATATTACTGTGC
ACACCTATTACTTTCGTTCGCCGAGGTATTA
CCCGATGCTTTTGATGTGTGGGGCCAAGGGAC
AATGGTCACCGTCTCTTCAG
TCCTATGAGCTGACTCAGCCACCCTCAGTGTCC
GTGTCCCCAGGACAGACAGCCAGCATCACCTGC
TCTGGAGATAAATTGGGGGAGACATATGCTAG
TTGGTATCAGCAGAAGCCAGGCCAGTCCCCTAT
Antibody 418 2 TCTAGTCATCTATCAAGATAACAAGCGGCCCTC
VL (nt) 41 AGGGATCCCTGAGCGATTCTCTGGCTCCAACTC
TGAGAACACAGCCACTCTGACCATCAGCGGGA
CCCAGACTATGGATGAGGCTGACTATTACTGTC
AGGCGTCCGACAAGACCATCGCCGCATTCGG
CGGAGGGACCAAGCTGACCGTCCTAG
EVQLVESGGGVVQPGRSLRLSCAASGFIFSTYGM
Antibody 418 3 HWVRQAPGKGLEWVAlIWYDGTKKYYADSVKG
VH (aa) 42RFTISRDNSKNTLYLQMNILRAEDTAVYYCASNR
YHYASSGYYQLDYWGQGTLVTVSS

SEQ
Sequence ID Sequence Description NO.
Antibody 418 3 CDRHI (aa) Antibody 418 3 CDRH2 (aa) Antibody 418 3 CDRH3 (aa) DIQMTQSPSSLSASVGDRVTITCQASQDISNSLNW
Antibody 418 3 YQQKPGKAPNLLIYDASNLETGVPSRFSGSGSGTD
VL (aa) 46 FTFTISSLQPEDVATYYCQHYDHLPLTFGGGTKV
EIK
Antibody 418 3 47 QDISNS
CDRLI (aa) Antibody 418 3 CDRL2 (aa) Antibody 418_3 CDRL3 (aa) GAGGTGCAGCTGGTGGAGTCGGGGGGAGGCGT
GGTCCAGCCTGGGAGGTCCCTGAGACTCTCCTG
TGCAGCGTCTGGATTCATCTTCAGTACCTATG
GCATGCACTGGGTCCGCCAGGCTCCAGGCAAG
GGGCTGGAGTGGGTGGCTATTATATGGTATGA
Antibody 418 3 TGGAACTAAAAAATACTATGCAGACTCCGTGA
VH (nt) 5 AGGGCCGATTCACCATCTCCAGAGACAATTCCA
AGAACACGCTGTATCTACAAATGAACATCCTGA
GAGCCGAGGACACGGCTGTGTATTACTGTGCG
AGTAACCGGTA ICACTATGCTAGTAGTGUI
ATTATCAACTTGACTACTGGGGCCAGGGAACC
CTGGTCACCGTCTCCTCAG
GACATCCAGATGACCCAGTCTCCATCCTCCCTG
TCTGCATCTGTTGGAGACAGAGTCACCATCACT
TGCCAGGCGAGTCAGGACATTAGCAACTCTTT
AAATTGGTATCAGCAGAAACCAGGGAAAGCCC
Antibody 418 3 CTAACCTCCTGATCTACGATGCATCCAATTTGG
VL (nt) 51 AAACAGGGGTCCCATCAAGGTTCAGTGGAAGT
GGATCTGGGACAGATTTTACTTTCACCATCAGC
AGCCTGCAGCCTGAAGATGTTGCAACATATTAC
TGTCAACATTATGATCATCTCCCTCTCACTTT
CGGCGGAGGGACCAAGGTGGAGATCAAAC
EVQLVESGGGVVQPGRSLRLSCAASGFTFSNYG
Antibody 418_4 52 MI-IWVRQAPGKGLEWVAVIWYDGSNKFYADSV

SEQ
Sequence ID Sequence Description NO.
VH (aa) KGRFTISRDNSKNSLYLQMNSLRAEDTAVYFCAR
AFPDSSSWSGFTIDYWGQGTLVTVSS
Antibody 418_4 53 GFTFSNYG
CDRH1 (aa) Antibody 418 4 54 IWYDGSNK
CDRH2 (aa) Antibody 418 4 55 ARAFPDSSSWSGF TIDY
CDRH3 (aa) SYELTQPPSVSVAPGQTARITCGGNNIERKSVHW
Antibody 418_4 CQQKPGQAPALVVYDDSDRPSGIPERFSGSNSGNT
VL (aa) 56 ATLTISRVEAGDEADYYCQVWDSGSDQVIFGGG
TKLTVL
Antibody 418 4 57 NIERKS
CDRL 1 (aa) Antibody 418_4 CDRL2 (aa) Antibody 418_4 59 QVWDSGSDQVI
CDRL3 (aa) GAGGTGCAGCTGGTGGAGTCTGGGGGAGGCGT
GGTCCAGCCTGGGAGGTCCCTGAGACTCTCCTG
TGCAGCGTCTGGATTCACCTTCAGTAATTATG
GCATGCACTGGGTCCGCCAGGCTCCAGGCAAG
GGACTGGAGTGGGTGGCAGTTATATGGTATGA
Antibody 418 4 TGGAAGTAATAAATTCTATGCAGACTCCGTGA
VH (nt) ¨ 60AGGGCCGATTCACCATCTCCAGAGACAATTCCA
AGAACAGTCTCTATCTGCAAATGAACAGCCTGA
GAGCCGAGGACACGGCTGTTTATTTCTGTGCGA
GGGCCTTTCCCGATAGCAGCAGCTGGTCCGG
CTTCACTATTGACTACTGGGGCCAGGGAACCC
TGGTCACCGTCTCCTCAG
TCCTATGAGCTGACTCAGCCACCCTCGGTGTCA
GTGGCCCCAGGACAGACGGCCAGGATTACCTGT
GGGGGAAACAACATTGAGAGGAAAAGTGTGC
ACTGGTGCCAGCAGAAGCCAGGCCAGGCCCCT
GCGCTGGTCGTCTATGATGATAGCGACCGGCC
Antibody 418 4 VL (nt) CTCTGGGAACACGGCCACCCTGACCATCAGCAG
GGTCGAAGCCGGGGATGAGGCCGACTATTACT
GTCAGGTGTGGGATAGTGGTAGTGATCAGGT
GATATTCGGCGGAGGGACCAAGCTGACCGTCC
TAG

SEQ
Sequence ID Sequence Description NO.
EVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGM
HWVRQAPGKGLEWVTVIWYDGSNRYYADSVKG
VU Antibody Anti4a)18-5 AGEWYFDYWGQGTLVTVSS
Antibody 418_5 63 GFTFSSYC
CDRH1 (aa) Antibody 418_5 64 IWYDGSNR
CDRH2 (aa) Antibody 418 5 65 ARAVAGEWYFDY
CDRH3 (aa) SYELTQPPSVSVSPGQTARITCSGDALAKHYAYW
YRQKPGQAPVLVIYKDSERPSGIPERFSGSSSGTTV
Antibody 418 5 TLTISGVQAEDEADYYCQSADSIGSSWVFGGGTK
VL (aa) 66LTVL
Antibody 418 5 67 ALAKHY
CDRL1 (aa) Antibody 418 5 68 KDS
CDRL2 (aa) Antibody 418_S 69 QSADSIGSSWV
CDRL3 (aa) GAGGTGCAGCTGGTGGAGTCTGGGGGAGGCGT
GGTCCAGCCTGGGAGGTCCCTGAGACTCTCCTG
TGCAGCGTCTGGATTCACCTTCAGTAGCTATG
GCATGCACTGGGTCCGCCAGGCTCCAGGCAAG
GGGCTGGAGTGGGTGACAGTTATTTGGTATGA
TGGAAGTAATCGATACTATGCAGACTCCGTGA
Antibody 418-5 VH (nt 70 AGGGCCGATTCACCATCTCCAGAGACAATTCCA
) AGAACACGCTGTATCTGCAAATGGACAGCCTGA
GAGCCGAGGACACGGCTGTTTATTACTGTGCGA
GAGCAGTGGCCGGGGAATGGTACTTTGACTA
CTGGGGCCAGGGAACCCTGGTCACCGTCTCCTC
AG
TCCTATGAGCTGACACAGCCACCCTCGGTGTCA
GTGTCCCCAGGACAGACGGCCAGGATCACCTGC
TCCGGAGATGCATTGGCAAAACACTATGCTTA
Antibody 418 5 VL (nt) GCTGGTGATATATAAAGACAGTGAGAGGCCCT
CAGGGATCCCTGAGCGATTCTCTGGCTCCAGCT
CAGGGACAACAGTCACGTTGACCATCAGTGGA

SEQ
Sequence ID Sequence Description NO.
GTCCAGGCAGAAGACGAGGCTGACTATTACTGT
CAATCAGCAGACAGCATTGGTAGTTCTTGGG
TGTTCGGCGGAGGGACCAAGCTGACCGTCCTA
QVQLQESGPGLVKPSETLSLTCTVSGGSVNSGSY
Antibody 418 6 YWSWIRQPPGKGLEWIGYIFYS GS TYYNP SLKSR
¨ 72 VTISIDTSKNQF SLKLS SVTAADTAVYYCAREVAP
VH (aa) VAGTAHQTTYYFDYWGQGTLVTVSS
Antibody 418_6 73 GGS VN SGS Y Y
CDRH1 (aa) Antibody 418_6 74 IFYSGST
CDRH2 (aa) Antibody 418 6 75 AREVAPVAGTAHQTTYYFDY
CDRH3 (aa) DIVMTQSPSSLSVSVGDRVSITCRASQSISTYLNW
Antibody 418 6 YQQKPGKAPKLLIYAASSLHSGVPSRFSGSGSGTD
VL ( ¨ 76 FTLTISSLQPEDFATYYCQQSRPLEEGICRYTFGQ
aa) GTKLEIK
Antibody 418 6 CDRL1 (aa) Antibody 418 6 78 AAS
CDRL2 (aa) Antibody 418 6 QQSRPLEEGICRYT

CDRL3 (aa) CAGGTGCAGCTACAGGAGTCGGGCCCAGGATT
GGTGAAGCCTTCGGAGACCCTGTCCCTCACCTG
CACTGTCTCTGGTGGCTCCGTCAACAGTGGCA
GTTACTACTGGAGCTGGATCCGGCAGCCCCCA
GGGAAGGGACTGGAGTGGATTGGGTATATCTT
TTACAGTGGGAGCACCTACTACAACCCCTCCC
Antibody 418-6 80 TCAAGAGTCGAGTCACCATATCAATAGACACGT
VH (nt) CCAAGAACCAGTTCTCCCTGAAGCTGAGCTCTG
TGACCGCTGCGGACACAGCCGTGTATTACTGTG
CGAGAGAGGTTGCGCCAGTGGCTGGTACTG
CCCACCAAACAACGTACTACTTTGACTACTG
GGGCCAGGGAACCCTGGTCACCGTCTCCTCAG
GACATCGTGATGACCCAGTCTCCATCCTCCCTG
Antibody 418 6 VL (nt) TGCCGGGCAAGTCAGAGCATTAGCACCTATTT

SEQ
Sequence ID Sequence Description NO.
AAATTGGTATCAGCAGAAACCAGGGAAAGCCC
CTAAGCTCCTGATCTATGCTGCATCCAGTTTGC
ACAGTGGGGTCCCATCAAGGTTCAGTGGCAGTG
GATCTGGGACAGATTTCACTCTCACCATCAGCA
GTCTGCAACCTGAAGATTTTGCAACTTACTACT
GTCAACAGAGTCGGCCGCTCGAAGAAGGCAA
AAGGTACACTTTTGGCCAGGGGACCAAGCTGG
AGATCAAAC
EVQLVQ SGAEVNKPGS SVKVSCKASGGTFSSYAI
Antibody 418 7 SWVRQAPGQGLEWMGGIIPIFHTANYAQKFHGR
VH ( ¨ 82 VTITADESTSTAYMELN SLRSEDTAVY Y CAGD SG
aa) SSTWLGPFDIWGQGTMVTVS S
Antibody 418 7 83 GGTFSSYA
CDRHI (aa) Antibody 418 7 84 IIPIFHTA
CDRH2 (aa) Antibody 418_7 85 AGDSGSSTWLGPFDI
CDRH3 (aa) VIWMTQ S P S TL SA SVGDRVTITC RA S QGISSYLAW
Antibody 418 7 YQQKPGKAPKLLIYDASTLQ SGVP SRFSGSGSGTE
¨ VL ( 86 FTLTI S SLQPGDF A TYYC QQYNSYP YTFGQGTKLE
aa) IK
Antibody 418 7 CDRL 1 (aa) Antibody 418 7 88 DAS
CDRL2 (aa) Antibody 418 7 89 QQYNSYPYT
CDRL3 (aa) GAGGTGCAGCTGGTGCAGTCTGGGGCTGAGGT
GAACAAGCCTGGGTCCTCGGTGAAGGTCTCCTG
CAAGGCTTCTGGAGGCACCTTCAGCAGCTAT
GCCATCAGCTGGGTGCGACAGGCCCCTGGACA
Antibody 418 7 90 AGGGCTTGAGTGGATGGGAGGGAT CATCCC TA
VH (nt) TCTTTCATACAGCAAACTACGCACAGAAGTTC
CACGGCAGAGTCACGATTACCGCGGACGAATC
CACGAGCACAGCCTACATGGAGCTGAACAGCC
TGAGATCTGAGGACACGGCCGTGTATTACTGTG
CGGGGGATAGTGGGAGCTCAACCTGGCTCG

SEQ
Sequence ID Sequence Description NO.
GACCTTTTGATATCTGGGGCCAAGGGACAATG
GTCACCGTCTCTTCAG
GTCATCTGGATGACCCAGTCTCCTTCCACCCTGT
CTGCATCTGTAGGAGACAGAGTCACCATCACTT
GCCGGGCCAGTCAGGGCATTAGCAGTTATTTA
GCCTGGTATCAGCAAAAACCAGGGAAAGCCCC
Antibody 418 7 TAAGCTCCTGATCTATGATGCATCCACTTTGCA
VL (nt 91 AAGTGGGGTCCCATCAAGGTTCAGCGGCAGTG
) GATCTGGGACAGAATTCACTCTCACCATCAGCA
GCCTGCAGCCTGGTGATTTTGCAACTTATTACT
GCCAACAGTATAATAGTTACCCGTACACTTTT
GGCCAGGGGACCAAGCTGGAGATCAAAC
QVQLVQSGAEVKKPGA SVKVSCKVSGYTLIEIS
Antibody 418 8 MHWVRQAPGKGLEWNIGGFDPEDAETIYAQKFQ
9") VH (aa) GRVTMTEDTSTDTAYMELSSLRSEDTAVYYCAT
QYAILTHSYFDYWGQGTLYTVSS
Antibody 418 8 CDRH1 (aa) Antibody 418_8 CDRH2 (aa) Antibody 418 8 CDRH3 (aa) DIQLTQSPSSLSASVGDRVITIVRASQGISNYLAW
Antibody 418 8 YQQKPGKVPKLLIYAASTLQSGVPSRFSGSGSGTD
VL (aa) 96 FTLTISSIAREDVATYYCQKYNSAPQTFGQGTKV
EIK
Antibody 418_8 CDRL1 (aa) Antibody 418 8 CDRL2 (aa) Antibody 418_8 CDRL3 (aa) CAGGTGCAGCTGGTACAGTCTGGGGCTGAGGICT
A A G.A A GC CTGGGGCCTCA. MU A A GCiTCTCCTGC
AAGGTTTCCG GA TACACC CTCATTGAA TTA TC
CATGCA CTGGGTGCGAC AGG CICCIGGAAAA G
Antibody 418_8 VH (nt) GATGCTGAAACAATCTACG CAC AGAACTIC CA
GGGCACiAGICACCATGACCGAGGACACATCTA
CAGACACAGCCTACATGGAGCTGAGCAG CCM
AG A TCTGAGGACACGG CCGTGTATTA CTGTG CA

SEQ
Sequence ID Sequence Description NO.
ACACAA TA CG CAA TCCTTACTCATTCCTACTT
Tc ACTACTGGGGCCAGGGAACCCTGGTCACCG
TCH:CTCAG
GACATCCAGTTGACCCAGTCTCCATCCTCCCTG
TCTGCATCTGTAGGAGACAGAGTcACCATCACT
TGCCGGGCGAGTCAGGGCATTAGCAATTATIFT
AGCCTGG'FATCAGCAGAAACCAGGGAAAGTTC
Antibody 418 8 CTAAGCTCCTGATCTATCCTGCATCCACTTTGC
VL (nt) 101AATCAGGGGTCCCATCTCGGTFCAGCGGCAGTG
GATCTGGGACAGATITCACTCTCACCATCAGCA
GCCTGCAGCCTGAAGATGTTG CAA CTTATTA CT
GTCAAAAGTATAACAGTGCCCCTCAGACGTT
CGGCCAAGGGACCAAGGTGGAAATCAAAC
EVQLVESGGGILVNPGGSLRLSCAASGFTFSDYTI
Antibody 418 9 VH (aa) 102 ,VISRDNAKNSLSWAINSLRAEDTAVYYCARDGN
AYKWLLAENVIUDYINGQGILVIVSS
Antibody 418 9 CDRH1 (aa) Antibody 418 9 CDRH2 (aa) Antibody 418 9 CDRH3 (aa) QTVVTQPASVSGSPGQSITISCTGTSSDVGGVNYV
Antibody 418 9 VL (aa) GGGTKILTV1, Antibody 418 9 CDRL1 (aa) Antibody 418_9 CDRL2 (aa) Antibody 418 9 CDRL3 (aa) GAGGTGCAGCTGGTGGAGTCTGGGGGAGGCCT
GGICAACCCTGGGGGGIFCCCTGAGACTCTCCTG
TGCAGCCTCTGGATTCACCTTCAGTGACTATA
CC.:ATFCACTGGG-FCCGCCACiGcrccAGGGAAG
Antibody 418 9 110 GGGCTGGACi.MGGICICATCCATIAGTAGIAG
VH (nt) TAGTAATTATATATACTACGCGGACTCAGTGA
AGGGCCGATTCACCATCTCC.AGAGACAA.COCCA
AGAA.CTCACIGTCTCMCAAATGAACA.GCCTGA
GAGCCGA GG A C A COGCT(IFCiTA "'TA Cf(iTGCG

SEQ
Sequence ID Sequence Description NO.
AGA GATGGTAATG CCTACAAGTGG TTA TTGG
GAGA G AACGTTCG TTIFTGACTACTGGGGCCA
GGGAACCCTGGTCACCGTCTCCTCAG
C AGA CTGTG GTGA CTCAGCCTGCCTC CGTGTCT
GGGTCTCCTGGACAGTCCiATC AC CATCTCCTGC
ACTGGAAC CAGCA GT GACGT T GGTGGTTATA
AC TAT GTCTC CIGGTAC CAACAACAC C CAGGCA

Antibody 418 9 VL (nt) GGCTCCAAGTCTGGCAACACGGCCTCCCTGACC
ATCTCTGGGCTC CAGGCTGAGGAC GAGGCTGAT
TATTACTG CA GCTC ATATACAA GCAGCAGCA C
CCCCAATTGGG TGTTCGGCGGAGGGACCAAGC
TGACCGTCCTAG
QVQLQQRGAGLLKPSETLSLTCDVYGGSLSGYY
Antibody 418 10 WSWIRQAPGKGLEWIGEINHRGSTNYNPSLKSRV
VH (aa) 112 TISIDTSKKQFSLKLSSVTAADTAVYYCARYVVVI
VHALPMPVNWFDPWGQGTLVTVSS
Antibody 418 10 CDRH1 (aa) Antibody 418 10 CDRH2 (aa) 114 INIIRGST
Antibody 418 10 CDRH3 (aa) QSVLTQPASVSGSPGQSITISCTGTSSDVGSYNLVS
Antibody 418 10 WYQQHPAKAPKLIWEGSKRPSGVSNRFSGSKSG
vL'(aa) ¨ 116 NTASLTISGLQAEDECDYYCCSYAGSSPLIVFGTG
TKVTVL
Antibody 418 10 CDRL1 (aa) Antibody 418 10 CDRL2 (aa) Antibody 418 10 CDRL3 (aa) CAGGTGCAGCTACAGCAGCGGGGCGCAGGACT
GTTGAAGCCTTCGGAGACCCTGTCCCTCACCTG
CGATGTCTATGGTGGGTCCCTCAGTGGTTACT
Antibody 418 10 VH (nt) GGGCTGGAGTGGATTGGGGAAATCAATCATCG
TGGAAGCACCAACTACAACCCGTCCCTCAAGA
GTCGGGTCACCATATCAATAGACACGTCCAAGA

SEQ
Sequence ID Sequence Description NO.
AGCAGTTCTCCCTGAAGCTGAGCTCTGTGACCG
CCGCGGACACGGCTGTGTATTACTGTGCGAGA
TACGTTGTGGTGATCGTACATGCCCTTCCAA
TGCCAGTTAACTGGTTCGACCCCTGGGGCCA
GGGAACCCTGGTCACCGTCTCCTCAG
CAGTCTGTGCTGACTCAGCCTGCCTCCGTGTCT
GGGTCTCCTGGACAGTCGATCACCATCTCCTGC
ACTGGAACCAGCAGTGATGTTGGGAGTTATA
ACCTTGTCTCCTGGTACCAACAACACCCAGCCA
AAGCCCCCAAACTCATCATTTATGAGGGCAGT
Antibody 418 10 121 AAGCGGCCCTCAGGAGTTICTAATCGCTICTCT
VL (nt) GGCTCCAAGTCTGGCAACACGGCCTCCCTGACA
ATCTCTGGACTCCAGGCTGAGGACGAATGTGAT
TATTACTGCTGCTCATATGCAGGTAGTAGCCC
CTTGATAGTCTTCGGAACTGGGACCAAGGTCA
CCGTCCTAG
EVQLLESGGGLIQPGGSLRLSCAASGFSVSSNYM
Antibody 418 11 NWVRQAPGKGLEWVSVIYSGGSAYYADSVKGRF
VH (aa) 122 TISRDISKNTLYLQMNSLRAEDTAVYYCARAPGS
WAYWYFDLWGRGTLVTVSS
Antibody 418 11 GFSVSSNY

CDRH1 (aa) Antibody 418 11 124 IYSGGSA
CDRH2 (aa) Antibody 418 11 125 ARAPGSWAYWYFDL
CDRH3 (aa) EIVMMQSPATLSVSPGERATLSCRASQSVRSNLA
A WYQQKPGQAPRLLIYGASTRATGIPARFSGSGSGT

Antibody 418

11 EIK
Antibody 418 11 127 QSVRSN
CDRL1 (aa) Antibody 418 11 GAS

CDRL2 (aa) Antibody 418 11 1,9 QQYNIWPT
CDRL3 (aa) GAGGTGCAGCTGTTGGAGTCTGGAGGAGGCTTG
Antibody 418 11 ATCCAGCCGGGGGGGTCCCTGAGACTCTCCTGT
VH (nt) 130GCAGCCTCTGGGTTCAGCGTCAGTAGCAACT
ACATGAACTGGGTCCGCCAGGCTCCAGGGAAG

SEQ
Sequence ID Sequence Description NO.
GGGCTGGAGTGGGTCTCAGTTATTTATAGCGG
TGGTAGTGCATACTACGCAGACTCCGTGAAGG
GCCGATTCACCATCTCCAGAGACATTTCCAAGA
ACACGCTGTATCTTCAAATGAACAGCCTGAGAG
CCGAGGACACGGCCGTGTATTACTGTGCGAGA
GCCCCCGGCAGTTGGGCCTACTGGTACTTCG
ATCTCTGGGGCCGTGGAACCCTGGTCACTGTCT
CCTCAG
GAAATAGTGATGATGCAGTCTCCAGCCACCCTG
TCTGTGTCTCCAGGGGAAAGAGCCACCCTCTCC
TGCAGGGCCAGTCAGAGTGTTAGAAGCAACTT
AGCCTGGTACCAGCAGAAACCTGGCCAGGCTCC
Antibody 418 11 CAGGCTCCTCATCTATGGTGCATCCACCAGGG
VL(VK) (nt) , , CCACTGGTATCCCAGCCAGGTTCAGTGGCAGTG
GGTCTGGGACAGAGITCACTCTCACCATCAGCA
GCATGCAGTCTGAAGATTTTGCAGTTTATTACT
GTCAGCAGTATAATATCTGGCCGACGTTCGGC
CAAGGGACCAAGGTGGAAATCAAAC
EVQLLESGGGLVQPGGSLRLSCAASGFTFSSHAM
Antibody 418 12 SWVRQAPGKGLEWVSVISGSGGDTYYADSVKGR
¨ 132 FTTSRDNSKNTLYLQ1VINSLRAEDTAVYYCAKGE
VH (aa) RIKMIVVVTMIDYWGQGTLVTVSS
Antibody 418_12 133 GFTFSSHA
CDRH1 (aa) Antibody 418_12 134 ISGSGGDT
CDRH2 (aa) Antibody 418 12 135 AKGERIKMIVVVTMIDY
CDRH3 (aa) QSVVTQPPSVSAAPGQKVTISCSGSSSNIGSNYVS
418 12 Antibody WYQQLPGTAPKLLIYDNNKRPSGIPDRFSGSKSGT
vL'(aa) ¨ 136 SATLGITGLQTGDEADYYCGTWDNSLSAGVFGG
GTKLTVL
Antibody 418_12 137 SSNIGSNY
CDRL1 (aa) Antibody 418 12 138 DNN
CDRL2 (aa) Antibody 418 12 139 GTWDNSLSAGV
CDRL3 (aa) SEQ
Sequence ID Sequence Description NO.
GAGGTGCAGCTGTTGGAGTCTGGGGGAGGCTTG
GTACAGCCTGGGGGGTCCCTGAGACTCTCCTGT
GCAGCCTCTGGATTCACCTTTAGCAGTCATGC
CATGAGCTGGGTCCGCCAGGCTCCAGGGAAGG
GGCTGGAGTGGGTCTCAGTTATTAGTGGTAGT
Antibody 418 12 GGTGGTGACACATACTACGCAGACTCCGTGAA
vH (nt) ¨ 140 GGGCCGGTTCACCATCTCCAGAGACAATTCCAA
GAACACGCTGTATTTGCAAATGAACAGCCTGAG
AGC CGAGGACACGGC CGTATATTACTGTGC GA
AAGGCGAACGTATTAAAATGATAGTAGTCGT
TACTATGATTGACTACTGGGGCCAGGGAACCC
TGGTCACCGTCTCCTCAG
CAGTCTGTCGTGACGCAGCCGCCCTCAGTGTCT
GCGGCCCCAGGACAGAAGGTCACCATCTCCTGC
TCTGGAAGCAGCTCCAACATTGGGAGTAATTA
TGTATCCTGGTACCAGCAGCTCCCAGGAACAGC
CC CCAAACTCCTCATTTATGACAATAATAAGCG
Antibody 418 12 141 ACCCTCAGGGATTCCTGACCGATTCTCTGGCTC
VL (nt) CAAGTCTGGCACGTCAGC CAC CCTGGGCATCA C
CGGACTCCAGACTGGGGACGAGGCCGATTATTA
CTGCGGAACATGGGATAACAGCCTGAGTGCT
GGGGTATTCGGCGGAGGGACCAAGCTGACCGT
CCTAG
QVTLRESGPALVKPTQTLTLTCTFSGFSLSTRGM
Antibody 418_13 CVNWIRQPPGKALEWLAFIDWDDDICYYSTSLKT

(aa) Antibody 418 13 143 GFSLSTRGMC
CDRH1 (aa) Antibody 418_13 144 IDWDDDK
CDRH2 (aa) Antibody 418_13 145 ARIRGVIPAAGTVPYYHYMDV
CDRH3 (aa) DIVMTQSPLSLPVTPGEPASISCRS SQSLL HSNGY
Antibody 418 13 NYLDWYLQKPGQ SPQLLIYLGSNRASGVPDRF SG
¨ VL(VK) (aa) 146 SGSGTDFTLKISRVEAEDVGVYYCMQAL QTLSIT
FGQGTRLEIK
Antibody 418 13 147 QSLLHSNGYNY
CDRL 1 (aa) SEQ
Sequence ID Sequence Description NO.
Antibody 418 13 LGS

CDRL2 (aa) Antibody 418 13 149 MQALQTLSIT
CDRL3 (aa) CAGGTCACCTTGAGGGAGTCTGGTCCTGCGTTG
GTGAAACCCACACAGACCCTCACACTGACCTGC
ACCTTCTCTGGGTTCTCACTCAGCACTCGTGG
AATGTGTGTGAACTGGATCCGTCAGCCCCCAG
GGAAGGCCCTGGAGTGGCTTGCATTCATTGATT
Antibody 418 13 GGGATGATGATAAATACTACAGCACATCTCTG
VH (nt) ¨ 150 AAGACCAGGCTCACCATCTCCAAGGACACCTCC
AAAAACCAGGTGGTCCTTACAATGACCAACATG
GACCCTGTGGACACAGCCACGTATTACTGTGCA
CGGATACGGGGGGTTATACCAGCAGCTGGTA
CAGTTCCCTACTACCACTACATGGACGTCTG
GGGC A A AGGGACCACGGTCACCGTCTCCTCA
GATATTGTGATGACTCAGTCTCCACTCTCCCTGC
CCGTCACCCCTGGAGAGCCGGCCTCCATCTCCT
GCAGGTCTAGTCAGAGCCTCCTGCATAGTAAT
CCATACAACTATTTGGATTGGTACCTGCAGAA
GCCAGGGCAGTCTCCACAGCTCCTGATCTATTT
Antibody 418 13 151 GGGTTCTAATCGGGCCTCCGGGGTCCCTGACA
VL(VK) (nt) GGTTCAGTGGCAGTGGATCAGGCACAGATTTTA
CACTGAAAATCAGCAGAGTGGAGGCTGAGGAT
GTTGGGGTTTATTACTGCATGCAAGCTCTACA
AACTCTTTCCATCACCITCGGCCAAGGGACAC
GACTGGAGATTAAAC
EVQLVESGGGLVKPGGSLRLSCAASGFTFSTYSM
Antibody 418 14 NWVRQAPGKGLEWVSSISSSSSYIYYADSVKGRF
( ¨ 152 TISRDNAKNSLFLQMNSLRAEDTAVYYCARWGY
aa VH ) SYDSRGYYPRELDYWGQGTLVTVSS
Antibody 418_14 153 GFTFSTYS
CDRH1 (aa) Antibody 418 14 154 ISSSSSYI
CDRH2 (aa) Antibody 418 14 155 ARWGYSYDSRGYYPRELDY
CDRH3 (aa) Antibody 418 14 DIVMTQSPATLSVSPGERATLSCTASQSVSNNLA
VL(VK) (aa) 156WYQQKPGQAPRLLIYGASTRATGIPARFSGSGSGT

SEQ
Sequence ID Sequence Description NO.
EFTLTISSLQSEDFAVYYCQHYYNWPPWTFGQGT
NVEIK
Antibody 418_14 157 QSVSNN
CDRL1 (aa) Antibody 418_14 GAS

CDRL2 (aa) Antibody 418 14 159 QHYYNWPPWT
CDRL3 (aa) GAGGTGCAACTGGTGGAGTCTGGGGGAGGCCT
GGTCAAGCCTGGGGGGTCCCTGAGACTCTCCTG
TGCAGCCTCCGGATTCACGTTCAGTACCTATA
GCATGAACTGGGTCCGCCAGGCTCCAGGGAAG
GGGCTGGAGTGGGTCTCATCCATTAGTAGTAG
Antibody 418 14 TAGTAGTTACATATACTACGCAGACTCAGTGA
vH-(nt) ¨ 160 AGGGCCGATTCACCATCTCCAGAGACAACGCCA
AGAACTCACTGTTTCTGCAAATGAACAGCCTGA
GAGCCGAGGACACGGCTGTTTATTACTGTGCGA
GGTGGGGTTATTCCTATGACAGTCGTGGCTA
TTACCCCCGGGAACTTGACTACTGGGGCCAG
GGAACCCTGGTCACCGTCTCCTCAG
GATATTGTGATGACTCAGTCTCCAGCCACCCTG
TCTGTGTCTCCAGGGGAAAGAGCCACCCTCTCC
TGCACGGCCAGTCAGAGTGTTAGCAACAACTT
AGCCTGGTACCAGCAGAAACCTGGCCAGGCTCC
Antibody 418 14 CAGGCTCCTCATCTATGGTGCATCCACCAGGG
¨ VL(VK) (nt) 161 CCACTGGTATCCCAGCCAGGTTCAGTGGCAGTG
GGTCTGGGACAGAGITCACTCTCACCATCAGCA
GCCTGCAGTCTGAAGATTTTGCAGTTTATTACT
GTCAGCACTATTATAACTGGCCTCCGTGGAC
CTTCGGCCAAGGGACCAACGTGGAAATCAAAC
EVQLVESGGGVVQPGGSLRLSCAASGFTFNSYG
Antibody 418 40 MHWVRQAPGKGLEWVAFIRYDGGNKYYADSV
VH ( ¨ 162 KGRFTISRDNSKNTLYLQMKSLRAEDTAVYYCAN
aa) LKDSRYSGSYYDYWGQGTLVTVSS
Antibody 418 40 163 GFTFNSYG
CDRH1 (aa) Antibody 418 40 164 IRYDGGNK
CDRH2 (aa) Antibody 418 40 165 ANLKDSRYSGSYYDY

SEQ
Sequence ID Sequence Description NO.
CDRH3 (aa) VIWMTQSPSSLSASVGDRVTITCQASQDIRFYLN
Antibody 418 40 WYQQKPGKAPKLLISDASNMETGVPSRFSGSGSG

VL(VK) (aa) VDFK
Antibody 418_40 167 QDIRFY

Antibody 418 40 Antibody 418_40 GAGGTGCAGCTGGTGGAGTCTGGGGGAGGCGT
GGTCCAGCCTGGGGGGTCCCTGAGACTCTCCTG
TGCAGCGTCTGGATTCACCTTCAATAGTTATG
GCATGCACTGGGTCCGCCAGGCTCCAGGCAAG
GGGCTGGAGTGGGTGGCATTTATACGGTATGA
Antibody 418 40 TGGAGGTAATAAGTACTATGCAGACTCCGTGA

VH(VK) (nt) AGAACACGCTGTATCTGCAAATGAAGAGCCTG
AGAGCTGAGGACACGGCTGTGTATTACTGTGC
GAACCTGAAAGATAGCAGATATAGTGGGAGC
TATTATGACTACTGGGGCCAGGGAACCCTGGT
CACCGTCTCCTCAG
GTCATCTGGATGACCCAGTCTCCATCCTCCCTGT
CTGCATCTGTAGGAGACAGAGTCACCATCACTT
GCCAGGCGAGTCAGGACATTAGGTTCTATTTA
AATTGGTATCAGCAGAAACCAGGGAAAGCCCC
Antibody 418 40 TAAGCTCCTGATCTCCGATGCATCCAATATGGA

VL (VK) (lit) GATCTGGGACCGATTTTACTTTCACCATCAGCA
GCCTTCAGCCTGAAGATATTGCAACATATTACT
GTCAACAGTATGATAATCTCCCTTTCACTTTC
GGCCCTGGGACCAAGGTGGATTTCAAAC
EVQLVQSGAEVKKPGASVKVSCKASGYTFT
GYHMI-IWVRQAPGQGLEWIVIGWINPNSGGT
Antibody 418 15 172 NYVQKF QGRVTMTRDT SI S TAYMEL SRLR SD
VH (aa) DTAVYYCAKVVAVAGPFDHWGQGTLVTVS
Antibody 418 15 173 GYTFTGYH

SEQ
Sequence ID Sequence Description NO.
CDRH1 (aa) Antibody 418_15 174 INPNSGGT
CDRH2 (aa) Antibody 418 15 175 AKVVAVAGPFDH
CDRH3 (aa) QSVLIQPASVSGSPGQSITISCIGTSSDVGSYNLVS
WYQQHPGKAPKLMIYEGSKRPSGVSNRFSGSKSG
Antibody 418-15 176 NTASLTISGLQAEDEADYYCFSYAGSSDWVEGGG
VL (aa) TKLTVL
Antibody 418_15 177 SSDVGSYNL

Antibody 418 15 178 EGS

Antibody 418_15 179 FSYAGSSDWV

GAGGTGCAGCTGGTGCAGTCTGGGGCTGAGGT
GAAGAAGCCTGGGGCCTCAGTGAAGGTCTCCTG
CAAGGCTTCTGGATACACCTTCACCGGCTACC
ATATGCACTGGGTGCGACAGGCCCCTGGACAA
GGGCTTGAGTGGATGGGATGGATCAACCCTAA
CAGTGGTGGCACAAACTATGTACAGAAGTTTC
Antibody 418-15 VH ( 180 AGGGCAGGGTCACCATGACCAGGGACACGTCC
nt) ATCAGCACAGCCTACATGGAGCTGAGCAGGCT
GAGATCTGACGACACGGCCGTGTATTACTGTGC
GAAGGTTGTAGCAGTGGCTGGCCCCTTTGAC
CACTGGGGCCAGGGAACCCTGGTCACCGTCTCC
TCAG
CAGTCTGTGTTGACTCAGCCTGCCTCCGTGTCTG
GGTCTCCTGGACAGTCGATCACCATCTCCTGCA
CTGGAACCAGCAGTGATGTTGGGAGTTATAA
CCTTGTCTCCTGGTACCAACAGCACCCAGGCAA
AGCCCCCAAACTCATGATTTATGAGGGCAGTA
Antibody 418 15 181 AGCGGCCCTCAGGGGTTTCTAATCGCTTCTCTG
VL (nt) GCTCCAAGTCTGGCAACACGGCCTCCCTGACAA
TCTCTGGGCTCCAGGCTGAGGACGAGGCTGATT
ATTACTGCTTCTCATATGCAGGTAGTAGTGAT
TGGGTGTTCGGCGGAGGGACCAAGCTGACCGT
CCTAG
EVQLVESGGGLVKPGGSLRLSCAASGFTFSTYSM
Antibody 418 16 182 NWVRQAPGKGLEWVSSVSISSSYIYYADSVKGRF

SEQ
Sequence ID Sequence Description NO.
VH (aa) TISRDNAKNSLYLQMNNVRAEDTAVYYCARVRP
HNYDSSGYYPDAFDIWGQGTMVTVS S
Antibody 418_16 183 GFTFSTYS
CDRH1 (aa) Antibody 418_16 184 VSISSSYI
CDRH2 (aa) Antibody 418 16 185 ARVRPHNYDSSGYYPDAFDI
CDRH3 (aa) VIWMTQ SPA TLSVSPGFRA TLSCRA SQSVSSNLA
Antibody 418 16 WYQ QKPGQAPRLLIYGASTRAT SVPARF S GS
GSG
¨ 186 TEFTLTIS SLQSEDFAVYYCQHYYNWPPWTFGQG
VL(VK) (aa) TKVEVK
Antibody 418 16 187 QSVSSN

Antibody 418 16 GAS

Antibody 418 16 189 QHYYNWPPWT

GAGGTGCAGCTGGTGGAGTCTGGGGGAGGC CT
GGTCAAGCCTGGGGGGTCCCTGAGACTTTCCTG
TGCAGCCTCTGGATTCACCTTCAGTACCTATA
GCATGAACTGGGTCCGCCAGGCTCCAGGGAAG
GGGCTGGAGTGGGTCTCATCCGTTAGTATTAG
Antibody 418 16 TAGTAGTTACATATATTACGCAGACTCAGTGA
¨ 190 AGGGCCGATTCAC CATCTCCAGAGACAA CGC CA
VH (nt) AGAACTCACTGTATCTGCAAATGAACAACGTGA
GAGCCGAGGACACGGCCGTGTATTACTGTGCG
AGAGTTCGCCCCCATAACTATGATAGTAGTG
GTTATTATCCGGATGCTTTTGATATCTGGGGC
CAAGGGACAATGGTCA CC GTCTCTTCAG
GTCATCTGGATGAC CCAGTCTCCAGC CAC CCTG
TCTGTGTCTCCAGGGGAAAGAGCCACCCTCTCC
TGCAGGGC CAGTCAGAGT GT TA GCA GCAA CTT
Antibody 418 16 191 AGCCTGGTACCAGCAGAAACCTGGCCAGGCTCC
VL (VK) (nt) CAGGCTCCTCATCTATGGTGCATCCACCAGGG
CCACCAGTGTCCCAGCCAGGTTCAGTGGCAGTG
GGTCTGGGACAGAGTTCACTCTCACCATCAGCA
GCCTGCAGTCTGAAGATTTTGCAGTTTATTACT

SEQ
Sequence ID Sequence Description NO.
GTCAGCATTATTATAACTGGCCTCCGTGGAC
GTTCGGCCAAGGGACCAAGGTGGAAGTCAAAC
EVQLVESGGGLVKPGGSLRL S CAA S GFTFSSYTM
Antibody 418 17 NWVRQAPGKGLEWVS SISSSGSYIYYADSVKGRF
¨ 192 TISRDSAKTSLYLQMNSLRAEDTAVYFCARDLMS
VH (aa) RSIFSGYYPDAFDIWGQGTMVTVSS
Antibody 418 17 193 GFTFSSYT
CDRH1 (aa) Antibody 418 17 194 IS SSGSYI
CDRH2 (aa) Antibody 418 17 195 ARDLMSRSIFSGYYPDAFDI
CDRH3 (aa) EIVMMQSPVTLSVSPGERATLSCRASQSVSSNLA
Antibody 418 17 WYQ QKPGQAPRLLIY GASTRATGIPARF S GSGS
GT
¨ 196 EFTLSISSMQSEDFAVYYCQHYYNWPPWTFGQG
VL(VK) (aa) TTVEIK
Antibody 418_17 1,7 QSVSSN

Antibody 418_17 198 GAS

Anti body 418_17 199 wyy-Nwppwi, GAGGTGCAGCTGGTGGAGTCTGGGGGAGGC CT
GGTCAAGCCTGGGGGGTCCCTGAGACTCTCCTG
TGCAGCCTCTGGATTCACCTTCAGTTCCTATA
CCATGAACTGGGTCCGC CAGGCTCCAGGGAAG
GGGCTGGAGTGGGTCTCATCCATTAGTAGTAG
Antibody 418 17 TGGTAGTTACATATATTACGCAGACTCAGTGA
¨ 200 AGGGCCGATTCAC CATCTCCAGAGACAGCGC CA
VH (nt) AGACCTCACTGTATCTACAAATGAACAGCCTGA
GAGC CGAGGACACGGCTGTGTATTTCTGTGC GA
GAGATCTTATGAGTAGGAGCATCTTCTCTGG
TTATTATCCTGATGCTTTTGATATCTGGGGCC
AAGGGACAATGGTCACCGTCTCTTCAG
GAAATAGTGATGATGCAGTCTCCAGTCACCCTG
TCTGTGTCTCCAGGGGAAAGAGCCACCCTCTCC
Antibody 418 17 VL (VK) (nt) AGCCTGGTACCAGCAGAAACCTGGCCAGGCTCC
CAGGCTCCTCATCTATGGTGCATCCACCAGGG

SEQ
Sequence ID Sequence Description NO.
CCACTGGTATCCCAGCCAGGTTCAGTGGCAGTG
GGTCTGGGACAGAGTTCACTCTCAGCATCAGCA
GCATGCAGTCTGAAGATTTTGCAGTTTATTACT
GTCAGCACTATTATAACTGGCCTCCGTGGAC
GTTCGGCCAAGGGACCACGGTGGAAATCAAAC
QVQLVESGGGLVKPGGSLRLSCAASGFTFRSYSI
Antibody 418 18 HWVRQAPGKGLEWVS SISRSSNYIYYADSVKGRF
VH ( ¨ 202 TV SRDNAKD S LYLQMN GLRAEDTAVYYCARDL
aa) QSSSGWYWDAFDIWGQGTMVTVSS
Antibody 418 18 203 GFTFRSYS
CDRH1 (aa) Antibody 418_18 204 ISRSSNYI
CDRH2 (aa) Antibody 418 18 205 ARDLQS SSGWYWDAFDI
CDRH3 (aa) QSVLTQPPSVSGAPGQRVTIS CTGSSSNIGAGYDV
Antibody 418 18 HWYQHLPGTAPKLLIYGNNNRP SGVPDRFSGSKS
¨ VL ( 206 GTSA SLAITGLQAEDEADYYCQSFDNTHVVFGGG
aa ) TKLTVL
Antibody 418_18 207 SSNIGAGYD
CDRL I
Antibody 418 18 Antibody 418_18 209 Q SFDN THV V

CAGGTACAGCTGGTGGAGTCTGGGGGAGGCCT
GGTCAAGCCTGGGGGGTCCCTGAGACTCTCATG
TGCAGCCTCTGGATTCACCTTCCGTAGTTATA
GCATACACTGGGTCCGCCAGGCTCCAGGGAAG
GGGCTGGAGTGGGTCTCTTCCATTAGTCGTAG
Antibody 418 18 TAGTAATTACATATACTACGCAGACTCAGTGA
¨ 210 AGGGCCGATTCAC CGTCTCCAGAGACAA CGC CA
VH (nt) AGGACTCACTGTATCTGCAAATGAACGGCCTGA
GAGCCGAGGACACGGCTGTGTATTACTGTGCG
AGAGATCTACAATCCAGCAGTGGCTGGTACT
GGGATGCTTTTGATATCTGGGGCCAAGGGACA
ATGGTCACCGTCTCTTCAG
CAGTCTGTGTTGACGCAGCCGCCCTCAGTGTCT
Antibody 418 18 211 GGGGCCCCAGGGCAGAGGGTCACCATCTCCTGC

SEQ
Sequence ID Sequence Description NO.
VL (nt) ACTGGGAGCAGCTCCAACATCGGGGCAGGTT
ATGATGTACACTGGTACCAGCACCTTCCAGGAA
CAGCCCCCAAACTCCTCATCTATGGTAACAACA
ATCGGCCCTCAGGGGTCCCTGACCGATTCTCTG
GCTCCAAGTCTGGCACCTCAGCCTCCCTGGCCA
TCACTGGGCTCCAGGCTGAGGATGAGGCTGATT
ATTACTGCCAGTCGTTTGACAACACCCATGTG
GTATTCGGCGGAGGGACCAAGCTGACCGTCCT
AG
QVQLQESGPGLVRPSETLSLTCAVSGYSISSGYY
Antibody 418 19 VH ( ¨

aa) GTVRLNWFDPWGQGTLVTVSS
Antibody 418 19 13 GYSISSGYY

CDRH1 (aa) Antibody 418 19 14 IYHSGSA

CDRH2 (aa) Antibody 418 19 5 AREAVAGTVRLNWFDP

CDRH3 (aa) AIRMTQSPSSLSASVGDRVTITCRPSQTISSYLNW
Antibody 418 19 YQQKPGKAPKLITYGASSLQSGVPSRFSGSESGID
¨ VL(VK) (aa) 216 FTLTISSLQPEDFATYYCQQSYSTPFTFGPGTKVDI
Antibody 418 19 17 QTISSY

Antibody 418 19 GAS

Antibody 418 19 219 QQSYSTPFT

CAGGTCCAGCTACAGGAGTCGGGCCCAGGACT
GGTGAGGCCTTCGGAGACCCTGTCCCTCACCTG
CGCTGTCTCTGGTTACTCCATCAGCAGTGGCT
ATTACTGGGGCTGGATCCGGCAGCCCCCAGGG
Antibody 418 19 AAGGGGCTGGAGTGGATTGGGAGTATCTATCA
VH (nt) AGAGTCGACTCACCATATCAGCAGACACGTCCA
AGAACCAGTTCTCCCTGAAGCTGAGCTCTGTGA
CCGCCGCAGACACGGCCGTGTATTACTGTGCGA
GAGAGGCAGTGGCTGGTACCGTGCGGCTGA

SEQ
Sequence ID Sequence Description NO.
ACTGGTTCGACCCCTGGGGCCAGGGGACCCTG
GTCACCGTTTCCTCAG
GCCATCCGGATGACCCAGTCTCCATCCTCCCTG
TCTGCATCTGTAGGAGACAGAGTCACCATCACT
TGCCGGCCAAGTCAGACCATTAGCAGCTATTT
AAATTGGTATCAGCAGAAACCAGGGAAAGCCC
Antibody 418 19 CTAAGCTCCTCATCTATGGTGCATCCAGTTTGC
¨ 221 AAAGTGGGGTCCCATCAAGGTTCAGTGGCAGTG
VL (VK) (nt) AATCTGGGATAGATTTCACTCTCACCATCAGCA
GTCTGCAACCTGAAGATTTTGCAACTTACTACT
GTCAACAGAGTTACAGTACTCCATTCACTTTC
GGCCCTGGGACCAAAGTGGATATCAAAC
QVQLQESGPGLVKPSETLSLTCTVSGGSISNY
Antibody 418-20 222 YWSWIRQPPGKGLEWIGYIYHSVSTNYNPSL
VH (aa) K SRVTI S VD T SKNQF SLKL S SVTAADTAVYYC

Antibody 418_20 223 GGSISNYY
CDRH1 (aa) Antibody 418 20 224 IYHSVST
CDRH2 (aa) Antibody 418_20 225 ARDHRFGEFGRMSWFDP
CDRH3 (aa) EIVMMQSPATLSVSPGERATLSCRASQSVSSNLA
Antibody 418 20 WYQQKPGQAPRLLIYGASTRATGIPARFSGSGSGT
¨ VL(VK)( ) aa VEIK
Antibody 418 20 QSVSSN

Antibody 418 20 GAS

Antibody 418 20 HQYNNWPRT

CAGGTACAGCTGCAGGAGTCGGGCCCAGGACT
GGTGAAGCCTTCGGAGACCCTGTCCCTCACCTG
Antibody 418 20 0 CACTGTCTCTGGTGGCTCCATCAGTAATTACT

VU (nt) ACTGGAGCTGGATCCGGCAGCCCCCAGGGAAG
GGACTGGAATGGATTGGGTATATCTATCACAG
TGTGAGCACCAACTACAACCCCTCCCTCAAGA

SEQ
Sequence ID Sequence Description NO.
GTCGAGTCACCATATCAGTAGACACGTCCAAGA
ACCAGTTCTCCCTGAAGCTGAGCTCTGTGACCG
CTGCGGACACGGC CGTGTATTACTGTGC GAGA
GATCATAGGTTCGGGGAGTTTGGGAGAATGA
GCTGGTTCGACCCCTGGGGCCAGGGAACCCTG
GTCACCGTCTCCTCAG
GAAATAGTGATGATGCAGTCTCCAGCCACCCTG
TCTGTGTCTCCAGGGGAAAGAGCCACCCTCTCC
TGCAGGGCCAGTCAGAGTGTTAGCAGCAACTT
AGCCTGGTACCAGCAGAAACCTGGCCAGGCTCC
CAGGCTCCTCATCTATGGTGCATCCACCAGGG
Antibody 418-20 231 CCACTGGTATCCCAGCCAGGTTCAGTGGCAGTG
VL (VK) (nt) GGTCTGGGACAGAGTTCACTCTCACCATCAGCA
GCCTGCAGTCTGAAGATTTTGCAGTTTATTACT
GTCACCAGTATAATAACTGGCCTCGGACGTT
CGGCCAAGGGACCAAGGTGGAAATCAAAC
QVQLQESGPGLVKPSETLSLSCTVSGGSISNYYWS
Antibody 418 21 WIRQPPG KG LEWIGYIYYTGSTYYNP SLKSRVTIS

VH (aa) TENWFDPWGQGTLVTVSS
Antibody 418_21 CDRH1 (aa) Antibody 418_21 234 1YYTGST
CDRH2 (aa) Antibody 418 21 235 AREV1-INWNTENWFDP
CDRH3 (aa) QPVLTQSPGTLSLSPGERATL SCRA SQSVSSTYLA
Antibody 418 21 WYQQKPGQAPRLLIYGASSRATGIPDRFSGSGSGT
¨ 236 DFTLTISRLEPEDFAVYYCQQYGG SPPLITFGQGT
VL(VK) (aa) RLEIK
Antibody 418_21 Antibody 418 21 GAS

Antibody 418_21 239 QQYGGSPPLIT

CAGGTGCAGCTGCAGGAGTCGGGCCCAGGACT
Antibody 418 21 VH (nt) CACTGTCTCTGGTGGCTCCATCAGTAATTACT

SEQ
Sequence ID Sequence Description NO.
ACTGGAGCTGGATCCGGCAGCCCCCAGGGAAG
GGACTGGAGTGGATTGGGTATATCTATTACAC
TGGGAGCACCTACTACAACCCCTCCCTCAAGA
GTCGAGTCACCATATCAGTAGACACGTCCAAGA
ACCAGTTCTCCCTGAAGCTGAGTTCTGTGACCG
CTGCGGACACGGCCGTCTATTACTGTGCGAGA
GAAGTTCATAACTGGAACACAGAAAACTGGT
TCGACCCCTGGGGCCAGGGAACCCTGGTCACC
GTCTCCTCAG
CAGCCTGTGCTGACTCAGTCTCCAGGCACCCTG
TCTITGICTCCAGGGGAAAGAGCCACCCICTCC
TGCAGGGCCAGTCAGAGTGTTAGCAGCACCT
ACTTAGCCTGGTACCAGCAGAAACCTGGCCAG
GCTCCCAGGCTCCTCATCTATGGTGCATCCAGC
Antibody 418 21 241 AGGGCCACTGGCATCCCAGACAGGTTCAGTGGC
VL (VK) (nt) AGTGGGTCTGGGACAGACTTCACTCTCACCATC
AGCAGACTGGAGCCTGAGGATTTTGCAGTGTAT
TACTGTCAGCAGTATGGTGGCTCACCTCCGC
TGATCACCTTCGGCCAAGGGACACGACTGGAG
ATTAAAC
QVQLVQSGAEVKKPGA SVKVSCKA SGYTFTSYA
Antibody 418 22 MHWVRQAPGQRLEWMGWINAGSGNTKYSQKF
VH ( ¨ 242 QGRVTITRDTSASTAYMELSSLRSEDTAVYYCAR
aa) EGTEGVRFLEYLFGTWFDPWGQGTLVTVSS
Antibody 418 22 243 GYTFTSYA
CDRHI (aa) Antibody 418 22 244 INAGSGNT
CDRH2 (aa) Antibody 418_22 245 AREGTEGVRFLEYLFGTWFDP
CDRH3 (aa) QAGLTQSPATLSLSPGERATLSCRASQSVISYLAW
Antibody 418 22 YQQKPGHAPRLLIYDASNRATGIPARFSGSGSGTD
¨ VL(VK) (aa) 246 FTLTISSLEPEDFAVYYCQQRSHWPETFGQGTKV
EIK
Antibody 418 22 247 QSVISY
CDRL I
Antibody 418_22 DAS

Antibody 418 22 249 QQRSHWPET

SEQ
Sequence ID Sequence Description NO.

CAGGTCCAGCTGGTGCAGTCTGGGGCTGAGGTG
AAGAAGCCTGGGGCCTCAGTGAAGGTTTCCTGC
AAGGCTTCTGGATACACCTTCACTAGCTATGC
TATGCATTGGGTGCGCCAGGCCCCCGGACAAA
GGCTTGAGTGGATGGGATGGATCAACGCTGGC
AGTGGTAATACAAAATATTCACAGAAGTTCCA
( Antibody 418-22 250 GGGCAGAGTCACCATTACCAGGGACACATCCG
VH nt) CGAGCACAGCCTACATGGAGCTGAGCAGCCTG
AGATCTGAAGACACGGCTGTGTATTACTGTGCG
AGAGAGGGGACCGAAGGCGTACGATTTTTG
GAGTACTTATTCGGAACCTGGTTCGACCCCT
GGGGCCAGGGAACCCTGGTCACCGTCTCCTCAG
CAGGCAGGGCTGACTCAGTCTCCAGCCACCCTG
TCTTTGTCTCCAGGGGAAAGAGCCACCCTCTCC
TGCAGGGCCAGTCAGAGTGTTATCAGCTACTT
AGCCTGGTACCAACAGAAACCTGGCCACGCTCC
CAGGCTCCTCATCTATGATGCATCCAACAGGG
Antibody 418-22 251 CCA CTGGCA TC CC A GC CAGGTTCAGTGGC AGTG
VL (VK) (nt) GGTCTGGGACAGACTTCACTCTCACCATCAGCA
GCCTAGAGCCTGAAGATTTTGCAGTTTATTACT
GTCAGCAGCGTAGCCACTGGCCTGAGACGTT
CGGCCAAGGGACCAAGGTGGAAATCAAAC

Antibody 418 23 GVSWIRQPPGKALEWLAHIFSNDEKSYSTSLKSR
VH ( 252 LTISKDTSKSQVVLTMTNMDPVDTATYYCARID
aa) WWSSYLVGDYWGQGTLVTVSS
Antibody 418 23 253 GFSLSNAKMG
CDRH1 (aa) Antibody 418 23 254 IFSNDEK
CDRH2 (aa) Antibody 418 23 255 ARIDWWSSYLVGDY
CDRH3 (aa) QSVLTQPPSVSGAPGQRVTISCTGSSSNIGAGYHV
HWYQQLPGTAPKLLIYGNSNRPSGVPDRFSGSKS
Antibody 418-23 256 GTSASLAITGLQAEDEADYYCQSYDSSLSVVFGG
VL (aa) GTKL'TVL
Antibody 418_23 SSNIGAGYH

SEQ
Sequence ID Sequence Description NO.
Antibody 418 23 GNS

Antibody 418 23 CAGGTCACCTTGAGGGAGTCTGGTCCTGTGCTG
GTAAAACCCACAGAGACCCTCACGCTGACCTGC
ACCGTCTCTGGGTTCTCACTCAGCAATGCTAA
AATGGGTGTGAGCTGGATCCGTCAGCCCCCAG
GGAAGGCCCTGGAGTGGCTTGCACACATTTTTT
Antibody 418 23 CGAATGACGAAAAATCCTACAGCACATCTCTG

VH (nt) AAAAGCCAGGTGGTCCTTACTATGACCAACATG
GACCCTGTGGACACAGCCACATATTACTGTGCA
CGGATAGATTGGTGGAGTAGTTATTTAGTTG
GTGACTACTGGGGCCAGGGAACCCTGGTCACC
GTCTCCTCAG
CAGTCTGTGCTGACGCAGCCGCCCTCAGTGTCT
GGGGCCCCAGGGCAGAGGGTCACCATCTCCTGC
ACTGGGAGCAGCTCCAACATCGGGGCGGGTT
ATCATGTACACTGGTACCAGCAGCTTCCAGGAA
CAGCCCCCAAACTCCTCATCTATGGTAACAGC
Antibody 418 23 61 AATCGGCCCTCAGGGGTCCCTGACCGATTCTCT

VL (nt) GGCTCCAAGTCTGGCACCTCAGCCTCCCTGGCC
ATCACTGGGCTCCAGGCTGAGGATGAGGCTGAT
TATTACTGCCAGTCCTATGACAGCAGTCTGAG
TGTGGTATTCGGCGGAGGGACCAAGCTGACCG
TCC TAG
EVQLVESGGGLVKPGGSLRLSCAASGFTFSSYSM
NWVRQAPGKGLEWVSSISSSRGYIYYADSVKGRF
Antibody 418-24 VH ( 262 TISRDNAKNSLYLQMNSLRAEDTAVYYCARWLT
aa) YYYDSSGYFPSPFDYWGQGTLVTVSS
Antibody 418 24 63 GFTFSSYS

CDRH1 (aa) Antibody 418 24 264 ISSSRGYI
CDRH2 (aa) Antibody 418_24 265 ARWLTYYYDSSGYFPSPFDY
CDRH3 (aa) Antibody 418 24 266 EIEMMQSPATLSVSPGERATLSCRASQSVSSNLA
VL(VK) (aa) WYQQKPGQAPRLLIYGASTRATGIPARFSGSGSGT

SEQ
Sequence ID Sequence Description NO.
EFTLTISSLQSEDFAVYYCQQYYNWPPWTFGQGT
KVEIK
Antibody 418_24 267 QSVSSN

Antibody 418_24 GAS

Antibody 418_24 269 QQYYNWPPWT

GAGGTGCAGCTGGTGGAGTCTGGGGGAGGCCT
GGTCAAGCCCGGGGGGTCCCTGAGACTCTCCTG
TGCAGCCTCTGGATTCACCTTCAGTAGCTATA
GTATGAACTGGGTCCGCCAGGCTCCAGGAAAG
GGGCTGGAGTGGGTCTCATCCATTAGTAGTAG
Antibody 418 24 TAGAGGTTACATATACTACGCAGACTCAGTGA

VH (nt) AGAACTCACTGTATCTGCAAATGAACAGCCTGA
GAGCCGAGGACACGGCTGTGTATTACTGTGCG
AGATGGCTTACATATTACTATGATAGTAGTG
GTTATTTCCCCTCGCCTTTTGACTACTGGGGC
CAGGGAACCCIGGICACCGTCTCCTCAG
GAAATAGAGATGATGCAGTCTCCAGCCACCCTG
TCTGTGTCTCCAGGGGAAAGAGCCACCCTCTCC
TGCAGGGCCAGTCACAC TC TTAC CAC CAACTT
AGCCTGGTACCAGCAGAAACCTGGCCAGGCTCC
CAGGCTCCTCATCTATGGTGCATCCACCAGGG
Antibody 418-24 271 CCACTGGTATCCCAGCCAGGTTCAGTGGCAGTG
VL (VK) (nt) GGTCTGGGACAGAGTTCACTCTCACCATCAGCA
GCCTGCAGTCTGAAGATTTTGCAGTTTATTACT
GTCAGCAGTATTATAACTGGCCTCCGTGGAC
GTTCGGCCAAGGGACCAAGGTGGAAATCAAAC
EVQLVESGGGLVKPGGSLRLSCAASGFTFSSYSM
Antibody 418 25 NWVRQAPGKGLEWVSSISSSRSFIVYADSVKGRI
¨ VH ( ) 272 TISRDNAKNSLYLQMNSLRAEDTAVYYCARVKIT
aa NYYDSSGYYPDAFDIWGQGTMVTVSS
Antibody 418_25 273 GFTFSSYS
CDRH1 (aa) Antibody 418_25 274 ISSSRSFI
CDRH2 (aa) Antibody 418_25 275 ARVKITNYYDSSGYYPDAFDI

SEQ
Sequence ID Sequence Description NO.
CDRH3 (aa) DIVMTQSPATLSVSPGERATLSCRASQSVSSNLAW
Antibody 418 25 YQQKPGQAPRLLIYGAS I RATGVPARFTGSGSGT
¨ 276 DFTLTISSMQSEDFAVYYCQQYYNWPPWTFGQG
VL(VK) (aa) TKVEIK
Antibody 418_25 Antibody 418 25 GAS

Antibody 418_25 279 wyyNwppwi-GAGGTGC A A CTGGTGGAGTCTGGGGGAGGCCT
GGTCAAGCCTGGGGGGTCCCTGAGACTCTCCTG
TGCAGCCTCTGGATTCACCTTCAGTAGCTATA
GCATGAACTGGGTCCGCCAGGCTCCAGGGAAG
GGGCTGGAGTGGGTCTCATCCATTAGTAGTAG
Antibody 418 25 TAGGAGTTTCATATACTACGCAGACTCAGTGA
VH (nt) ¨ 280 AGGGCCGAATCACCATCTCCAGAGACAACGCC
AAGAACTCACTGTATCTGCAAATGAACAGCCTG
AGAGCCGAGGACACGGCTGTGTATTACTGTGC
GAGAGTGAAAATTACGAATTACTATGATAGT
AGTGGTTATTACCCTGATGCTTTTGATATCTG
GGGCCAAGGGACAATGGTCACCGTCTCTTCAG
GATATTGTGATGACTCAGTCTCCAGCCACCCTG
TCTGTGTCTCCAGGGGAAAGAGCCACCCTCTCC
TGCAGGGCCAGTCAGAGTGTTAGCAGCAACTT
AGCCTGGTACCAGCAGAAACCTGGCCAGGCTCC
Antibody 418-25 281 CAGGCTCCTCATCTATGGTGCATCCACCAGGG
CCACTGGTGTCCCAGCCAGGTTCACTGGCAGTG
VL (VK) (lit) GGTCTGGGACAGATTTCACTCTCACCATCAGCA
GCATGCAGTCTGAAGATTTTGCAGTTTATTACT
GTCAGCAGTATTATAACTGGCCTCCGTGGAC
GTTCGGCCAAGGGACCAAGGTGGAAATCAAAC
EVHLVQSGAEVKKPGASVKVSCKVSGYTLTELS
Antibody 418 26 MHWVRQAPGKGLEWMGGFDPQDAETIYAQKFQ
VH ( ¨ 282 GRVTMTEDTSTDTAYMELSSLRSEDTAVYYCVT
aa) ATAVAGTPDLYYYHYGLDVWGQGTTV'TVSS
Antibody 418_26 83 GYTLTELS

CDRH1 (aa) SEQ
Sequence ID Sequence Description NO.
Antibody 418 26 284 FDPQDAET
CDRH2 (aa) Antibody 418_26 285 VTATAVAGTPDLYYYHYGLDV
CDRH3 (aa) QTVVTQTPLSSPVTLGQPASISCRSSQSLVHSDGN
TYLSWLQQRPGQPPRLLIYKISNRFSGVPDRFSGS
Antibody 418-26 ( 286 GAGTDFTLQISRVEAEDVGVYYCMQATQFPRTF
VL(VK) aa) GQGTKVEIK
Antibody 418_26 287 QSLVHSDGNTY

Antibody 418 26 KIS

Antibody 418 26 289 MQATQFPRT

GAGGTGCATCTGGTACAATCTGGGGCTGAGGTG
AAGAAGCCTGGGGCCTCAGTGAAGGTCTCCTGC
AAGGTTTCCGGATACACCCTCACTGAATTATC
CATGCACTGGGTGCGACAGGCTCCTGGAAAAG
GGCTTGAGTGGATGGGAGGTTTTGATCCTCAA
Antibody 418 26 GATGCTGAAACAATCTACGCACAGAAGTTCCA
¨ 290 GGGCAGAGTCACCATGACCGAGGACACATCTA
VH (nt) CAGACACAGCCTACATGGAACTGAGCAGCCTG
AGATCTGAGGACACGGCCGTGTATTACTGTGTA
ACAGCGACAGCAGTGGCTGGCACCCCAGAC
CTATACTACTACCACTACGGTTTGGACGTCT
GGGGCCAAGGGACCACGGTCACCGTCTCCTCA
CAGACTGTGGTGACCCAGACTCCACTCTCCTCA
CCGGTCACCCTTGGACAGCCGGCCTCCATCTCC
TGCAGGTCTAGTCAAAGCCTCGTCCACAGTGA
TGGAAACACCTACTTGAGTTGGCTTCAGCAGA
GGCCAGGCCAGCCTCCAAGACTCCTAATTTATA
Antibody 418 26 291 AGATTTCTAACCGGTTCTCTGGGGTCCCAGACA
VL (VK) (nt) GATTCAGTGGCAGTGGGGCAGGGACAGATTTC
ACACTGCAAATCAGCAGGGTGGAAGCTGAGGA
TGTCGGGGTTTATTACTGCATGCAAGCTACAC
AG TTTCCTCG TACG TTCGGCCAAGGGACCAAG
GTGGAAATCAAAC
Antibody 418 27 EVQLVESGGGLVRPGGSLRLSCAASGFTFSSCGM
VH (aa) 292 NWVRQAPGKGLEWVSSISRSSNYIYYADSVKGRF

SEQ
Sequence ID Sequence Description NO.
TISRDNAKNSLYLQMNSLRAEDTAVYYCARIPHT
SLYGDYRDDYYYYYGMDVWGQGTTVTVSS
Antibody 418_27 293 GFTFSSCG
CDRH1 (aa) Antibody 418_27 294 ISRSSNYI
CDRH2 (aa) Antibody 418_27 295 ARIPHTSLYGDYRDDYYYYYGMDV
CDRH3 (aa) EIVMTQSPATLSVSPGERATLSCRASQSVSSNLAW
Antibody 418 27 YQQKPGQAPRLLIYGASTRATGIPARFSGSGSGTE
¨ VL(VK) (aa) 296 FTLTISSLQSEDFAVYYCQQYNNWPPLTFGGGTK
VEIK
Antibody 418 27 Antibody 418 27 GAS

Antibody 418 27 GAGGTGCAGCTGGTGGAGTCTGGGGGAGGCCT
GGTCAGGCCTGGGGGGTCCCTGAGACTCTCCTG
TGCAGCCTCTGGATTCACCTTCAGTAGCTGTG
GCATGAACTGGGTCCGCCAGGCTCCAGGGAAG
GGGCTGGAGTGGGTCTCATCCATTAGTAGGAG
TAGTAATTATATATACTACGCAGACTCAGTGA
Antibody 418 27 300 AGGGCCGATTCACCATCTCCAGAGACAACGCCA
VH (nt) AGAACTCACTGTATCTGCAAATGAACAGCCTGA
GAGCCGAGGACACGGCTGTATATTACTGTGCG
AGAATCCCCCACACCTCACTCTACGGTGACT
ACCGGGATGATTACTACTATTACTACGGTAT
GGACGTCTGGGGCCAAGGGACCACGGTCACCG
TCTCCTCA
GAAATAGTGATGACGCAGTCTCCAGCCACCCTG
TCTGTGTCTCCAGGGGAAAGAGCCACCCTCTCC
TGCAGGGCCAGTCAGAGTGTTAGCAGCAACTT
Antibody 418 27 AGCCTGGTACCAGCAGAAACCTGGCCAGGCTCC
VL (VK) (nt) .. 301CAGACTCCTCATCTATGGTGCATCCACCAGGG
CCACTGGTATCCCAGCCAGGTTCAGTGGCAGTG
GGTCTGGGACAGAGITCACTCTCACCATCAGCA
GCCTGCAGTCTGAAGATTTTGCAGTTTATTACT

SEQ
Sequence ID Sequence Description NO.
GTCAGCAGTATAATAACTGGCCTCCGCTCAC
TTTCGGCGGAGGGACCAAGGTGGAGATCAAAC
QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYAI
Antibody 418 28 HWVRQAPGKGLEWVAVISYDRINKYYADSVKG
¨ 302 RFTISRDNSKNTLYLQMNSLRAEDTAVYYCARDE
VH (aa) LPSPYSGYDGGFLYYFDSWGQGTLVTVSS
Antibody 418 28 303 GFTFSSYA
CDRH1 (aa) Antibody 418 28 304 ISYDRINK
CDRH2 (aa) Antibody 418 28 305 ARDELP SPY SGYDGGFLYYFD S
CDRH3 (aa) NIQMTQSPSSLSASVGDRVTITCRASQGISSALAW
Antibody 418 28 YQQKPGKAPKLLIYDASSLASGVPSRFSGSGSGTD
¨ VL(VK) (aa) 306 FTLTISSLQPEDFATYYCQQFNSYPPTFGQGTKVE
IK
Antibody 418_28 307 QGIS SA

Antibody 418_28 DAS

Antibody 418_28 309 QQFNSYPPT

CAGGTGCAGCTGGTGGAGTCTGGGGGAGGCGT
GGTCCAGCCTGGGAGGTCCCTGAGACTCTCCTG
TGCAGCCTCTGGATTCACCTTCAGTAGCTATG
CTATACACTGGGTCCGCCAGGCTCCAGGCAAG
GGGCTAGAGTGGGTGGCAGTTATATCATATGA
TAGAATTAATAAATACTACGCAGACTCCGTGA
Antibody 418 28 310 AGGGCCGATTCACCATCTCCAGAGACAATTCCA
VH (nt) AGAACACGCTGTATCTGCAAATGAACAGCCTGA
GAGCTGAGGACACGGCTGTGTATTACTGTGCG
AGAGATGAGCTCCCGTCCCCATATAGTGGCT
ACGATGGGGGATTTTTATACTACTTTGACTC
CTGGGGCCAGGGAACCCTGGTCACCGTCTCCTC
AG
AACATCCAGATGACCCAGTCTCCATCCTCCCTG
Antibody 418 28 3 TCTGCATCTGTAGGAGACAGAGTCACCATCACT

VL (VK) (nt) .. TGCCGGGCAAGTCAGGGCATTAGCAGTGCTTT
AGCCTGGTATCAGCAGAAACCAGGGAAAGCTC

SEQ
Sequence ID Sequence Description NO.
CTAAACTCCTGATCTATGATGCCTCCAGTTTGG
CAAGTGGGGTCCCATCAAGGTTCAGCGGCAGTG
GATCTGGGACAGATTTCACTCTCACCATCAGCA
GCCTGCAGCCTGAAGATTTTGCAACTTATTACT
GTCAACAGTTTAATAGTTACCCTCCGACGTTC
GGCCAAGGGACCAAGGTGGAAATCAAAC
QVQLVQSGAEVKKPGASVKVSCKVSGYSLIEVS
Antibody 418 29 MEIWVRQAPGKGLEWMGGFDPENVETIYAQKFQ
VH ( 312 GRVTMTEDTSADTAYMELSSLRSEDTAVYYCAT
aa) TFAFGATTRNLVDYWGQGTLVTVSS
Antibody 418_29 CDRH1 (aa) Antibody 418_29 314 FDPENVET
CDRH2 (aa) Antibody 418 29 315 ATTFAFGATTRNLVDY
CDRH3 (aa) SYELTQPPSASGTPGQRVTISCSGSSSNIGSNYVY
WYQQVPGTAPKLLIFKNYQRPSGVPDRFSGSKSG
VL ( Antibody 418-29 316 TSASLAISGLRSEDEADYYCAAWDDTLSGVLFGG
aa) GTKLTVL
Antibody 418 29 317 SSNIGSNY

Antibody 418 29 KNY

Antibody 418 29 319 AAWDDTLSGVL

CAGGTGCAGCTGGTGCAGTCTGGGGCTGAGGTG
AAGAAGCCTGGGGCCTCAGTGAAGGTCTCCTGC
AAGGTTTCCGGATACTCCCTCATTGAAGTATC
CATGCACTGGGTGCGACAGGCTCCTGGAAAAG
GACTTGAGTGGATGGGAGGTTTTGATCCTGAA
Antibody 418 29 AATGTGGAAACAATCTACGCACAGAAGTTCCA

VH (nt) CAGACACAGCCTACATGGAGCTGAGCAGCCTG
AGATCTGAGGACACGGCCGTATATTACTGTGCA
ACAACCTTCGCCTTCGGAGCTACAACGAGGA
ACTTAGTAGACTACTGGGGCCAGGGAACCCTG
GTCACCGTCTCCTCAG

SEQ
Sequence ID Sequence Description NO.
TCCTATGAGCTGACACAGC CAC C CTCAGCGTCT
GGGACCCCCGGGCAGAGGGTCACCATCTCTTGC
TCTGGAAGCAGCTCCAACATCGGAAGTAATTA
TGTATACTGGTACCAGCAGGTCCCAGGAACGGC
CC CCAAACTCCTCATCTTTAA GAATTAT CAGCG
Antibody 418 29 321 GCCCTCAGGGGTCCCTGACCGATTCTCTGGCTC
VL (nt) CAAGTCTGGCACCTCAGCCTCCCTGGCCATCAG
TGGGCTCCGGTCCGAGGATGAGGCTGATTATTA
CTGTGCAGCATGGGATGACACCCTGAGTGGT
GTGCTATTCGGCGGAGGGACCAAGCTGACCGT
CCTAG
QVQLVQSGADVKKPGASVKVSCKASGYTFISYY
Antibody 418 30 MEIWVRQAPGQGLEWMGIINP SS GSTIYAQKFQG
VEI
¨ 322 RVTMTTDTSTSTVYMDLS SLTSEDTAVYYCARD
aa) ( Antibody 4 I 8 30 323 GYTFISYY
CDRH 1 (aa) Antibody 418_30 CDRH2 (aa) Antibody 418 30 325 ARDGRPREMIERDSSGPYFDY
CDRH3 (aa) SYELTQPPSVSVSPGQTARITCSGDALPKQYAYW
YQQKPGQAPVLVIYKDSERPSGIPERFSGS SSGTT
VL ( Antibody 418-30 326 VTLTISGVQAEDEADYYCQSTDSSGTHVVFGGGT
aa) KLTVL
Antibody 418_30 Antibody 418 30 KDS

Antibody 418_30 329 Q STD S SGTHVV

CAGGTACAGCTGGTGCAGTCTGGGGCTGATGTG
AAGAAGCCTGGGGCCTCAGTGAAGGTTTCCTGC
AAGGCATCTGGATACACCTTCATCAGTTACTA
Antibody 418 30 30 TATGCATTGGGTGCGACAGGCCCCTGGACAAG

VH (nt) GGCTTGAGTGGATGGGAATAATCAACCCTAGT
AGTGGTAGCACAATCTACGCACAGAAGTTC CA
GGGCAGAGTCACCATGACCACGGACACGTCCA
CGAGCACAGTTTACATGGACTTGAGCAGCCTGA

SEQ
Sequence ID Sequence Description NO.
CATCTGAGGACACGGCCGTGTATTACTGTGCGA
GAGATGGGCGACCGCGAGAGATGATC GAAC
GTGATAGTAGTGGGCCTTACTTTGACTACTG
GGGCCAGGGAACCCTGGTCACCATCTCCTCAG
TCCTATGAGCTGACACAGCCACCCTCGGTGTCA
GTGTCCCCAGGACAGACGGCCAGGATCACCTGC
TCTGGAGATGCA TT GCCAAAGCAA TATGCTTA
TTGGTACCAGCAGAAGCCAGGCCAGGCCCCTGT
Antibody 418 30 GCTGGTGATATATAAAGACAGTGAGAGGCCCT
VL (nt) ¨

CAGGGACAACAGTCACGTTGACCATCAGTGGA
GTCCAGGCAGAAGATGAGGCTGACTATTACTGT
CAATCAACAGACAGCAGTGGTACCCACGTGG
TATTCGGCGGAGGGACCAAGCTGACCGTCCTAG
QVQLQESGPGLVKPSQTLSLTCTVSGGSISSTSYY
Antibody 418 31 WSWIRQPAGKGLEWIGRIYNSGSTNYNPSLKSRV
VH ( aa) YDFWSGYSDWYFDLWGRGTQVTVSS
Antibody 418 31 CDRH1 (aa) Antibody 418 31 CDRH2 (aa) Antibody 418 31 335 A RDLDYYDFWSGY SDWYFDL
CDRH3 (aa) SYELTQPPSVSKGLRQTATLTCTGNSNNVGDQGA
Antibody 418 31 AWLQQHQGHPPKLL SYRNNNRP SGI SERF SA SRS
VL ( ¨

aa ) GGGTKLTVL
Antibody 418 31 SNNVGDQG

Antibody 418_31 RNN

Antibody 418 31 33, SAWDTSLSAWV

CAGGTACAGCTGCAGGAGTCGGGCCCAGGACT
GGTGAAGCCTTCACAGACCCTGTCCCTCACCTG
Antibody 418 31 VH (nt) GTTACTACTGGAGCTGGATCCGGCAGCCCGCC
GGGAAGGGACTGGAGTGGATTGGGCGTATATA

SEQ
Sequence ID Sequence Description NO.
TAACAGTGGGAGCACCAACTACAATCCCTCCC
TCAAGAGTCGAGTCACCATATCAGTAGACACGT
CCAAGAACCAGTTCTCCCTGAAGCTGACCTCTG
TGACCGCCGCCGACACGGCCGTCTATTACTGTG
CGAGAGATCTTGACTACTACGATTTTTGGAG
TGGTTATTCTGACTGGTACTTCGATCTCTGGG
GCCGTGGCACCCAGGTCACTGTCTCCTCAG
TCCTATGAGCTGACTCAGCCACCCTCGGTGTCC
AAGGGCTTGAGACAGACCGCCACACTCACCTGC
ACTGGGAACAGCAACAATGTTGGCGACCAAG
GAGCAGCTTGGCTGCAGCAGCACCAGGGCCAC
CCTCCCAAACTCCTATCCTACAGGAATAACAAC
Antibody 418 31 341 CGGCCCTCAGGGATCTCAGAGAGATTCTCTGCA
VL (nt) TCCAGGTCAGGAAACACAGCCTCCCTGACCATT
ACTGGACTCCAGCCTGAGGACGAGGCTGACTAT
TACTGCTCAGCATGGGACACCAGCCTCAGTG
CTTGGGTGTTCGGCGGAGGGACCAAACTGACC
GTCCTAA
QVQLQESGPGLVKPSETLSLTCTVSDDSISSYYWS
WIRQPPGKGLEWIGYIYYSGSTNYNPSLKSRVTIS
Antibody 418-33 342 VDTSKNQFSLNLSSVTAADTAVYYCARDRGWD
VH (aa) GYNLGFDYWGQGTLVTVSS
Antibody 418_33 CDRH1 (aa) Antibody 418_33 IYYSGST

CDRH2 (aa) Antibody 418 33 345 ARDRGWDGYNLGFDY
CDRH3 (aa) SYELTQPPSVSVAPGQTARITCGGNKIGSKSVHW
Antibody 418 33 YQQMPGQAPVLVVYDDSDRPSGIPERFSGSNSGN
¨ 346 TATLTISRVEAGDEADYYCQVVVDNNSDQGVFGG
VL (aa) GTKLTVL
Antibody 418_33 347 KIGSKS

Antibody 418 33 DDS

Antibody 418 33 349 QVWDNNSD QG V

SEQ
Sequence ID Sequence Description NO.
CAGGTGCAGCTGCAGGAGTCGGGCCCAGGACT
GGTGAAGCCTTCGGAGACCCTGTCCCTCACTTG
CACTGTCTCTGATGACTCCATCAGTAGTTACT
ACTGGAGCTGGATTCGGCAGCCCCCAGGGAAG
GGACTGGAGTGGATTGGGTATATCTATTACAG
Antibody 418 33 TGGGAGCACCAACTACAACCCCTCCCTCAAGA
¨ 350 GTCGAGTCACCATATCAGTAGACACGTCCAAGA
VH (nt) ACCAGTTCTCCCTGAACCTGAGCTCTGTGACCG
CTGCGGACACGGCCGTATATTACTGTGCGAGA
CATAGAGGATGCGATGGCTACAATTTAGGCT
TTGACTACTGGGGCCAGGGAACCCTGGTCACC
GTCTCCTCAG
TCCTATGAGCTGACTCAGCCACCCTCGGTGTCA
GTGGCCCCAGGACAGACGGCCAGGATAACCTG
TGGGGGAAACAAGATTGGAAGTAAAAGTGTG
CACTGGTACCAGCAGATGCCAGGCCAGGCCCC
GGTGCTGGTCGTCTATGATGATAGCGACCGGC
Antibody 418 33 351 CCTCAGGGATCCCTGAGCGATTCTCTGGCTC CA
VL (nt) ACTCTGGGAACACGGCCACCCTGACCATCAGCA
GGGTCGAAGCCGGGGATGAGGCCGACTATTAC
TGTCAGGTGTGGGATAATAATAGTGACCAGG
GGGTGTTCGGCGGAGGGACCAAGCTGACCGTC
CTAG
QITLKESGPALVKPTQTLTLTCTFSGFSLSTTGMR
Antibody 418 34 VSWIRQPPGKALEWLARIDWDDDKFYSTSLKTRL
VTI
¨ 352 TISKDTSKNQVVLTMTNMDPVDTGTYYCARAYG
(aa) DHEDYWGQGTLVTVSS
Antibody 418 34 353 GFSLSTTGMR
CDRHI (aa) Antibody 418_34 354 IDWDDDK
CDRH2 (aa) Antibody 418_34 355 ARAYGDHEDY
CDRH3 (aa) DIVMTQSPDSLAVSLGERATINCKSSQSVLYSSNN
KNYLVWYQQKPGQPPKLLIYWASTRESGVPDRFS
Antibody 418-34 356 GSGSGTDFTLTISSLQAEDVAVYYCQQYYSTPITF
VL(VK) (aa) GQGTRLEIK
Antibody 418 34 CDRLI

SEQ
Sequence ID Sequence Description NO.
Antibody 418 34 Antibody 418 34 359 QQYYSTPIT

CAGATCACCTTGAAGGAGTCTGGTCCTGCGCTG
GTGAAACCCACACAGACCCTCACACTGACCTGC
ACCTTCTCGGGGTTCTCACTCAGCACTACTGG
AATGCGTGTGAGCTGGATCCGTCAGCCCCCAG
GGAAGGCCCTGGAGTGGCTTGCACGCATTGAT
Antibody 418 34 60 TGGGATGATGATAAATTCTACAGCACATCTCT

VH (nt) GAAGACCAGGCTCACCATCTCCAAGGACACCTC
CAAAAACCAGGTGGTCCTTACAATGACCAACAT
GGACCCTGTGGACACAGGCACGTATTACTGTGC
ACGGGCCTACGGTGATCACGAAGACTACTGG
GGCCAGGGAACCCTGGTCACCGTCTCCTCAG
GATATTGTGATGACTCAGTCTCCAGACTCCCTG
GCTGTGTCTCTGGGCGAGAGGGCCACCATCAAC
TGCAAGTCCAGCCAGAGTGTTTTATACAGCTC
CAACAATAAGAATTACTTAGTTTGGTACCAGC
AGAAACCAGGACAGCCTCCTAAGCTGCTCATTT
Antibody 418 34 361 ACTGGGCATCTACGCGGGAATCCGGGGTCCCT
VL(VK) (nt) GACCGATTCAGTGGCAGCGGGTCTGGGACAGA
TTTCACTCTCACCATCAGCAGCCTGCAGGCTGA
AGATGTGGCAGTTTATTACTGTCAGCAATATTA
TAG TACTCCCATCACCTTCGGCCAAGGGACAC
GACTGGAGATTAAAC
EVQLVESGGGLVQPGGSLRLSCAASGFTFSSYEM
Antibody 418 35 NWVRQAPGKGLEWVSYISSSGSTIYYADSVKGRF
¨ 362 TISRDNAKNSLYLQMNSLRAEDTAVYYCARGEG
VH (aa) SGYYIFYYYGMDVWGRGTTVTVSS
Antibody 418_35 363 GFTFSSYE
CDRH1 (aa) Antibody 418_35 364 ISSSGSTI
CDRH2 (aa) Antibody 418_35 365 ARGEGSGYYIFYYYGMDV
CDRH3 (aa) Antibody 418 35 DIVMTQSPLSLSVTPGQPASISCKSSESLLHSDGK

VL(VK) (aa) TYLSWYVQKPGQPPRLLIHELSNRFPGVPDRFSGS

SEQ
Sequence ID Sequence Description NO.
GSETDFTLRISRVEAEDVGVYYCMQFGEKFTFGP
GTKVDIK
Antibody 418_35 367 ESLLHSDGKTY

Antibody 418_35 68 ELS

Antibody 418_35 369 MQFGEKFT

GAGGTGCAGCTGGTGGAGTCTGGGGGAGGCTT
GGTACAGCCTGGAGGGTCCCTGAGACTCTCCTG
TGCAGCCTCTGGATTCACCTTCAGTAGTTATG
AAATGAACTGGGTCCGCCAGGCTCCAGGGAAG
GGTCTGGAGTGGGTTTCATACATTAGTAGTAG
Antibody 418 35 TGGTAGTACCATATACTACGCAGACTCTGTGA

VH (nt) AGAACTCACTGTATCTGCAAATGAACAGCCTGA
GAGCCGAGGACACGGCTGTTTATTACTGTGCGA
GAGGGGAGGGCAGTGGCTACTATATTTTCTA
CTACTACGGTATGGACGTCTGGGGCCGAGGG
ACCACGGTCACCGTCTCCTCA
GATATTGTGATGACTCAGTCTCCACTCTCTCTGT
CCGTCACCCCTGGACAGCCGGCCTCCATCTCCT
GCAAGTCTAGTCACACCCTCCTACATACTCAT
GGAAAGACCTATTTGTCTTGGTACGTGCAGAA
GCCAGGCCAGCCTCCACGGCTCCTGATCCATGA
Antibody 418 35 371 ACTTTCCAACCGGTTCCCTGGAGTGCCAGATAG
VL(VK) (nt) GTTCAGTGGCAGCGGGTCAGAGACAGACTICAC
ACTGAGGATCAGCCGGGTGGAGGCTGAGGATG
TTGGCGTTTATTACTGCATGCAATTCGGGGAG
AAATTCACTTTCGGCCCTGGGACCAAAGTGGA
CATCAAAC
QVQLQESGPGLVKPSGTLSLTCDVSGASISSSNW
Antibody 418 37 WSWVRQPPGKGLEWIGEIYHSGNTNYNPSLKSR
VH

aa () SGRYNWFDPWGQGTLVTVSS
Antibody 418 37 GASISSSNW

CDRH1 (aa) Antibody 418 37 1YHSGNT

CDRH2 (aa) SEQ
Sequence ID Sequence Description NO.
Antibody 418 37 A SRVSGRYNWFDP

CDRH3 (aa) QSVLTQPASVSGSPGQSITISCTGTSSDVGSYNLVS
Antibody 418 37 WYQQHPGKAPKLMIYEGSKRPSGISNRFSGSKSG

VL (aa) TKLTVL
Antibody 418 37 SSDVGSYNL

Antibody 418 37 EGS

Antibody 418_37 FSYAGFSTWV

CAGGTGCAGCTGCAGGAGTCGGGCCCAGGACT
GGTGAAGCCTTCGGGGACCCTGTCCCTCACCTG
CGATGTCTCTGGTGCCTCCATCAGCAGTAGTA
ACTGC 1G GAM_ I GGG I CCGCCAGCCCCCAGGG
AAGGGGCTGGAGTGGATTGGGGAAATCTATCA
Antibody 418 37 TAGTGGGAACACCAACTACAACCCGTCCCTCA
VH

(nt) AGA ACCAGTTCTCCCTGACGGTGAGCTCTGTGA
CCGCCGCGGACACGGCCGTGTACTACTGTGCG
AGCCGAGTTTCAGGGAGGTACAACTGGTTCG
ACCCCTGGGGCCAGGGAACCCTGGTCACCGTC
TCCTCAG
CAGTCTGTGTTGACTCAGCCTGCCTCCGTGTCTG
GGTCTCCTGGACAGTCGATCACCATCTCCTGCA
CTGGAACCAGCAGTGATGTTGGCAGTTATAA
CCTTGTCTCCTGGTACCAACAGCACCCAGGCAA
AGCCCCCAAACTCATGATTTATGAGGGCAGTA
Antibody 418 37 381 AGCGGCCCTCAGGAATTTCTAATCGCTTCTCTG
VL (nt) GCTCCAAGTCTGGCAACACGGCCTCCCTGACAA
TCTCTGGGCTCCAGGCTGAGGACGAGGCTGATT
ATTACTGCTTCTCATATGCAGGTTTTAGCACT
TGGGTGTTCGGCGGAGGGACCAAGCTGACCGT
CCTAG
QVQLVESGGGVVQPGKSLRLSCAASGFTFNNYG
Antibody 418 38 MHWVRQAPGKGLEWVAVIWYDGSNKYYTDSV
( ) 382 KGRFTISRDNSKNTLYLQIVENSLRAEDTAVYYCAR
aa ETSDYGDYIRLRRNAFDIWGQGTMVTVSS

SEQ
Sequence ID Sequence Description NO.
Antibody 418 38 383 GFTFNNYG
CDRH1 (aa) Antibody 418 38 384 IWYDGSNK
CDRH2 (aa) Antibody 418_38 385 ARETSDYGDYIRLRRNAFDI
CDRH3 (aa) EIVMMQ SP ATLSVSPGA RVTLS CRA SQSISNNLA
WYQQKPGQAPRLLIYGASTRASGIPARFSGSGSGT
Antibody 418-38 386 EFTLTISSLQ SEDFAVYYCQQYDKWPPWTFGQG
VL(VK) (aa) TKVEIK
Antibody 418_38 Antibody 418 38 GAS

Antibody 418 38 389 QQYDKWPPWT

CAGGTGCAGCTGGTGGAGTCTGGGGGAGGCGT
GGTCCAGCCTGGGAAGTCCCTGAGACTCTCCTG
TGCAGCGTCTGGATTCACCTTTAATAACTATG
GCATGCACTGGGTCCGCCAGGCTCCAGGCAAG
GGGCTGGAGTGGGTGGCAGTTATATGGTAT GA
Antibody 418 38 TGGAAGTAATAAATAC TATA CAGAC TC CGTGA
¨ 390 AGGGCCGATTCAC CATCTCCAGAGACAATTC CA
VH (nt) AGAACACGCTGTATCTGCAAATGAACAGCCTGA
GAGCCGAGGACACGGCTGTCTATTACTGTGCG
AGAGAGACAAGTGACTACGGTGACTACATAC
GCTTGCGCAGGAATGC T TT TGATATCTGGGG
CCAAGGGACAATGGTCACCGTCTCTICAG
GAAATAGTGATGATGCAGTCTCCAGCCACCCTG
TCTGTGTCTCCAGGGGCGAGAGTCACCCTCTCC
TGCAGGGCCAGTCAGAGTATTAGCAACAACTT
AGCCTGGTACCAGCAGAAACCTGGCCAGGCTCC
Antibody 418 38 CAGGCTCCTCATCTATGGTGCATCCACCAGGG
vL(vi() (nt) 391 CCTCTGGTATCCCAGCCAGGTTCAGTGGCAGTG
GGTCTGGGACAGAGTTCACTCTCACCATCAGCA
GCTTGCAGTCTGAAGATTTTGCAGTTTATTACTG
TCAGCAGTATGATAAGTGGCCTCCGTGGACG
TTCGGCCAAGGGACCAAGGTGGAAATCAAAC

SEQ
Sequence ID Sequence Description NO.
QVQLVQSGAEVKKPGS SVKVSCKASGGTISSYAI
SWVRQAPGQGLEWMGGIMRIFGTPNYAQKFQG
VH ( Antibody 418-39 392 RVTITADESTSTAYMELSSLRSEDTAVYYCAREG
aa) YC SS SNCYDDALDIWGQGTMVTVS S
Antibody 418_39 393 GTIS SYA
CDRH1 (aa) Antibody 418_39 CDRH2 (aa) Antibody 418 39 395 AREGYCSS SNCYDDALDI
CDRH3 (aa) QPVLTQSPSASASLGASVKLTCTLSSGHSSYAIAW
Antibody 418 39 HQ Q QPEKGPRYLMKLNSD GSHSKGDGIPDRFSGS
¨ 396 SSGAERYLTISSLQSEDEADYYCQTWGIGIRVFG
VL (aa) GGTKLTVL
Antibody 418 39 397 SGHS SYA

Antibody 418 39 398 LNSDGSH

Antibody 418_39 399 QTWGIGIRV

CAGGTGCAGCTGGTGCAGTCTGGGGCTGAGGTG
AAGAAGCCTGGGTCCTCGGTGAAGGTCTCCTGC
AAGGCTTCTGGAGGCACCATCAGCAGCTATG
C TATCAGC TGGGTGC GA CAGGC C CCTG GA CAA
GGGCTTGAGTGGATGGGAGGGATCAT GC GTAT
CTTTGGTACACCAAACTACGCACAGAAGTTCC
VH t Antibody 418-39 400 AGGGCAGAGTCACGATTACCGCGGACGAATCC
n) ( ACGAGCACAGCCTACATGGAGCTGAGCAGCCT
GAGATCTGAGGACACGGCCGTGTATTACTGTGC
GAGGGAAGGATATTGTAGTAGTAGTAACTGT
TATGAC GAT GC T T TAGATATC TGGGGCCAAGG
GACAATGGTCACCGTCTCTTCAG
CAGCCTGTGCTGACTCAATCGCCCTCTGCCTCT
GCCTCCCTGGGAGCCTCGGTCAAGCTCACCTGC
ACTCTGAGCAGTGGGCACAGCAGCTACGCCA
Antibody 418 39 VL (nt) CCTCGGTACTTGATGAAGCTTAACAGTGATGG
CAGCCACAGCAAGGGGGACGGGATCCCTGATC
GCTTCTCAGGCTCCAGCTCTGGGGCTGAGCGTT

SEQ
Sequence ID Sequence Description NO.
ACCTCACCATCTCCAGCCTCCAGTCTGAGGATG
AGGCTGACTATTACTGTCAGACGTGGGGCATT
GGCATTCGGGTATTCGGCGGAGGGACCAAACT
GACCGTCCTAG
EVQLVQSGAEVKKPGASVKVSCKVSGYTLPELSI
Antibody 418 41 HWVRQAPGKGLEWMGGFDPEDGETIYAQKFQG
¨ 402 RVTMTEDTSTDTAYMELTSLRSDDTAVYYCATSP
VH (aa) AVVRKNWFDPWGQGTLVTVSS
Antibody 418_41 403 GYTLPELS
CDRH1 (aa) Antibody 418 41 404 FDPEDGET
CDRH2 (aa) Antibody 418_41 405 ATSPAVVRKNWFDP
CDRH3 (aa) SYELTQPP SVSVSPGQTA SITC SGDKL GD KD A CW
Antibody 418 41 YQQKPGQSPVLVIYEDNKRPSGIPERFSGSNSGNT
VL ( ¨ 406 ATLTISGTQAMDEADYYCQAWDSSTHVVFGGGT
aa) KLTVL
Antibody 418 41 407 KLGDKD

Antibody 418 41 EDN

Antibody 418_41 409 QAWDSSTHVV

GAGGTGCAGCTGGTACAGTCTGGGGCTGAGGT
GAAGAAGCCTGGGGCCTCAGTGAAGGTCTCCTG
CAAGGTTTCCGGATACACCCTCCCTGAATTAT
CCATACACTGGGTGCGACAGGCTCCTGGAAAA
GGGCTTGAGTGGATGGGAGGTTTTGATCCTGA
Antibody 418 41 AGA TGGTGAAACAATCTATGCACAGAAGTTCC
¨ 410 AGGGCAGAGTCACCATGACCGAGGACACATCT
VH (nt) ACAGACACAGCCTACATGGAGCTGACCAGCCT
GAGATCTGACGACACGGCCGTCTATTACTGTGC
AACCTCCCCGGCTGTGGTACGAAAGAACTGG
TTCGACCCCTGGGGCCAGGGAACCCTGGTCAC
CGTCTCCTCAG
TCCTATGAGCTGACTCAGCCACCCTCAGTGTCC
Antibody 418 41 VL (nt) TCTGGAGATAAATTGGGGGATAAAGATGCCTG

SEQ
Sequence ID Sequence Description NO.
CTGGTATCAGCAGAAGCCAGGCCAGTCCCCTGT
GCTGGTCATCTATGAAGATAACAAGCGGCCCT
CAGGGATCCCTGAGCGATTCTCTGGCTCCAACT
CTGGGAACACAGCCACTCTGACCATCAGCGGG
ACCCAGGCTATGGATGAGGCTGACTATTACTGT
CAGGCGTGGGACAGCAGCACTCATGTGGTAT
TCGGCGGAGGGACCAAGCTGACCGTCCTAG
QVQLQESGPGLVKPSQTLSLTCTVSGDSISSGDHY
Antibody 418 42 WSWIRQPPGKGLEWIGYIYYSGNTYYNPSLKSRL
¨ 412 TISVDTSNNQFSLKLSSVTAADTAVYYCARAIVG
VH (aa) MVRGVILLWYFDPWGRGTLVTVSS
Antibody 418 42 413 GDSISSGDHY
CDRH1 (aa) Antibody 418_42 414 IYYSGNT
CDRH2 (aa) Antibody 418 42 415 ARAIVGMVRGVILLWYFDP
CDRH3 (aa) QSVLTQPPSVSAAPGQKVTISCSGNRSNIGNNYVS
Antibody 418 42 WYQQFPGTAPKLLIYDINKRPSGIPDRFSGSKSGTS
¨ 416 ATLGITGLQTGDEADYYCGTWDSSLSGPVFGGG
VL (aa) TKL'TVL
Antibody 418 42 417 RSNIGNNY

Antibody 418 42 Antibody 418 42 419 GTWDSSLSGPV

CAGGTGCAGCTACAGGAGTCGGGCCCAGGACT
GGTGAAGCCTTCACAGACCCTGTCCCTCACCTG
CACTGTCTCTGGTGACTCCATCAGCAGTGGTG
ATCACTACTGGAGTTGGATCCGCCAGCCCCCA
GGGAAGGGCCTGGAGTGGATTGGTTACATCTA
TTACAGTGGCAACACCTACTACAACCCGTCCC
Antibody 418-42 420 TCAAGAGTCGACTTACCATATCAGTAGACACGT
VH (nt) CCAATAATCAGTTCTCCCTGAAGCTGAGCTCTG
TGACTGCCGCAGACACGGCCGTGTATTACTGTG
CCAGAGCAATCGTGGGTATGGTTCGGGGAGT
TATTCTTCTCTGGTACTTCGATCCCTGGGGCC
GTGGCACCCTGGTCACTGTCTCCTCAG

SEQ
Sequence ID Sequence Description NO.
CAGTCTGTGTTGACGCAGCCGCCCTCAGTGTCT
GCGGCCCCAGGACAGAAGGTCACCATCTCCTGC
TCTGGAAACAGATCCAACATAGGGAATAATTA
TGTATCCTGGTACCAGCAGTTCC CAGGAACAGC
CCCCAAACTCCTCATTTATGACATTAATAAGCG
Antibody 418 42 421 ACCCTCAGGGATTCCTGACCGATTCTCTGGCTC
VL (nt) CAAGTCTGGCACGTCAGCCACCCTGGGCATCAC
CGGACTCCAGACTGGGGACGAGGCCGATTATTA
CTGCGGAACATGGGATAGCAGCCTGAGTGGT
CCTGTATTCGGCGGAGGGACCAAGCTGACCGT
CCTAG
EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAM
Antibody 418 43 SWVRQAPGKGLEWVSSISGDGGSTYYADSVKGR
VI-I
¨ 422 FTVSRDNSKNTVYLQMNSLRVEDTAVYYCAKGD
a a) ( TFMVPYNWFDPWGQGTLVTVSS
Antibody 418 43 423 GFTFSSYA
CDRHI (aa) Antibody 418 43 424 ISGDGGST
CDRH2 (aa) Antibody 418 43 AKGDTFMVPYNVVFDP

CDRH3 (aa) EIVLTQSPATLSLSPGERATLSCRASQSISSRLAWY
QQKPGQAPRLLIYDASNRATGIPARFSGSGSGTDF
Antibody 418-43 VL(VK) (aa) 426 TLTISGLEPEDFAVYYCQQRSNWPGTFGQGTKVE
IK
Antibody 418_43 427 QSISSR
CDRLI
Antibody 418 43 DAS

Antibody 418 43 429 QQRSNWPGT

GAGGTGCAGCTGTTGGAGTCTGGGGGAGGCTTG
GTACAGCCTGGGGGGTCCCTGAGACTCTCCTGT
GCAGCCTCTGGATTCACCTTTAGCAGCTATGC
Antibody 418 43 3 CATGAGCTGGGTCCGCCAGGCTCCAGGGAAGG

VH (nt) GGCTGGAGTGGGTCTCATCTATTAGTGGTGAT
GGTGGTAGCACATATTACGCAGACTCCGTGAA
GGGCCGGTTCACCGTCTCCAGAGACAATTCCAA
GAACACGGTATATCTGCAAATGAACAGCCTGA

SEQ
Sequence ID Sequence Description NO.
GAGTCGAGGACACGGCCGTATATTACTGTGCG
AAAGGGGATACATTTATGGTTCCGTACAACT
GGTTCGACCCCTGGGGCCAGGGAACCCTGGTC
ACCGTCTCCTCAG
GAAATTGTGCTGACTCAGTCTCCAGCCACCCTG
TCTTTGTCTCCAGGGGAAAGAGCCACCCTCTCC
TGCAGGGCCAGTCAGAGTATTAGCAGCCGCTT
AGCCTGGTACCAGCAGAAACCTGGCCAGGCTCC
Antibody 418 43 CAGGCTCCTCATCTATGATGCATCCAACAGGG
¨ 431 CCACTGGCATCCCAGCCAGGTTCAGTGGCAGTG
VL(VK) (nt) GGTCTGGGACAGACTTCACTCTCACCATCAGCG
GCCTAGAGCCTGAAGATTTTGCTGTTTATTACT
GTCAGCAGCGTAGCAACTGGCCGGGGACGTT
CGGCCAAGGGACCAAGGTGGAAATCAAAC
EVQLQESGPGLVKSSETLSLTCTVSGGSISSDYWN
Antibody 418-44 432 WIRQPPGKGPEWIGYIYYSGSTHYNPSLKSRVTIS
VH ( VDTSKSQFSLKLSSVTAADTAVYYCARLLYYYDS
aa) SGYSIGGAFDIWGQGTMVTVSS
Antibody 418 44 CDRH1 (aa) Antibody 418 44 IYYSGST

CDRH2 (aa) Antibody 418 44 435 ARLLYYYDSSGYSIGGAFDI
CDRH3 (aa) SYELTQPPSVSVSPGQTARITCSGDALAKQYAYW
Antibody 418 44 YQQKPGQAPVLVIYKDTERPSGIPERFSGSSSGTT
VL ( ¨ 436 VTLTISGVQ AEDEADYYCQSADSSSTYVVFGGGT
aa ) RLTVL
Antibody 418 44 437 ALAKQY

Antibody 418_44 KDT

Antibody 418_44 43, QSADSSSTYVV

GAGGTGCAGCTGCAAGAGTCGGGCCCAGGACT
GGTGAAGTCTTCGGAGACCCTGTCCCTCACTTG
Antibody 418 44 VH (nt) ACTGGAATTGGATTCGGCAGCCCCCAGGGAAG
GGACCGGAGTGGATTGGGTATAT C TAT TACA G

SEQ
Sequence ID Sequence Description NO.
TGGGAGCACCCACTACAACCCCTCCCTCAAGA
GTCGAGTCACCATATCAGTAGACACGTCCAAGA
GCCAGTTCTCCCTAAAGCTGAGCTCTGTGACCG
CTGCGGACACGGCCGTCTATTACTGTGCGAGG
CTTTTATATTACTATGATAGTAGTGGTTATTC
CATAGGAGGTGCTTTTGATATCTGGGGCCAA
GGGACAATGGTCACCGTCTCTTCAG
TCCTATGAGCTGACACAGCCACCCTCGGTGTCA
GTGTCCCCAGGACAGACGGCCAGGATCACCTGC
TCTGGAGATGCATTGGCAAAGCAATATGCTTA
TIGGTACCAACAGAAGCCAGGCCAGGCCCCIGT
Antibody 418 44 GCTGGTGATATATAAAGACACTGAGAGGCCCT

VL (nt) CAGGGACAACAGTCACGTTGACCATCAGTGGA
GTCCAGGCAGAAGACGAGGCTGACTATTACTGT
CAATCAGCAGACAGCAGTTCTACTTATGTGG
TATTCGGCGGAGGGACCAGGCTGACCGTCCTAG
QVQLVQSGAEVKKPGASVKVSCKASGYPFTSYGI
SARS-CoV-2 SWVRQAPGQGLEWMGWISTYNGNTNYAQKFQG
S309 mAb VU 442 RVTMTTDTSTTTGYMELRRLRSDDTAVYYCARD
(aa) YTRGAWFGESLIGGEDNWGQGTLVTVSS
SARS-CoV-2 S309 mAb 443 GYPFTSYG
CDRH1 (aa) SARS-CoV-2 S309 mAb 444 ISTYNGNT
CDRH2 (aa) SARS-CoV-2 S309 mAb 445 ARDYTRGAWFGESLIGGFDN
CDRH3 (aa) EIVLTQSPGTLSLSPGERATLSCRASQTVSSTSLAW
SARS-CoV-2 YQQKPGQAPRLLIYGASSRATGIPDRFSGSGSGTD
S309 mAb VL 446 FTLTISRLEPEDFAVYYCQQHDTSLTFGGGTKVEI
(VK) (aa) SARS-CoV-2 S309 mAb CDRL1 447 QTVSSTS
(aa) SARS-CoV-2 S309 mAb CDRL2 448 GAS
(aa) SEQ
Sequence ID Sequence Description NO.
SARS-CoV-2 S309 mAb CDRL3 449 QQHDTSLT
(aa) QVQLVQSGPEVKKPGTSVRVSCKASGFTFTSSAV
QWVRQARGQRLEWVGWIVVGSGNTNYAQKFHE
SARS-CoV-2 RVTITRDMSTSTAYMELSSLRSEDTAVYYCASPY
S2E12 mAb VH 450 CSGGSCSDGFDIWGQGTMVTVSS
(aa) SARS-CoV-2 S2E12 mAb 451 GFTFTSSA
CDRH1 (aa) SARS-CoV-2 S2E12 mAb 452 IVVGSGNT
CDRH2 (aa) SARS-CoV-2 S2E12 mAb 453 ASPYCSGGSCSDGFDI
CDRH3 (aa) DIVLTQTPGTLSLSPGERATLSCRASQSVSSSYLA
WYQQKPGQAPRLLIYGASSRATGIPDRFSGSGSGT
SARS-CoV-2 DFTLTISRLEPEDFAVYYCQQYVGLTGWTFGQG
S2E12 mAb 454 TKVEIK
VL(VK) (aa) SARS-CoV-2 S2E12 mAb 455 QSVSSSY
CDRL I (aa) SARS-CoV-2 S2E12 mAb 456 GAS
CDRL2 (aa) SARS-CoV-2 S2E12 mAb 457 QQYVGLTGWT
CDRL3 (aa) EVQLVQSGAEVKKPGASVKVSCKASGYTFTGYY
MHWVRQAPGQGLEWMGWINPISSGTSYAQTFQ
SARS-CoV-2 GRVTMTSDTSITTAYMELSRLRSDDTAVYYCARA
S2M11 m Ab VH 458 APFYDFWSGYSYFDYWGQGTLVTVSS
(aa) SEQ
Sequence ID Sequence Description NO.
SARS-CoV-2 S2M11 mAb 459 GYTFTGYY
CDRH1 (aa) SARS-CoV-2 S2M11 mAb 460 INPISSGT
CDRH2 (aa) SARS-CoV-2 ARAAPFYDFWSGYSYFDY
S2M11 mAb 461 CDRH3 (aa) EIVMMQSPGTLSLSPGERATLSCRASQSVSSSYLA
WYQQKPGQAPRLLIYGASSRATGIPDRFSGSGSGT
SARS-CoV-2 DFTLTISRLEPEDFAVYYCQQYGSSAWTFGQGTK
S2M11 mAb 462 VEIK
VL(VK) (aa) SARS-CoV-2 S2M11 mAb 463 QSVSSSY
CDRL1 (aa) SARS-CoV-2 S2M11 mAb 464 GAS
CDRL2 (aa) SARS-CoV-2 QQYGSSAWT
S2M11 mAb 465 CDRL3 (aa) QVQLVQSGAEVKKPGASVKVSCKASGYPFTSYGI
SARS-CoV-2 SWVRQAPGQGLEWMGWISTYQGNTNYAQKFQG
S309 N55Q mAb 466 RVTMTTDTSTTTGYMELRRLRSDDTAVYYCARD
VH (aa) YTRGAWFGESLIGGFDNWGQGTLVTVSS
SARS-CoV-2 S309 N55Q mAb 467 ISTYQGNT
CDRH2 (aa) EXAMPLES

RECOMBINANT EXPRESSION OF CERTAIN ANTIBODIES
Antibodies were recombinantly expressed in ExpiCHO cells transiently co-transfected with plasmids expressing the heavy and light chains as previously described (Stettler et at. (2016)). Specificity, cross-reactivity, and function of antibodies elicited by Zika virus infection. Science, 353(6301), 823-826). The concentration of antibody in cell culture supernatant was measured for antibodies as shown in Table 2.
Table 2.
Monoclonal Antibody Expression in CHO cells (mg/m1) Antibody 418 1 0.078 Antibody 418 2 0.722 Antibody 418 3 0.541 Antibody 418 4 0.366 Antibody 418j4 0.14 Antibody 418 42 0.17 Antibody 418 43 0.20 CHARACTERIZATION OF CERTAIN ANTIBODIES
Certain antibodies of the present disclosure were characterized by identification of the germline VH and VL genes, their EC50 and KD for binding to SARS-CoV-2 Domain A, and whether they exhibit neutralizing activity against SARS-CoV-2.
The results are shown in Table 3. The notation "nn" indicates that the antibody was not neutralizing by this assay. Blank cells in the table indicate that no measurement was made.
Table 3.
mAb IgVH IgVL DomA KD N T
Maximal gene gene ELISA Domain A (IC50 (1/0 neutr.
(EC50 (M) ng/ml) ng/ml) 418_7 1-69 K1-9 1468 <1E-12 119.3 95%
418 13 2-70 K2-28 148.7 <1E-12 nn nn 418_14 3-21 1(3-15 8.943 <1E-12 nn nn 418 39 1-69 L4-69 507.4 <1E-12 nn nn 418_40 3-30 K1-33 447.7 <1E-12 2982 98%
418 8 1-24 K1-27 602.3 <1E-12 nn nn 418_41 1-24 L3-10 47.72 <1E-12 1941.
77%
418 9 3-21 L2-14 34.26 7.10E-11 76.38 77%
418_42 4-30 L1-51 22.1 <1E-12 nn nn 418 43 3-23 K3-11 26.06 <1E-12 nn nn 418 44 4-59 L3-25 63.72 <1E-12 24.88 91%
418_6 4-61 K1-39 698.7 <1E-12 nn nn 418_5 3-33 L3-25 31.66 <1E-12 53.73 88%
418_22 1-24 L1-47 12.3 nn nn 41S1 3-30 L3-10 337.6 5.52E-10 90 91%
418 23 2-26 L1-40 18.5 nn nn 418_24 3-21 K3-15 14.2 nn nn 418_25 3-21 K3-15 19.4 nn nn 418 37 4-4 L2-23 11.5 nn nn 418 26 1-24 K2-24 60.5 40.3 87%
418_38 3-21 K3-15 13.4 nn nn mAb IgVH IgVL DomA KD N T
Maximal gene gene ELISA Domain A (IC50 % neutr.
(EC50 (M) ng/ml) ng/ml) 418_11 3-3 K3-15 8.7 nn nn 418 27 3-53 K3-15 20 nn nn 418 21 4-59 K3-20 24.3 nn nn 418 20 4-59 K3-15 12.4 nn nn 418_19 4-38 K1-39 69 7.9 65%
418_28 3-30 1(1-13 30 8.9 68%
418 18 3-21 L1-40 24 nn nn 418_29 1-24 L1-47 18.7 32.2 96%
418 30 1-46 L3-25 16.4 17 83%
418_31 4-61 L10-54 51.8 0.67 98%
418 12 3-23 L1-51 10.9 nn nn 418_17 3-21 1(3-15 26.7 nn nn 418 33 4-59 L3-21 52.8 nn nn 418 16 3-21 K3-15 16.8 nn nn 418_34 2-70 1(4-1 474.6 nn nn 418_15 1-2 L2-23 18.3 nn nn 418 2 2-5 L3-1 14.2 <1E-12 79.1 85%
418_10 4-34 L2-23 8 nn nn 418 35 3-48 K2D-29 97.8 nn nn 418_3 3-33 1(1-33 26.9 <1E-12 147.9 42%
418 4 3-33 L3-21 7.6 2.75E-11 43.3 94%
Additional characterization of monoclonal antibodies 418_i, 418 2, 418 3, and 4i8_4 was performed. The kon, kdis, neutralization activity against SARS-CoV-2 with or without tosyl phenylalanyl chloromethyl ketone (TPCK), ability to block SARS-CoV-2 binding to ACE2, ability to induce antibody-dependent cellular phagocytosis (ADCP) (i.e., FcyRIIa activation), ability to induce antibody-dependent cell-mediated cytotoxicity (ADCC) (i.e., Fc7RIIIa activation), and measurement of antibody-mediated shedding of SARS-CoV-2 Si protein from infected cells for each of these antibodies is shown in Table 4.
Table 4.

mAb 418_i 418_2 418_3 418_4 kon (1/Ms) 2.4E+05 4.5E+05 1.7E+05 2.4E+05 kdis (1 /s) 1.34E-04 <1.0E-07 <1.0E-7 6.67E-06 Neutr IgG + TPCK (IC50, 90.0 79.1 147.1 43.3 ng/ml) Neutr IgG ¨ TPCK (IC50, 139.4 92.2 97.1 239.6 ng/ml) ACE2 blockade No No No No ADCP (FcyRIIa activation) + +++
ADCC (FcyRIIIa activation) +++
Si shedding no Additional characterization was carried out for six antibodies, as shown in Table 5. EC50 values were measured by ELISA for binding to SARS-CoV-2 Spike protein Domain A. KD, km, and kdis values were measured by BLI for binding to SARS-CoV-Spike protein Domain A.
Table 5.
mAb EC50 (ng/ml) KD (M) k.. (1/Ms) kdis (Vs) 418 37 11.5 3.35E-09 1.80E+05 6.05E-mAb EC50 (ng/ml) KD (M) k0 (1/Ms) kdis (Vs) 481A 8.7 3.73E-08 2.14E+04 7.97E-418 20 12.4 1.39E-08 5.02E+04 6.96E-418 22 12.3 1.87E-09 1.72E+05 3.22E-418 12 10.9 1.49E-08 3.77E+04 5.63E-418 10 8 5.58E-09 9.58E+04 5.35E-FURTHER STUDIES USING NTD-SPECIFIC ANTIBODIES
Introduction The emergence of SARS-CoV-2 coronavirus at the end of 2019 resulted in the ongoing COVID-19 pandemic. The lack of pre-existing immunity to SARS-CoV-2 combined with its efficient human-to-human transmission has already resulted in more than 86 million infections and over 1.85 million fatalities as of January 2021.
Prophylactic and/or therapeutic anti-viral drugs may be helpful for unvaccinated individuals or those who respond poorly to vaccination as well as upon waning of immunity or emergence of antigenically distinct strains.
SARS-Coli-2 infects host cells through attachment of the viral transinembrane spike (S) glycoprotein to angiotensin-converting enzyme 2 (ACE) followed by fusion of the viral and host membranes (Letko et al., 2020; Walls et al., 2020c;
Wrapp et al., 2020; Zhou et al., 2020). SARS-CoV-2 S also engages cell-surface heparan-sulfates (Clausen et al., 2020), neuropilin-1 (Cantuti-Castelvetri et al., 2020; Daly et al., 2020) and L-SIGN/DC-SIGN (Chiodo et al., 2020; Gao et al., 2020; Soh et al., 2020;
Thepaut et al., 2020) which were proposed to serve as co-receptors, auxiliary receptors, or adsorption factors. SARS-CoV-2 S is the main target of neutralizing Abs in infected individuals and the focus of the many nucleic acid, vectored, and protein subunit vaccines currently deployed or in development (Corbett et al., 2020a; Corbett et al., 2020b, Erasmus et al., 2020; Hassan et al., 2020; Keech et al., 2020; Mercado et al., 2020; Walls et al., 2020b). Besides blocking ACE2 attachment (Piccoli et al., 2020;
Tortorici et al., 2020), some neutralizing Abs may interfere with heparan-sulfate, neuropilin-1 or L-SIGN/DC-SIGN interactions.
The SARS-CoV-2 S protein comprises an N-terminal Si subunit responsible for virus¨receptor binding, and a C-terminal Sz subunit that promotes virus¨cell membrane fusion (Walls et al., 2020c; Wrapp et al., 2020). The Si subunit comprises an N-terminal domain (NTD) and a receptor-binding domain (RBD), also known as domain A and B, respectively (Tortorici and Veesler, 2019) Antibodies targeting the RBD
account for 90% of the neutralizing activity in COVID-19 convalescent sera (Piccoli et al., 2020) and numerous monoclonal antibodies (mAbs) recognizing this domain have been isolated and characterized (Barnes et al., 2020a; Barnes et al., 2020b, Baum et al., 2020b; Brouwer et al., 2020; Hansen et al., 2020; Ju et al., 2020; Piccoli et al., 2020;
Pinto et al., 2020; Tortorici et al., 2020; Wang et al., 2020; Wu et al., 2020). Several RBD-specific mAbs capable of protecting small animals and non-human primates from SARS-CoV-2 challenge are able to neutralize viral infection by targeting multiple distinct antigenic sites (Baum et al., 2020a; Hansen et al., 2020; Jones et al., 2020; Pinto et al., 2020; Rogers et al., 2020; Tortorici et al., 2020; Zost et al., 2020).
A subset of these mAbs is currently being evaluated in clinical trials or have recently received emergency use authorization from the FDA
The apparent limited immunogenicity of the SARS-CoV-2 NTD in COVID-19 patients (Piccoli et al., 2020; Rogers et al., 2020) has been hypothesized to result from its N-linked glycan shielding (Walls et al., 2020c; Watanabe et al., 2020).
However, some studies have reported on the isolation of NTD-targeted mAbs and their ability to neutralize SARS-CoV-2 infection in vitro suggesting they could be useful for COVID-19 prophylaxis or treatment (Chi et al., 2020; Liu et al., 2020a). Although the NTD has been proposed to interact with auxiliary receptors in cell types that do not express ACE2 (e.g. DC-SIGN/L-SIGN), its role and the mechanism of action of NTD
targeted neutralizing mAbs remain unknown (Soh et al., 2020). Understanding the immunogenicity of different S domains and the function of mAbs targeting them, including the NTD, is important to understanding immunity during the pandemic.
Ab responses in three COVID-19 convalescent individuals were analyzed and 41 NTD-specific human mAbs were identified. Integrating cryo-electron microscopy (cryoEM), binding assays, and antibody escape mutants analysis a SARS-CoV-2 NTD
antigenic map was defined, and a supersite recognized by potent neutralizing mAbs was identified. These mAbs exhibit neutralization activities on par with potent RBD-specific mAbs and efficiently activate Fc-mediated effector functions. Immunologically important variations of the SARS-CoV-2 NTD were also identified, suggesting that the S glycoprotein is under selective pressure from the host humoral immune response. A
highly potent NTD mAb was shown to provide prophylactic protection against lethal SARS-CoV-2 challenge of Syrian hamsters.
NTD-specific mAbs with potent neutralizing activity To discover mAbs targeting diverse SARS-CoV-2 epitopes, IgG- memory B
cells from peripheral blood mononuclear cells (PBMCs) of three COVID-19 convalescent individuals (L, M, X) were sorted using biotinylated prefusion SARS-CoV-2 S as a bait. The percentage of SARS-CoV-2 S-reactive IgG B cells ranged between Ll - L3 % of IgG+ memory B cells. A total of 278 mAbs were isolated and recombinantly produced as human IgG I (Figure 20). Characterization by ELISA
showed that most mAbs isolated from the three donors recognize the RBD (65-77%), with a smaller fraction targeting the NTD (6-20%). The remaining mAbs (4-20%) are expected to bind to either the Sz subunit or the C-D domains within the Si subunit (Figure 20). The low proportion of NTD-specific mAbs isolated from these donors is in line with the previously observed limited NTD immunogenicity in SARS-CoV-2 exposed individuals (Piccoli et al., 2020; Rogers et al., 2020). Overall, 41 mAbs recognizing the SARS-CoV2 NTD were identified, with EC50s ranging between 7.6 -698 ng/ml and nanomolar binding affinities, as evaluated using ELISA and biolayer interferometry, respectively (Figures 21, 24A-24D, and 28A-28F, and Tables 6 and 7). These NTD-specific mAbs use a large repertoire of V genes, with an over-representation of IGHV3-21 and IGK3-15 genes (Figure 25 and Tables 6 and 7).
These mAbs harbor few somatic hypermutations (VH and VL are 97.57% and 97.54%
identical to V germline genes, respectively; (Figure 26, Tables 6 and 7), as previously described for most SARS-CoV-2 neutralizing mAbs binding to the RBD (Piccoli et al., 2020; Seydoux et al., 2020). Antibody 418_i is also referred to herein as S2X28.
Antibody 418 2 is also referred to herein as S2X303. Antibody 418 3 is also referred to herein as S2X320. Antibody 418 4 is also referred to herein as S2X333.
Antibody 418 5 is also referred to herein as S2M28. Antibody 418_6 is also referred to herein as S2M24 or S2M24v2. Antibody 418_7 is also referred to herein as S2L7. Antibody 418 8 is also referred to herein as S2L24. Antibody 4i89 is also referred to herein as S2L28. Antibody 418 10 is also referred to herein as S2X310. Antibody 418 11 is also referred to herein as S2X94. Antibody 418_12 is also referred to herein as S2X169. Antibody 418 13 is also referred to herein as S2L11. Antibody 418 14 is also referred to herein as S2L12. Antibody 418 15 is also referred to herein as S2X186. Antibody 418_16 is also referred to herein as S2X175. Antibody 418 17 is also referred to herein as S2X170. Antibody 418_18 is also referred to herein as S2X125. Antibody 418 19 is also referred to herein as S2X107. Antibody 418 20 is also referred to herein as S2X105. Antibody 418 21 is also referred to herein as S2X102. Antibody 418_22 is also referred to herein as S2X15. Antibody 418_23 is also referred to herein as S2X49. Antibody 418_24 is also referred to herein as S2X51.
Antibody 418 25 is also referred to herein as S2X72. Antibody 418_26 is also referred to herein as S2X91. Antibody 418 27 is also referred to herein as S2X98.
Antibody 418 28 is also referred to herein as S2X124. Antibody 418_29 is also referred to herein as S2X158. Antibody 418 30 is also referred to herein as S2X161. Antibody 418 is also referred to herein as S2X165. Antibody 418_33 is also referred to herein as S2X173. Antibody 418 34 is also referred to herein as S2X176. Antibody 418_35 is also referred to herein as S2X316. Antibody 418 37 is also referred to herein as S2X90. Antibody 418_38 is also referred to herein as S2X93. Antibody 418 39 is also referred to herein as S2L14. Antibody 418_40 is also referred to herein as S2L20 or S2L20v1. Antibody 418 41 is also referred to herein as S2L26. Antibody 4i8_42 is also referred to herein as S2L35. Antibody 418 43 is also referred to herein as S2L38.
Antibody 418 44 is also referred to herein as S2L50.

CDRI-13 lengths of these mAbs range between 10 and 24 amino acid residues (Figure 26). Collectively, these data indicate that the Ab response to the SARS-CoV-2 NTD is polyclonal.
Table 6.
NT IgG vs IgVH HCDR3 IgVL ELISA vs NTD Antigenic ND
Mx %
# Donor mAb VH % G L VL ./0 G L
MLV-S2 pp NT
gene length gene (EC50 ng/ml) site NTD (M) (IC50 ng/m1) 1 S2L11 2-70 21 98.63 K2-28 100 148.7 ii 7.25E-09 MI nn 2 S2L 12 3-21 19 98.26 K3-15 97.49 8.943 0.1 na nn MI
3 S2L14 1-69 18 98.61 L4-69 98.64 507.4 iii 1.18E-08 66 nn 4 S2L20 3-30 15 97.92 K1-33 96.42 447.7 iv 3.05E-08 2982 98%
S2L24 1-24 14 98.96 K1-27 99.28 602.3 i 1.17E-08 49.1 83.50%
L
is 182L26 1-24 14 97.22 L7-10 9/1.21 47.72 i 4L56E-119 19.41 77%
7 S2L28 3-21 19 96.53 L2-14 97.57 34.26 i 1.20E-07 76.38 77%
8 S2L35 4-30 21 96.9 Li-Si 98.25 22.1 iii 7.63E-09 66 nn 9 S2L38 3-23 17 97.22 K3-11 97.13 26.06 iii 1.62E-09 nn on S2L50 4-59 20 95.44 L3-25 98.92 63.72 i 5.96E-09 24.88 91%
11 S2M24 4-61 20 97.25 K1-39 95.34 698.7 vi 6.98E-09 no no M

12 S2M28 3-33 12 97.57 L3-25 97.85 31.66 i 6.89E-09 53.73 88%

13 S2X15 1-3 21 98.98 K3-11 96.14 12.3 iii 1.87E-09 nn nn

14 S2X28 3-30 18 97.92 L3-10 99.64 337.6 i na 90 91%
S2X19 2-26 11 98.66 Li-10 98.98 18.5 iii 1.97E-09 no no 16 S2X51 3-21 20 98.95 K3-15 98.29 14.2 iii 3.24E-08 no no 17 S2X72 3-21 21 97.21 K3-15 98.93 19.4 iii 2.19E-08 iiii no 18 S2X90 4-4 13 97.61 L2-23 96_94 11.5 iii 3_35E-09 no on 19 S2X9 I 1-24 21 97.28 K2-24 97.64 60.5 i 3.39E-09 40.3 87%
S2X93 3-33 20 97.97 K3-15 96.51 13.4 iii 6.71E-08 66 no 21 S2X94 3-53 14 97.6 K3-15 98.59 8.7 iii 3.73E-08 nn no X
22 S2X98 3-21 24 97.61 K3-15 99.65 20 iii 6.14E-08 MI no 23 S2X102 4-59 15 97.81 K3-20 96.81 24.3 0.1 8.23E-09 nn MI
24 S2X105 4-59 17 98.29 K3-15 99.3 12.4 iii 1.39E-08 66 no S2X107 4-38 16 96.95 K1-39 96.83 69 i 4.08E-08 7.9 65%
26 S2X124 3-30 22 98.98 K1-13 98.25 30 i 4.70E-08 8.9 68%
27 S2X125 3-21 17 95.6 L1-40 97.57 24 111 3.39E-09 MI no 28 S2X158 1-24 16 96.25 L1-47 95.91 18.7 i 8.54E-09 32.2 96%
29 S2X161 1-46 21 95.46 L3-25 99.64 16.4 i 8.42E-09 17 83%
S2X165 4-61 20 96.65 L10-54 96.59 51.8 i 3.46E-08 61.07 98%
_ 31 S2X169 3-23 17 97.96 L1-51 98.64 10.9 iii 1.49E-08 nn nn 32 S2X170 3-21 20 96.96 K3-15 97.9 26.7 iii 1.62E-08 nn nn 33 S2X173 4-59 15 97.95 L3-21 97.21 52.8 v 1.22E-08 nn nn 34 S2X175 3-21 20 97.72 K3-15 96.167 16.8 Ill 8.94E-09 nn nn 35 S2X176 2-70 10 98.65 K4-1 98.01 474.6 ii 1.08E-08 nn nn 36 S2X186 1-2 12 98.97 L2-23 98.62 18.3 iii 9.58E-09 nn nn 37 S2X303 2-5 17 95.88 L3-1 95.34 14.2 1 6.30E-09 79.1 85"/o 38 S2X310 4-34 20 97.14 1,2-23 96.53 8 iii 5.5812-09 nn nn 39 S2X316 3-48 18 99.66 K2-29 94.73 97.8 v 1.77E-08 nn nn 40 S2X320 3-33 17 96.53 K1-33 97.85 26.9 i 1.81E-08 147.9 42%
41 S2X333 3-33 17 96.53 L3-21 97.49 7.6 i 2.89E-08 43.3 94%
-Table 7.
NT IgG vs NT Fab vs Mx %
# Donor mAb live virus live virus Mx % NT
NT
IC50 (ng/m1) IC50 (Hg/ml) 7 S2L28 26.2 98.20% 928 55.60%

NT IgG vs NT Fab vs Mx %
# Donor mAb live virus live virus Mx % NT
NT
IC50 (ng/ml) IC50 (ng/ml) 12 S2M28 5 98.50% 26.3 66%

14 S2X28 9.1 99.10% 248.9 87.30%

NT IgG vs NT Fab vs Mx %
# Donor mAb live virus live virus Mx % NT
NT
IC50 (ng/ml) IC50 (ng/ml) 41 S2X333 3 98.70% 6.1 82.20%

In vitro neutralization activity of the NTD-specific mAbs was evaluated using a SARS-CoV-2 S pseudotyped murine leukemia virus system (Millet and Whittaker, 2016; Walls et al., 2020c). Out of 41 mAbs, 9 are potent neutralizers (IC0 <
50 ng/mL) and 6 are moderate neutralizers (IC50 of 50-150 ng/mL) (Figure 21). The remaining 25 mAbs were non-neutralizing. Most of the mAbs plateaued around 80-90% maximum neutralization in this assay (Figures 21 and 29A-29F). Evaluation of the neutralization potency of a subset of NTD-specific mAbs measured 6 hours post-infection of Vero E6 cells infected with authentic SARS-CoV-2 virus confirmed that these mAbs did not completely block viral entry and instead plateaued at 80-90% neutralization, as opposed to the RBD-specific mAbs S309, S2E12 and S2M11 that achieved 100%
neutralization (Figure 22) (Pinto et al., 2020; Tortorici et al., 2020). When the activity was measured at 24 hours post-infection, however, all mAbs tested achieved 95-100%
neutralization with a marked enhancement of neutralization potency (Figure 23). For instance, S2X333 neutralized SARS-CoV-2 with an IC50 of 2 ng/ml and an IC90 of 12 ng/ml, on par with the potent RBD-targeting mAbs S2E12 and S2M11 (Figure 23).
Previous studies established that SARS-CoV-2 infection of Vero E6 cells proceeds through cathepsin-activated endosomal fusion, as opposed to TMPRSS2-dependent entry which is thought to occur at the level of the plasma membrane and to be the most relevant route of lung cells infection (Hoffmann et al., 2020a;
Hoffmann et al., 2020b; Hoffmann et al., 2020c) Antibodies S2L28, S2M28, S2X28 and S2X333 efficiently block cell-cell fusion (Figure 27) Definition of a SARS-CoV-2 NTD antigenic map Competition biolayer interferometry binding assays were carried out using recombinant SARS-CoV-2 S The data indicated that the mAbs recognize six distinct antigenic sites, designated i, ii, iii, iv, v and vi. Most mAbs clustered within antigenic sites i and iii, whereas sites ii, iv, v and vi each accounted for only one or a small number of mAbs from the panel (Figures 30 and 31A-31I). All potently neutralizing mAbs tested competed for binding to the NTD site i (Figures 21, 22, and 30) Mechanism of action of NTD-specific neutralizing mAbs The ability of these mAbs to block ACE2 binding was evaluated, as this step correlates with neutralization titers in SARS-CoV-2 exposed individuals (Piccoli et al., 2020). None of the site i-targeting mAbs (S2L28, S2M28, S2X28, and S2X333) blocked binding of SARS-CoV-2 S to immobilized human recombinant ACE2 as measured by biolayer interferometry (Figure 32), indicating that interference with engagement of the main entry receptor is unlikely as the mechanism of action.
Moreover, these mAbs did not promote shedding of the Si subunit from cell-surface-expressed full-length SARS-CoV-2 S (Figure 8C), suggesting that premature S
triggering does not occur, unlike what was previously shown for a SARS-CoV and several SARS-CoV-2 RBD-specific mAbs (Huo et al., 2020; Piccoli et al., 2020;
Walls et al., 2019; Wec et al., 2020; Wrobel et al., 2020a).
The neutralization potency of each of 5L28, 52M28, 52X28, and 52X333 was evaluated, in both Fab and IgG formats, against authentic SARS-CoV-2-Nlue (Figure 33). NTD-specific Fabs displayed a potency reduction, both in terms of IC50 values and maximal neutralization plateau reached (Tables 6 and 7), as compared to IgGs, possibily due to reduced avidity as observed by surface plasmon resonance (Figure 34).
Since the Fabs could still partially neutralize SARS-CoV-2, at least part of the observed neutralization activity may result from direct interaction with their respective epitopes It is possible that NTD-specific mAb-mediated neutralization further relies on steric hindrance provided by Fc positioning, similar to what was observed for anti-hcmagglutinin influenza A virus neutralizing mAbs (Xiong et al., 2015).
Potential additive, antagonistic or synergistic effects of NTD- and RBD-targeting mAbs was examined, as mAb synergy was previously described for SARS-CoV and SARS-CoV-2 neutralization (Pinto et al., 2020; ter Meulen et al., 2006).
Cocktails of S2X333 with S309, S2EI2, or S2M11 additively prevented entry of SARS-CoV-2 S-MLV pseudotyped virus in Vero E6 cells (Figures 9A-9C). This additive effect was also observed between 52X333 and S309 using authentic SARS-CoV-2 at hours post-infection in Vero E6 cells (Figure 36). These results are consistent with RBD- and NTD-targeting mAbs mediating inhibition by distinct mechanisms and demonstrate that they could be used as cocktails for prophylaxis or therapy.
Since Fc-mediated effector functions can contribute to protection by promoting viral clearance and anti-viral immune responses in vivo (Bournazos et al., 2020;
Bournazos et al., 2016; Schafer et al., 2021; Winkler et al., 2020), the ability of site i-targeting mAbs to trigger activation of FcyRIIa and FcyRIIIa was evaluated as a proxy for Ab-dependent cellular phagocytosis (ADCP) and Ab-dependent cellular cytotoxicity (ADCC), respectively. S2L28, S2M28, S2X28, and S2X333 promoted dose-dependent FcyRIIa and FcyRIIIa-mediated signaling to levels comparable to those of the highly effective mAb S309 (Pinto et al., 2020) (Figure 35). In contrast, the non-neutralizing site vi-targeting S2M24 mAb did not promote FcyR-mediated signaling, possibly due to the different orientation relative to the membrane of the effector cells in comparison to site i-specific mAbs (Figure 35). These findings suggest that besides their neutralizing activity, mAbs recognizing site i can exert a protective activity via promoting Fc-mediated effector functions.
NTD neutralizing mAbs protect against SARS-CoV-2 challenge in hamsters The S2X333 mAb was selected for a prophylactic study in a Syrian hamster model (Boudewijns et al., 2020). The mAb was administered at 4 and 1 mg/kg via intraperitoneal injection 48 hours before intranasal SARS-CoV-2 challenge.
Four days later, lungs were collected for the quantification of viral RNA and infectious virus titers. Prophylactic administration of S2X333 decreased the amount of viral RNA
detected in the lungs by ¨3 orders of magnitude, compared to hamsters receiving a control mAb (Figure 37A) and completely abrogated viral replication in the lungs of most animals at both doses tested (Figure 37B). Although all animals had similar scrum mAb concentrations within each group, no reduction in the amount of viral RNA
or infectious virus was observed for one hamster at each dose compared to those administered with a control mAb (Figures 37C-37D). Based on the aforementioned variability and mutation tolerance of the SARS-CoV-2 NTD, it may be that escape mutants were selected in these animals. Overall, these data suggest that low doses of anti-NTD mAbs provide prophylactic activity in vivo, comparable to RBD-specific mAbs S2E12 and S2M11 (Tortorici et al., 2020), consistent with their potent in vitro neutralizing activity. The protection efficacy of S2X333 (and related NTD mAbs) may be further enhanced in humans by engineering to enhance interactions with with human Fey receptors.
Discussion The data herein suggest that neutralizing NTD-targeting mAbs represent one aspect of immunity to SARS-CoV-2 and account for 5-20% of SARS-CoV-2 S-specific mAbs cloned from memory B cells isolated from the PBMCs of three COVID-19 individuals. Analysis of a large panel of neutralizing and non-neutralizing mAbs defined an antigenic map of the heavily glycosylated SARS-CoV-2 NTD, in which antigenic sites (i-vi) were identified. All the neutralizing mAbs from the three donors investigated targeted the same antigenic supersite (site i). The neutralizing mAbs described here along with the mAbs 4A8 (Chi et al., 2020), FC05 (Zhang et al., 2020) and CM25 (Voss et al., 2020), which also target this antigenic supersite, use various germline V genes to recognize overlapping epitopes, thereby providing examples of convergent solutions to NTD-targeted mAb neutralization. A highly potent NTD
mAb provides prophylactic protection against SARS-CoV-2 challenge of Syrian hamsters demonstrating that this class of mAbs can be a critical barrier to infection.
These data show that site i-targeting NTD neutralizing mAbs efficiently activate FcyRIIa and FcyRIIIa in vitro. Fc-mediated effector functions can be affected by the epitope specificity of the mAbs (Piccoli et al., 2020), highlighting the importance of the orientation of the S-bound Fe fragments for efficient FeyR cross-linking and engagement. The site vi-targeting NTD mAb S2M24 did not activate either FcyRIIa or FcyRIIIa. The contribution of Fe-mediated effector functions could further enhance the prophylactic activity of potent NTD-specific mAbs against SARS-CoV-2 in humans.
As several examples of single amino acid mutations reducing or completely abrogating neutralization by immune sera have been reported (Li et al., 2020;
Liu et al., 2020b; Weisblum et al., 2020), combinations of mAbs targeting distinct domains may reduce the likelihood of emergence of escape mutants.
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MATERIALS AND METHODS
Affinity determination using Octet (BLI, biolayer interferometry) For KD determination of full-length antibodies, protein A biosensors (Pall ForteBio) were used to immobilize recombinant antibodies at 2.7 ug/m1 for 1 minute, after a hydration step for 10 minutes with Kinetics Buffer (KB). Association curves were recorded for 5min by incubating the antibody-coated sensors with SARS-CoV-Domain A analyte at 10 ug/m1 (66.6 nM) in KB for 5 minutes (association phase), followed by dissociation with KB for 9 minutes. Signals were recorded and analysed with Octet Systems Software.
ELISA binding The reactivities of mAbs with SARS-CoV Spike Si Subunit Protein (strain WH20) protein were determined by enzyme-linked immunosorbent assays (ELISA).
Briefly, 96-well plates were coated with 3 ug/m1 of recombinant SARS-CoV Spike Si Subunit Protein (Sino. Biological). Wells were washed and blocked with PB
S+1%B SA
for 1 h at room temperature and were then incubated with serially diluted mAbs for 1 h at room temperature. Bound mAbs were detected by incubating alkaline phosphatase-conjugated goat anti-human IgG (Southern Biotechnology: 2040-04) for 1 h at room temperature and were developed by 1 mg/ml p-nitrophenylphosphate substrate in 0.1 M
glycine buffer (pH 10.4) for 30 min at room temperature. The optical density (OD) values were measured at a wavelength of 405 nm in an ELISA reader (Powerwave 340/96 spectrophotometer, BioTek).
Pseudoparticle neutralization assay Unless otherwise indicated, Murine leukemia virus (MLV) pseudotyped with SARS-CoV-2 Spike protein (SARS-CoV-2pp) was used. DBT cells stably transfected with ACE2 (DBT-ACE2) were used as target cells. SARS-CoV-2pp was activated with trypsin TPCK at lOug/ml. Activated SARS-CoV-2pp was added to a dilution series of antibodies (starting 5Oug/m1 final concentration per antibody, 3-fold dilution). DBT-ACE2 cells were added to the antibody-virus mixtures and incubated for 48h.
Luminescence was measured after aspirating cell culture supernatant and adding steady-GLO substrate (Promega).
In some cases, pseudoparticle neutralization assays use a VSV-based luciferase reporter pscudotyping system (Kcrafast). VSV pseudoparticles and antibody arc mixed in DMEM and allowed to incubate for 30 minutes at 37C. The infection mixture is then allowed to incubate with Vero E6 cells for lh at 37C, followed by the addition of DMEM with Pen-Strep and 10% FBS (infection mixture is not removed). The cells are incubated at 37C for 18-24 hours. Luciferase is measured using an Ensight Plate Reader (Perkin Elmer) after the addition of Bio-Glo reagent (Promega).
Expression of recombinant antibodies Recombinant antibodies were expressed in ExpiCHO cells transiently co-transfected with plasmids expressing the heavy and light chain as previously described (Stettler et al. (2016) Specificity, cross-reactivity, and function of antibodies elicited by Zika virus infection. Science, 353(6301), 823-826) Authentic SARS-CoV-2 neutralization assay Vero E6 cells cultured in DMEM supplemented with 10% FBS (VWR) and lx Penicillin/Streptomycin (Thermo Fisher Scientific) were seeded in white 96-well plates at 20,000 cells/well and attached overnight. Serial 1:4 dilutions of the monoclonal antibodies were incubated with 200 pfu of SARS-CoV-2 (isolate USA-WA1/2020, passage 3, passaged in Vero E6 cells) for 30 minutes at 37 C in a BSL-3 facility. Cell supernatant was removed and the virus-antibody mixture was added to the cells.

hours post infection, cells were fixed with 4% paraformaldehyde for 30 minutes, followed by two PBS (pH 7.4) washes and permeabilization with 0.25% Triton X-in PBS for 30 minutes. After blocking in 5% milk powder/PBS for 30 minutes, cells were incubated with a primary antibody targeting SARS-CoV-2 nucleocapsid protein (Sino Biological, cat. 40143-R001) at a 1:2000 dilution for 1 hour. After washing and incubation with a secondary Alexa647-labeled antibody mixed with 1 tg/ml Hoechst33342 for 1 hour, plates were imaged on an automated cell-imaging reader (Cytation 5, Biotek) and nucleocapsid-positive cells were counted using the manufacturer's supplied software. Data were processed using Prism software (GraphPad Prism 8.0).
Cell lines Cell lines were obtained from ATCC (FIEK293T and Vero-E6)or ThermoFisher Scientific (Expi CHO cells, FreeStyleTm 293-F cells and Expi293FTM cells).
Sample donors Samples were obtained from three SARS-CoV-2 recovered individuals (L, M
and X) under study protocols approved by the local Institutional Review Boards (Canton Ticino Ethics Committee, Switzerland, the Ethical committee of Luigi Sacco Hospital, Milan, Italy). All donors provided written informed consent for the use of blood and blood components (such as PBMCs, sera or plasma).
Samples were collected 14 and 52 days after symptoms onset for donor L and M, respectively. Blood drawn from donor X was obtained at day 36, 48, 75 and after symptoms onset Cloning and mutant generation SARS-CoV-2 NTD was sub-cloned with E. coli DHIOB Competent Cells into pCMV using primers NTD fwd and NTD rev. The resulting construct was mutated by PCR mutagenesis to generate N149Q, D253G/Y, T19A, R246A, L18F, H146Y, A222V, Y144del, S254F, K147T, C136Y, and the NTD construct with native signal peptide with and without S12P, using the eponymously named primers (Key Resources Table). The genes encoding for the Sarbecovirus S proteins tested were cloned in the phCMV1 or pcDNA.3 vectors, and the gene for the C-terminally his-tagged ectodomain of P-GD S was cloned into pCMV (Key Resources Table). Plasmid sequences were verified by Genewiz sequencing facilities (Brooks Life Sciences).
Recombinant ectodomains production All SARS-CoV-2 S spike ectodomains were produced in 500 mL cultures of FreeStyleTM 293-F cells (ThermoFisher Scientific) grown in suspension using FreeStyle 293 expression medium (ThermoFisher Scientific) at 37 C in a humidified 8% CO2 incubator rotating at 130 r.p.m. Cells grown to a density of 2.5 million cells per mL
were transfected using PEI (9 pg/mL) and pCMV::SARS-CoV-2 S ecto hexapro, pCMV::SARS-CoV-2 S ecto 2P DS, pCMV::P-GD S ecto, pCMV::SARS-CoV-2 S ecto avi, pCMV::SARS-CoV-2 S D614G ecto avi and cultivated for 4 days.
The supernatant was harvested and cells were resuspended for another three days, yielding two harvests. S ectodomains were purified from clarified supernatants using a Cobalt affinity column (Cytiva, HiTrap TALON crude), washing with 20 column volumes of 20 mM Tris-HC1 pH 8.0 and 150 mM NaCl and eluted with a gradient of 600 mM imidazole. The same protocol was followed for P-GD spike ectodomain purification, except that 25 mM sodium phosphate pH 7 and 300 mM sodium chloride were used instead of 20 mM Tris-HC1 pH 8.0 and 150 mM NaCl. At this stage, SARS-CoV-2 S with the avi tag (from pCMV::SARS-CoV-2 S ecto avi) was biotinylated (BirA biotin-protein ligase standard reaction kit, Avidity) and further purified by size exclusion chromatography (Superose6, GE Healthcare). All purified proteins were then concentrated using a 100 kDa centrifugal filter (Amicon Ultra 0.5 mL
centrifugal filters, MilliporeSigma), residual imidazole was washed away by consecutive dilutions in the centrifugal filter unit with 20 mM Tris-HC1 pH 8.0 and 150 mM NaC1, and finally concentrated to 5 mg/ml and flash frozen.
All SARS-CoV-2 S NTD domain constructs (residues 14-307) with a C-terminal 8XHis-tag were produced in 100 mL culture of Expi293FTM Cells (ThermoFisher Scientific) grown in suspension using Expi293TM Expression Medium (ThermoFisher Scientific) at 37 C in a humidified 8% CO2 incubator rotating at 130 r.p.m.) (Walls et al., 2020) (Walls et al., 2020) (Walls et al., 2020) (Walls et al., 2020) (Walls et al., 2020) (Walls et al., 2020) (Walls et al., 2020) (Walls et al., 2020) (Walls et al., 2020) (Walls et al., 2020) (Walls et al., 2020). Cells grown to a density of 3 million cells per mL were transfected using pCMV::SARS-CoV-2 S NTD derivative mutants with the ExpiFectamineTM 293 Transfection Kit (ThermoFisher Scientific) with and cultivated for five days at which point the supernatant was harvested. His-tagged NTD
domain constructs were purified from clarified supernatants using 2 ml of cobalt resin (Takara Bio TALON), washing with 50 column volumes of 20 mM HEPES-HC1 pH 8.0 and 150 mM NaCl and eluted with 600 mM imidazole. Purified protein was concentrated using a 30 kDa centrifugal filter (Amicon Ultra 0.5 mL centrifugal filters, MilliporeSigma), the imidazole was washed away by consecutive dilutions in the centrifugal filter unit with 20 mM HEPES-HC1 pH 8.0 and 150 mM NaCl, and finally concentrated to 20 mg/ml and flash frozen. For crystallization, the purified NTD was not frozen but was further purified by size exclusion chromatography (Superdex Increase 75 10/300 G, GE Healthcare), concentrated using a new 30 kDa centrifugal filter, and used immediately.
Intact mass spectrometry analysis of purified NTD constructs The purpose of intact MS was to verify the n-terminal sequence on four constructs. N-linked glycans were removed by PNGase F after overnight non-denaturing reaction at room temperature. 4ug of deglycosylated protein was used for each injection on the LC-MS system to acquire intact MS signal after separation of protease and protein by LC (Agilent PLRP-S reversed phase column). Thermo MS
(Q
Exactive Plus Orbitrap) was used to acquire intact protein mass under denaturing condition. BioPharma Finder 3.2 software was used to deconvolute the raw m/z data to protein average mass.
Isolation of peripheral blood mononuclear cells (PBMCs), plasma and sera PBMCs were isolated from blood draw performed using tubes pre-filled with heparin, followed by Ficoll density gradient centrifugation. PBMCs were either used freshly along SARS-CoV2 Spike protein specific memory B cells sorting or stored in liquid nitrogen for later use. Sera were obtained from blood collected using tubes containing clot activator, followed by centrifugation and stored at -80 C.
B-cell isolation and recombinant mAb production Starting from freshly isolated PBMCs or upon cells thawing, B cells were enriched by staining with CD19 PE-Cy7 (BD Bioscience 341113) and incubation with anti-PE bead (Miltenyi Biotec, cat. 130- 048-801), followed by positive selection using LS columns. Enriched B cells were stained with anti-IgM, anti-IgD, anti-CD14 and anti-IgA, all PE labelled, and prefusion SARS-CoV-2 S with a biotinylated avi tag conjugated to Streptavidin Alexa-Fluor 647 (Life Technologies). SARSCoV-2 S-specific IgG+ memory B cells were sorted by flow cytometry via gating for PE
negative and Alexa-Fluor 647 positive cells. Cells were cultured for the screening of positive supernatants. Antibody VH and VL sequences were obtained by RT-PCR and mAbs were expressed as recombinant human Fab fragment or as IgG1 (G1m3 allotype) carrying the half-life extending M428L/N434S (LS) mutation in the Fc region.
ExpiCHO cells were transiently transfected with heavy and light chain expression vectors as previously described (Pinto et al., 2020).
Affinity purification was performed on AKTA Xpress FPLC (Cytiva) operated by UNICORN software version 5.11 (Build 407) using HiTrap Protein A columns (Cytiva) for full length human and hamster mAbs and CaptureSelect CHI -XL
MiniChrom columns (ThermoFisher Scientific) for Fab fragments, using PBS as mobile phase. Buffer exchange to the appropriate formulation buffer was performed with a HiTrap Fast desalting column (Cytiva). The final products were sterilized by filtration through 0.22 urn filters and stored at 4 C.
Enzyme-linked immunosorbent assay (ELISA) To determine specificity of recombinantly produced mAbs, 96 half area well-plates (Corning) were coated over-night at 4 C with of SARS-CoV-2 S, NTD or RBD
proteins prepared 1 ug/ml, 2 ug/m1 and 5 ug/m1 in PBS pH 7.2, respectively.
Plates were then blocked with PBS 1% BSA (Sigma) and subsequently incubated with mAbs serial dilutions for 1 h at room temperature. After 2 washing steps with PBS
0.05%
Tween 20 (PBS-T) (Sigma-Aldrich) goat anti-huma IgG secondary antibody (Southern Biotech) was added in incubated for 1 h at room temperature. Plates were then washed again with PBS-T and 4-NitroPhenyl phosphate (pNPP, Sigma-Aldrich) substrate added. After 30 min incubation, absorbance at 405 nm was measured by a plate reader (Biotek) and data plotted using Prism GraphPad.
For all other applications reported, the following ELISA procedure was followed: 30 ?Al of ectodomains (stabilized prefusion trimer) of S or NTD from SARS-CoV-2 were coated on 384 well ELISA plates at 1 ng/til for 16 hours at 4 C.
Plates were washed with a 405 TS Microplate Washer (BioTek Instruments) then blocked with 80 ttl SuperBlock (PBS) Blocking Buffer (Thermo Scientific) for 1 hour at 37 C. Plates were then washed and 30 pi antibodies were added to the plates at concentrations between 0.001 and 100,000 ng/ml and incubated for 1 h at 37 C. Plates were washed and then incubated with 30 tl of 1/5000 diluted goat anti-human Fc IgG-HRP
(invitrogen A18817). Plates were washed and then 30 ul Substrate T1VIB
microwell peroxidase (Seracare 5120-0083) was added for 4 min at room temperature. The colorimetric reaction was stopped by addition of 30 pi of 1 N HC1. A450 was read on a Varioskan Lux plate reader (Thermo Scientific).
MLV-based pseudovirus production and neutralization To generate SARS-CoV-2 S murine leukemia virus pseudotyped virus, HEK293T cells were seeded in 10-cm dishes in DMEM supplemented with 10% FBS.
The next day cells were transfected with a SARS-CoV-2 S glycoprotein-encoding plasmid harboring the D19 C-terminal truncation (Ou et al., 2020) using the X-tremeGENE HP DNA transfection reagent (Roche) according to the manufacturer's instructions. Cells were then incubated at 37 C with 5% CO2 for 72 h.
Supernatant was harvested and cleared from cellular debris by centrifugation at 400 X g, and stored at -80 C.
For neutralization assays, Vero E6 cells were seeded into white 96-well plates (PerkinElmer) at 20,000 cells/well and cultured overnight at 37 C with 5 %
CO2 in 100 pi DMEM supplemented with 10% FBS and 1% penicillin/streptomycin. The next day, MLV-SARS-CoV-2 pseudovirus was activated with 10 Fl g/m1 TPCK treated-Trypsin (Worthington Biochem) for 1 h at 37 C. Then recombinant antibodies at various concentrations were incubated with activated pseudovirus for 1 h at 37 C. The Vero E6 cells were then washed with DMEM, and the 50 01 of pseudovirus/mAbs mixes were added and incubated for 2 h at 37 C with 5 % CO2. After incubation, 50 .1 of DMEM containing 20% FBS and 2 % penicillin/streptomycin was added and the cells were incubated 48 h at 37 C with 5 % CO2. Following these 48 h of infection, culture medium was removed from the cells and 50 litl/wellof Bio-Glo (Promega) (diluted 1:2 with PBS with Ca2+Mg2+ (Thermo Fisher) was added to the cells and incubated in the dark for 15 min before reading on a Synergy H1 Hybrid Multi-Mode plate reader (Biotek). Measurements were done in duplicate and RLU values were converted to percentage of neutralization and plotted with a nonlinear regression curve fit in Graph Prism.
Neutralization of authentic SARS-CoV-2-Nluc virus Neutralization of authentic SARS-CoV-2 by entry-inhibition assay Neutralization was determined using SARS-CoV-2-Nluc, an infectious clone of SARSCoV-2 (based on strain 2019-nCoV/USA WA1/2020) which encodes nanoluciferase in place of the viral ORF7 and demonstrated comparable growth kinetics to wildtype virus (Xie et al., 2020). Vero E6 cells were seeded into black-walled, clear-bottom 96-well plates at 2 x 104 cells/well and cultured overnight at 37 C.
The next day, 9-point 4-fold serial dilutions of mAbs were prepared in infection media (DIVLEM
+ 10% FBS). SARS-CoV-2-Nluc was diluted in infection media at a final MOI of 0.1 or 0.01 PFU/cell, added to the mAb dilutions and incubated for 30 minutes at 37 C.
Media was removed from the Vero E6 cells, mAb-virus complexes were added and incubated at 37 C for 6 or 24 hours. Media was removed from the cells, Nano-Glo luciferase substrate (Promega) was added according to the manufacturer's recommendations, incubated for 10 minutes at room temperature and the luciferase signal was quantified on a VICTOR Nivo plate reader (Perkin Elmer).
Binding and affinity determination by Biolayer Interferometry (BLI) BLI measurements were performed using an Octet Red96 (ForteBio). All reagents were prepared in kinetics buffer (PBS plus 0.01% BSA) at the indicated concentrations BLI was used to assess antibody binding affinity to SARS-CoV-2 NTD. IgG
antibodies were prepared at 2.7 1.tg/m1 and captured on pre-hydrated Protein A
biosensors (Sartorius) for 1 min. The biosensors with immobilized antibodies were moved into kinetics buffer with SARS-CoV-2 NTD (concentrations tested: 333.3, 166.6, 83.3, 41.7, 20.8, 10.4, 5.2 nM) for 5 min (i.e. association). The dissociation of the SARS-CoV-2 NTD was then recorded for 9 min in wells containing kinetics buffer.
Affinity constants were calculated using a global fit model and results were plotted using GraphPad Prism.
BLI was also used to assess antibody competition studies to define the NTD
antigenic map. Biotinylated SARS-CoV-2 S protein was prepared at 10 p.g/m1 in kinetics buffer and loaded on pre-hydrated High Precision Streptavidin SAX
Biosensors (Sartorius) for 3 min. NTD mAbs at 20 pg/ml in kinetics buffer were then sequentially added to observe binding competition and signal recorded for 5 min (or 7 min) BLI was also used to assess mAb-mediated inhibition of SARS-CoV-2 S
binding to human recombinant ACE2. Before the assay SARS-CoV2 S ectodomain trimer (5 g/ml) was incubated with tested mAbs (30 p.g/m1) or no mAb for 30 minutes at 37 C. Biotinylated recombinant human ACE2 protein (2 ug/m1) was immobilized on High Precision Streptavidin SAX Biosensors (Sartorius). Next, an association step with S/mAb complexes was performed for 10 minutes. Results were plotted using GraphPad Prism.
Affinity determination by Surface Plasmon Resonance (SPR) SPR binding measurements were performed using a Biaeore T200 instrument where purified avi-tagged SARS-CoV-2 S D614G ectodomain trimer was captured using anti-AviTag pAb covalently immobilized on a CM5 sensor chip. The running buffer was Cytiva HBS-EP-F pH 7.4; measurements were performed at 25 C.
Affinity/avidity determinations were run as single-cycle kinetics, with a 3-fold dilution series of mAb starting from 300 nM, and each concentration injected for 180 sec.
Double reference-subtracted data were fit to a 1:1 binding model using Biacore Evaluation software. Fit results for IgG yielded apparent equilibrium dissociation constants due to avidity. For dissociation rates that were too slow to fit, equilibrium dissociation constants are reported as an upper limit Transient Expression of Sarbecovirus S protein in ExpiCHO-S Cells.
Immediately before transfection, ExpiCHO-S cells were seeded at 6 x 106 cells cells/mL in a volume of 5 mL in a 50 mL bioreactor. Spike coding plasmids were diluted in cold OptiPRO SFM, mixed with ExpiFectamine CHO Reagent (Life Technologies) and added to the cells. Transfected cells were then incubated at with 8% CO2 with an orbital shaking speed of 120 RPM (orbital diameter of 25 mm) for 42 hours Binding to cell surface expressed Sarbecovirus S proteins by Flow Cytometry Transiently transfected ExpiCHO cells were harvested and washed two times in wash buffer (PBS 1% BSA, 2 mM EDTA). Cells were counted and distributed into round bottom 96-well plates (Corning) and incubated with the NTD antibodies at the final concentration of 5 mg/mi. Alexa Fluor647-labelled Goat Anti-Human IgG
secondary Ab (Jackson Immunoresearch) was prepared at 1.5 g/ml added onto cells after two washing steps. Cells were then washed twice and resuspended in wash buffer for data acquisition at ZE5 cytometer (Biorad).
Fusion inhibition assay Vero E6 cells were seeded in 96 well plates at 15,000 cells per well in 70 1.11 DMEM with high glucose and 2.4% FBS (Hyclone). After 16 h at 37 C with 8 %
CO2, the cells were transfected with SARS-CoV-2-S-D19_pcDNA3.1 as follows: for 10 wells, 0.57 lig plasmid SARS-CoV-2- S-D19_pcDNA3.1 were mixed with 1.68 tl X-tremeGENE HP in 30 tl OPTIMEM. After 15 minutes incubation, the mixture was diluted 1:10 in DMEM medium and 30111 was added per well. A 4-fold serial dilution mAbs was prepared and added to the cells, with a starting concentration of 20 [tg/ml.
The following day, 30 [11 5X concentrated DRAQ5 in DMEM was added per well and incubated for 2 hours at 37 C. Nine images of each well were acquired with a Cytation 5 equipment for analysis.
Measurement of Fc-effector functions mAb-dependent activation of human FcyRIIIa was performed with a bioluminescent reporter assay. ExpiCHO cells stably expressing full-length wild-type SARS-CoV-2 S (target cells) were incubated with different amounts of mAbs.
After a

15-minute incubation, .Turkat cells stably expressing FcyRIIIa receptor (V158 variant) or FcyRIIa receptor (H131 variant) and NFAT-driven luciferase gene (effector cells) were added at an effector to target ratio of 6:1 for FcyRIIIa and 5:1 for FcyRIIa.
Signaling was quantified by the luciferase signal produced as a result of NFAT
pathway activation. Luminescence was measured after 20 hours of incubation at 37 C
with 5%
CO2 with a luminometer using the Bio-Glo-TM Luciferase Assay Reagent according to the manufacturer's instructions (Promega, Cat. Nr.: G9798, G7018 and G9995).
Cell-surface mAb-mediated Si shedding CHO cells stably expressing wild-type SARS-CoV-2 S were resuspended in wash buffer (PBS 1 % BSA, 2 mM EDTA) and treated with 10 l.t.g/mL TPCK-trypsin (Worthington Biochem) for 30 min at 37 C. Cells were then washed and distributed into round bottom 96-well plates (90,000 cells/well). MAbs were added to cells at 15 mg/mL final concentration for 180 min at 37 C. Cells were collected at different time points (5, 30, 60, 120 and 180), washed with wash buffer at 4 C, and incubated with 1.5 mg/mL secondary goat anti-human IgG, Fc fragment specific (Jackson ImmunoResearch) on ice for 20 min. Cells were washed and resuspended in wash buffer and analyzed with ZE5 FACS (Bio-rad).
Generation of stable overexpression cell lines Lentiviruses were generated by co-transfection of Lenti-X 293T cells (Takara) with lentiviral expression plasmids encoding DC-SIGN (CD209), L-SIGN (CLEC4M), SIGLEC1, TMPRSS2 or ACE2 (all obtained from Genecopoeia) and the respective lentiviral helper plasmids. Forty-eight hours post transfection, lentivirus in the supernatant was harvested and concentrated by ultracentrifugation for 2 h at 20,000 rpm. Lenti-X 293T (Takara), Vero E6 (ATCC), MRCS (Sigma-Aldrich), A549 (ATCC) were transduced in the presence of 6 ug/mL polybrene (Millipore) for 24 h.
Cell lines overexpressing two transgenes were transduced subsequently. Selection with puromycin and/or blastici din (Gibco) was started two days after transduction and selection reagent was kept in the growth medium for all subsequent culturing.
Single cell clones were derived from the A549-ACE2-TMPRSS2 cell line, all other cell lines represent cell pools.
SAPS-Co V-2 neutralization Vero E6 or Vero E6-TMPRSS2 cells cultured in DMEM supplemented with 10% FBS (VWR) and lx Penicillin/Streptomycin (Thermo Fisher Scientific) were seeded in black 96-well plates at 20,000 cells/well. Serial 1:4 dilutions of the monoclonal antibodies were incubated with 200 pfu of SARS-CoV-2 (isolate USA-WA1/2020, passage 3, passaged in Vero E6 cells) for 30 min at 37 C in a BSL-3 facility. Cell supernatant was removed and the virus-antibody mixture was added to the cells. 24 h post infection, cells were fixed with 4% paraformaldehyde for 30 min, followed by two PBS (pH 7.4) washes and permeabilization with 0.25% Triton X-in PBS for 30 min. After blocking in 5% milk powder/PBS for 30 min, cells were incubated with a primary antibody targeting SARS-CoV-2 nucleocapsid protein (Sino Biological, cat. 40143-R001) at a 1:2000 dilution for lh. After washing and incubation with a secondary Alexa647-labeled antibody mixed with 1 ug/ml Hoechst33342 for hour, plates were imaged on an automated cell-imaging reader (Cytation 5, Biotek) and nucleocapsid-positive cells were counted using the manufacturer's supplied software.
SAPS-CoV-2-Nhic neutralization Neutralization was determined using SARS-CoV-2-Nluc, an infectious clone of SARS-CoV-2 (based on strain 2019-nCoV/USA WA1/2020) encoding nanoluciferase in place of the viral ORF7, which demonstrates comparable growth kinetics to wild type virus (Xie et al., Nat Comm, 2020, https://doi.org/10.1038/s41467-020-19055-7). Cells were seeded into black-walled, clear-bottom 96-well plates at 20,000 cells/well (293T
cells were seeded into poly-L-lysine-coated wells at 35,000 cells/well) and cultured overnight at 37 C. The next day, 9-point 4-fold serial dilutions of antibodies were prepared in infection media (DMEM + 10% FBS). SARS-CoV-2-Nluc was diluted in infection media at the indicated MOT, added to the antibody dilutions and incubated for min at 37 C. Media was removed from the cells, mAb-virus complexes were added, and cells were incubated at 37 C for 24 h. Media was removed from the cells, Nano-Glo luciferase substrate (Promega) was added according to the manufacturer's recommendations, incubated for 10 min at RT and luciferase signal was quantified on a 25 VICTOR Nivo plate reader (Perkin Elmer).
SAPS-Co V-2 pseudotyped VSV production and neutralization To generate SARS-CoV-2 pseudotyped vesicular stomatitis virus, Lenti-X 293T
cells (Takara) were seeded in 10-cm dishes for 80% next day confluency The next day, cells were transfected with a plasmid encoding for SARS-CoV-2 5-glycoprotein 30 (YP 009724390.1) harboring a C-terminal 19 aa truncation using TransIT-Lenti (Minis Bio) according to the manufacturer's instructions. One day post-transfection, cells were infected with VSV(G*AG-luciferase) (Kerafast) at an MOI of 3 infectious units/cell.
Viral inoculum was washed off after one hour and cells were incubated for another day at 37 C. The cell supernatant containing SARS-CoV-2 pseudotyped VSV was collected at day 2 post-transfection, centrifuged at 1000 x g for 5 minutes to remove cellular debris, aliquoted, and frozen at -80 C.
For viral neutralization, Cells were seeded into black-walled, clear-bottom 96-well plates at 20,000 cells/well (293T cells were seeded into poly-L-lysine-coated wells at 35,000 cells/well) and cultured overnight at 37 C. The next day, 9-point 4-fold serial dilutions of antibodies were prepared in media. SARS-CoV-2 pseudotyped VSV was diluted 1:30 in media in the presence of 100 ng/mL anti-VSV-G antibody (clone 8G5F11, Absolute Antibody) and added 1:1 to each antibody dilution.
Virus:antibody mixtures were incubated for 1 hour at 37 C. Media was removed from the cells and 50 pL of virus:antibody mixtures were added to the cells. One hour post-infection, 100 pL
of media was added to all wells and incubated for 17-20 hours at 37 C. Media was removed and 50 pi, of Bio-Glo reagent (Promega) was added to each well. The plate was shaken on a plate shaker at 300 RPM at room temperature for 15 minutes and RLUs were read on an EnSight plate reader (Perkin-Elmer).
Transfection-based attachment receptor screen Lenti-X 293T cells (Takara) were transfected with plasmids encoding the following receptor candidates (all purchased from Genecopoeia): ACE2 (NM
021804), DC-SIGN (NM 021155), L-SIGN (BC110614), LGALS3 (NM 002306), SIGLEC1 (NM 023068), SIGLEC3 (XM 057602), SIGLEC9 (BC035365), SIGLEC10 (NM 033130), MGL (NM 182906), MINCLE (NIVI 014358), CD 147 (NMI 98589), ASGR1 (NM 001671.4), ASGR2 (NM 080913), NRP1 (NM 003873). One day post transfection, cells were infected with SARS-CoV-2 pseudotyped VSV at 1:20 dilution in the presence of 100 ng/mL anti-VSV-G antibody (clone 8G5F11, Absolute Antibody) at 37 C. One hour post-infection, 100 pL of media was added to all wells and incubated for 17-20 hours at 37 C Media was removed and 50 pL of Bio-Glo reagent (Promega) was added to each well. The plate was shaken on a plate shaker at 300 RPM at room temperature for 15 minutes and RLUs were read on an EnSight plate reader (Perkin-Elmer).

Trans-infection Parental HeLa cells or HeLa cells stably expressing DC-SIGN, L-SIGN or SIGLEC1 were seeded at 5,000 cells per well in black-walled clear-bottom 96-well plates. One day later, cells reached about 50% confluency and were inoculated with SARS-CoV-2 pseudotyped VSV at 1:10 dilution in the presence of 100 ng/mL anti-VSV-G antibody (clone 8G5F11, Absolute Antibody) at 37 C for 2 h. For antibody-mediated inhibition of trans-infection, cells were pre-incubated with 10 ug/mL
anti-SIGLEC1 antibody (Biolegend, clone 7-239) for 30 min. After 2 h inoculation, cells were washed four times with complete medium and 10,000 VeroE6-TIVIPRSS2 cells per well were added and incubated 17-20 h at 37 C for trans-infection. Media was removed and 50 [it of Bio-Glo reagent (Promega) was added to each well. The plate was shaken on a plate shaker at 300 RPM at room temperature for 15 minutes and RLUs were read on an EnSight plate reader (Perkin-Elmer).
Cell-cell fusion of CHO-S cells CHO cells stably expressing SARS-CoV-2 S-glycoprotein were seeded in 96 well plates for microscopy (Thermo Fisher Scientific) at 12'500 cells/well and the following day, different concentrations of mAbs and nuclei marker Hoechst (final dilution 1:1000) were added to the cells and incubated for additional 24h hours. Fusion degree was established using the Cytation 5 Imager (BioTek) and an object detection protocol was used to detect nuclei as objects and measure their size. The nuclei of fused cells (i.e., syncytia) are found aggregated at the center of the syncitia and are recognized as a unique large object that is gated according to its size. The area of the objects in fused cells divided by the total area of all the object multiplied by 100 provides the percentage of fused cells Immunofluorescence analysis HEK 293T cells were seeded onto poly-D-Lysine-coated 96-well plates (Sigma-Aldrich) and fixed 24 h after seeding with 4% paraformaldehyde for 30 min, followed by two PBS (pH 7.4) washes and permeabilizati on with 0.25% Triton X-100 in PBS for min. Cells were incubated with primary antibodies anti-DC-SIGN/L- SIGN
30 (Biolegend, cat. 845002, 1:500 dilution), anti-DC-SIGN (Cell Signaling, cat. 13193S, 1:500 dilution), anti-SIGLEC1 (Biolegend, cat. 346002, 1:500 dilution) or anti-(R&D Systems, cat. AF933, 1:200 dilution) diluted in 3% milk powder/PBS for 2 h at room temperature. After washing and incubation with a secondary Alexa647-labeled antibody mixed with 1 ug/ml Hoechst33342 for 1 hour, plates were imaged on an inverted fluorescence microscope (Echo Revolve).
ACE2/TIVIPRSS2 RT-qPCR
RNA was extracted from the cells using the NucleoSpin RNA Plus kit (Macherey-Nagel) according to the manufacturer's protocol. RNA was reverse transcribed using the High Capacity cDNA Reverse Transcription kit (Applied Biosystems) according to the manufacturer's instructions. Intracellular levels of ACE2 (Forward Primer: CAAGAGCAAACGGTTGAACAC, Reverse Primer:
CCAGAGCCTCTCATTGTAGTCT), HPRT (Forward Primer:
CCTGGCGTCGTGATTAGTG, Reverse Primer: ACACCCTTTCCAAATCCTCAG), and TMPRSS2 (Forward Primer: CAAGTGCTCCRACTCTGGGAT, Reverse Primer:
AACACACCGRTTCTCGTCCTC) were quantified using the Luna Universal qPCR
Master Mix (New England Biolabs) according to the manufacturer's protocol.
Levels of ACE2 and TMPRSS2 were normalized to HPRT. Hela cells were used as the reference sample. All qPCRs were run on a QuantStudio 3 Real-Time PCR System (Applied Biosystems).
SARS2 D614G Spike Production and biotinylation Prefusion-stabilized SARS2 D614G spike (comprising amino acid sequence Q14 to K1211) with a C-terminal TEV cleavage site, T4 bacteriophage fibritin foldon, 8x His-, Avi- and EPEA-tag was transfected into HEK293 Freestyle cells, using 293fectin as a transfection reagent. Cells were left to produce protein for three days at 37 C. Afterwards, supernatant was harvested by centrifuging cells for 30 minutes at 500 xg, followed by another spin for 30 minutes at 4000 xg. Cell culture supernatant was filtered through a 0.2 um filter and loaded onto a 5 mL C-tag affinity matrix column, pre-equilibrated with 50 mM Tris pH 8 and 200 mM NaCl. SARS2 D614G spike was eluted, using 10 column volumes of 100 mM Tris, 200 mM NaCl and 3.8 mM SEPEA
peptide. Elution peak was concentrated and injected on a Superose 6 increase GL gel filtration column, using 50 mM Tris pH 8 and 200 mM NaCl as a running buffer. SEC fractions corresponding to monodisperse SARS2 D614G spike were collected and flash frozen in liquid nitrogen for storage at -80 C. Purified D614G spike protein was biotinylated using BirA500 biotinylation kit from Avidity. To 50 ug of spike protein, 5 ug of BirA, and 11 uL of BiomixA and BiomixB was added.
Final spike protein concentration during the biotinylation reaction was ¨1 uM.
The reaction was left to proceed for 16 hours at 4 C. Then, protein was desalted using two Zeba spin columns pre-equilibrated with lx PBS pH 7.4.
_How cytometry analysis for DC-,S'IGN, L-SIGN, ,SYGLEC1 and ACE-2 HEK 293T cells expressing DC-SIGN, L-SIGN, SIGLEC1 or ACE2 were resuspended at 4x106 cells/mL and 100 pL per well were seeded onto V-bottom 96-well plates (Corning, 3894). The plate was centrifuged at 2,000 rpm for 5 minutes and washed with PBS (pH 7.4). The cells were resuspended in 200 p1_, of PBS
containing Ghost violet 510 viability dye (Cell Signaling, cat. 13-0870-T100, 1:1,000 dilution), incubated for 15 minutes on ice and then washed. The cells were resuspended in 100 pL
of FACS buffer prepared with 0.5% BSA (Sigma-Aldrich) in PBS containing the primary antibodies at a 1:100 dilution: mouse anti-DC/L-SIGN (Biolegend, cat.
845002), rabbit anti-DC-SIGN (Cell Signaling, cat. 13193), mouse anti-SIGLEC1 (Biologend, cat. 346002) or goat anti-ACE2 (R&D Systems, cat. AF933). After 1 h incubation on ice, the cells were washed two times and resuspended in FACS
buffer containing the Alexa Fluor-488-labeled secondary antibodies at a 1:200 dilution: goat anti-mouse (Invitrogen cat. A11001), goat anti-rabbit (Invitrogen cat. Al1008) or donkey anti-goat (Invitrogen cat. A11055). After incubation for 45 min on ice, the cells were washed three times with 200 L of FACS buffer and fixed with 2004, of 4%
PFA
(Alfa Aesar) for 15 mins at room temperature. Cells were washed three times, resuspended in 200pL of FACS buffer and analyzed by flow cytometry using the CytoFLEX flow cytometer (Beckman Coulter).
Flow cytometry of SARS-CoV-2 Spike and RBD binding to cells Biotinylated SARS-CoV-2 Spike D6 14G protein (Spikebiotin, in-house generated) or the biotinylated SARS-CoV-2 Spike receptor-binding domain (RBDbiotin, Sino Biological, 40592-V08B) were incubated with Alexa Fluor 647 streptavidin (AF647-strep, Invitrogen, S21374) at a 1:20 ratio by volume for 20 min at room temperature. The labeled proteins were then stored at 4 C until further use. Cells were dissociated with TrpLE Express (Gibco, 12605-010) and 105 cells were transferred to each well of a 96-well V bottom plate (Corning, 3894). Cells were washed twice in flow cytometry buffer (2% FBS in PBS (w/o Ca/Mg)) and stained with Spikebiotin-AF647-strep at a final concentration of 20 [tg/m1 or RBDbiotin-AF647-strep at a final concentration of 7.5 g/ml for lh on ice. Stained cells were washed twice with flow cytometry buffer, resuspended in I% PFA (Electron Microscopy Sciences, 15714-S) and analyzed with the Cytoflex LX (Beckman Coulter).
Recombinant expression of SARS-CoV-2-specific mAbs.
Human mAbs were isolated from plasma cells or memory B cells of SARS-CoV-2 immune donors, as previously described. Recombinant antibodies were expressed in ExpiCHO cells at 37 C and 8% CO2. Cells were transfected using ExpiFectamine. Transfected cells were supplemented 1 day after transfection with ExpiCHO Feed and ExpiFectamine CHO Enhancer. Cell culture supernatant was collected eight days after transfection and filtered through a 0.2 p.m filter.
Recombinant antibodies were affinity purified on an AKTA xpress FPLC device using 5 mL
HiTrapTm MabSelectTM PrismA columns followed by buffer exchange to Histidine buffer (20 mM Histidine, 8% sucrose, pH 6) using HiPrep 26/10 desalting columns SARS-CoV-2 infection model in hamster Virus preparation The SARS-CoV-2 strain used in this study, BetaCov/Belgium/GTM-03021/2020 (EPI ISL 109 40797612020-02-03), was recovered from a nasopharyngeal swab taken from an RT-qPCR confirmed asymptomatic patient who returned from Wuhan, China in February 2020. A close relation with the prototypic Wuhan-Hu-1 201 9-nCoV
(GenBank accession 112 number MN908947.3) strain was confirmed by phylogenetic analysis. Infectious virus was isolated by serial passaging on HuH7 and Vero E6 cells, passage 6 virus was used for the study described here. The titer of the virus stock was determined by end-point dilution on Vero E6 cells by the Reed and Muench method.
Cells Vero E6 cells (African green monkey kidney, ATCC CRL-1586) were cultured in minimal essential medium (Gibco) supplemented with 10% fetal bovine serum (Integro), 1% L- glutamine (Gibco) and 1% bicarbonate (Gibco). End-point titrations were performed with medium containing 2% fetal bovine serum instead of 10%.
SARS-Co V-2 infection model in hamsters The hamster infection model of SARS-CoV-2 has been described before. The specific study design is shown in the schematic below. In brief, wild-type Syrian Golden hamsters (Mesocricetus auratus) were purchased from Janvier Laboratories and were housed per two in ventilated isolator cages (IsoCage N Biocontainment System, Tecniplast) with ad libitum access to food and water and cage enrichment (wood block).
The animals were acclimated for 4 days prior to study start. Housing conditions and experimental procedures were approved by the ethics committee of animal experimentation of KU Leuven (license P065- 2020). Female 6-8 week old hamsters were anesthetized with ketamine/xylazine/atropine and inoculated intranasally with 50 pL containing 2x106 TCID50 SARS-CoV-2 (day 0).
Treatment regimen Animals were prophylactically treated 48h before infection by intraperitoneal administration (i.p.) and monitored for appearance, behavior, and weight. At day 4 post infection (p.i.), hamsters were euthanized by i.p. injection of 500 uL
Dolethal (200 mg/mL sodium pentobarbital, Vetoquinol SA). Lungs were collected and viral RNA

and infectious virus were quantified by RT-qPCR and end-point virus titration, respectively. Blood samples were collected before infection for PK analysis.
SARS-CoV-2 RT-qPCR
Collected lung tissues were homogenized using bead disruption (Precellys) in RLT buffer (RNeasyMinikit, Qiagen)and centrifuged (10.000 rpm, 5 min) to pellet the cell debris. RNA was extracted according to the manufacturer's instructions.
Of 50 uL eluate, 4 uL was used as a template in RT-qPCR reactions. RT-qPCR was performed on a LightCycler96 platform (Roche) using the iTaq Universal Probes One-Step RT-qPCR kit (BioRad) with N2 primers and probes targeting the nucleocapsid.
Standards of SARS-CoV-2 cDNA (IDT) were used to express viral genome copies per mg tissue or per mL serum.
End-point virus titrations Lung tissues were homogenized using bead disruption (Precellys) in 350 uL
minimal essential medium and centrifuged (10,000 rpm, 5min, 4 C) to pellet the cell debris. To quantify infectious SARS-CoV-2 particles, endpoint titrations were performed on confluent Vero E6 cells in 96- well plates. Viral titers were calculated by the Reed and Muench method using the Lindenbach calculator and were expressed as 50% tissue culture infectious dose (TCID50) per mg tissue.
Histology For histological examination, the lungs were fixed overnight in 4%
formaldehyde and embedded in paraffin. Tissue sections (5 prn) were analyzed after staining with hematoxylin and eosin and scored blindly for lung damage by an expert pathologist. The scored parameters, to which a cumulative score of 1 to 3 was attributed, were the following: congestion, intra-alveolar hemorrhagic, apoptotic bodies in bronchus wall, necrotizing bronchiolitis, perivascular edema, bronchopneumonia, perivascular inflammation, peribronchial inflammation and vasculitis.
Binding of immunocomplexes to hamster monocytes Immunocomplexes (IC) were generated by complexing S309 mAb (hamster IgG, either wt or N297A) with a biotinylated anti-idiotype fab fragment and Alexa-488-streptavidin, using a precise molar ratio (4:8:1, respectively). Pre-generated fluorescent IC were serially diluted incubated at 4 C for 3 hrs with freshly revitalized hamster splenocytes, obtained from a naïve animal. Cellular binding was then evaluated by cytometry upon exclusion of dead cells and physical gating on monocyte population.
Results are expressed as Alexa-488 mean florescent intensity of the entire monocyte population.
Bionyarmatic analyses Processed Human Lung Cell Atlas (HLCA) data and cell-type annotations were downloaded from Github (github.com/krasnowlab/HLCA). Processed single-cell transcriptome data and annotation of lung epithelial and immune cells from SARS-CoV-2 infected individuals were downloaded from NCBI GEO database (ID:
GSE158055) and Github (github.com/zhangzlab/covid balf). Available sequence data from the second single-cell transcriptomics study by Liao et al. were downloaded from NCBI SRA (ID: PRJNA608742) for inspection of reads corresponding to viral RNA.

The proportion of sgRNA relative to genomic RNA was estimated by counting TRS-containing reads supporting a leader-TRS junction. Criteria and methods for detection of leader-TRS junction reads were adapted from Alexandersen et al. The viral genome reference and TRS annotation was based on Wuhan-Hu-1 NC 045512.2/MN908947.
Only 2 samples from individuals with severe COVID-19 had detectable leader-TRS
junction reads (SRR11181958, SRR11181959).

ACE2-INDEPENDENT MECHANISM OF SARS-CoV2 NEUTRALIZATION
In the following experiments, unless otherwise indicated, S309 antibody (VH of SEQ ID NO.442, VL of SEQ ID NO. :446) was expressed as recombinant IgG1 with M428L and N434S Fe mutations. In certain experiments, antibody S2X333 (VH of SEQ ID NO.52, VL of SEQ ID NO. :56) and/or 52E12 (VH of SEQ ID NO.:450, VL of SFQ TD NO .454) (also expressed as rTgG1) were tested Other tested antibodies included S2M11 (VH of SEQ ID NO.:458, VL of SEQ ID NO.462), S2D106, and 52X58 (Starr et al, Nature 597:97-102 (2021), which antibodies are incorporated herein by reference).
The effect of ACE2 overexpression on S309 antibody neutralization of infection was investigated. Vero E6 or Vero E6-TMPRSS2 cells were infected with SARS-CoV-2 (isolate USA-WA1/2020) at MOI 0.01 in the presence of S309 (10 g/ml). Cells were fixed 24h post infection, viral nucleocapsid protein was immunostained and quantified. Nucleocapsid staining was effectively absent in antibody-treated cells.
S309 had an IC50 (ng/mL) in Vero E6 cells of 65 and in Vero E6-TMPRSS2 of 91 (data not shown).
A panel of 7 cell lines (HeLa, 293T (wt), Vero E6, Huh7, 293T ACE2, MRC 5-ACE2-IMPRSS2, A549-ACE2-IMPRSS2 clone 5, A549-ACE2-IMPRSS2 clone 10) were infected with SARS-CoV-2-Nluc or VSV pseudotyped with the SARS-CoV-2 spike protein in the presence of S309. Luciferase signal was quantified 24h post infection. S309 maximum neutralization values were as shown in Table 8.

Table 8. Maximum Neutralization Values of S309 Virus/Pseudotype Cell Type SARS- VSV Pseudotype CoV-2-Nlue Vero E6 >99% >99%
Vero E6-TMPRSS2 >99% 96%
Huh7 98% 78%
293T ACE2 26% 34%
MRC5-ACE2-TMF'RSS2 87% 45%
A549-ACE2-TMPRSS2 89% 65%
clone 5 A549-ACE2-TMPRS S2 81% 42%
clone 10 Binding of purified, fluorescently-labeled SARS-CoV-2 spike protein binding to these cell lines was quantified by flow cytometry. HeLa and 239T WT cells had he lowest MFIs, followed by Huh7 and VeroE6 cells. 293T ACE2 cells (highest), MRC

ACE2-TMPRSS2 (third-highest), A549-ACE2-TMPRSS2 clone 5 (fourth-highest), and A549-ACE2-TMPRSS2 clone 10 (second-highest) had higher MFIs. Correlation analysis between spike binding maximum neutralization potential of S309 was determined; S309 Spearman correlation values were: r = -0.94 for both viral models. p = 0.017.
To further characterize SARS-CoV-2-susceptible cell lines, the seven cell lines described above were incubated with purified, fluorescently-labeled SARS-CoV-2 spike protein or RBD protein and protein binding was quantified by flow cytometry. In descending order of MFI, the cell lines were: A549-ACE2-TMPRSS2 clone 10; 293T
ACE2; MRC 5-ACE2-TMPRSS2; A549-ACE2-TMPRSS2 clone 5; Vero E6; Huh7;
293T (wt), and HeLa.
Selected lectins and published receptor candidates were screened using HEK293T cells infected with SARS-CoV-2 VSV pseudoviruses. ACE2, DC-SIGN, L-SIGN, and SIGLEC-1 gave the highest signals. ACE2 provided a signal of approximately 105 relative luminescence units (RLUs), and DC-SIGN, SIGLEC-1, and L-SIGN had signals of approximately 104RLUs. All other lectins/candidates tested gave signals of approximately 102¨ 103 RLUs.
HEK 293T, HeLa and MRCS cells were transiently transduced to overexpress DC-SIGN, L-SIGN, SIGLEC1 or ACE2 and infected with SARS-CoV-2 VSV
pseudoviruses. Uninfected cells and untransduced cells were included as controls. In 1-1EK2931 cells, ACE2, DC-SIGN, SIGLEC-1, and L-SIGN all provided substantial increases in infection. In HeLa and MRC5 cells, only ACE2 increased infection.
Stable HEK293T cell lines overexpressing DC-SIGN, L-SIGN, SIGLEC-1 or ACE2 were infected with authentic SARS-CoV-2 (MOI 0.1), fixed and immunostained at 24 hours for the SARS-CoV-2 nucleoprotein. Wild-type cells (infected and uninfected) were used as controls. Increased staining was observed in cells overexpressing DC-SIGN, L-SIGN, or SIGLEC-1, and staining was significantly increased in cells overexpressing ACE2.
Stable cell lines were infected with SARS-CoV-2-Nluc and luciferase levels were quantified at 24 hours. In ascending order of RLUs. uninfected (approx.

RLUs); parental 293T (approx. 104RLUs); DC-SIGN (approx. 105RLUs); L-SIGN
(approx. 105RLUs), SIGLEC-1 (approx. 105-106RLUs), ACE2 (>107 RLUs).
Stable cell lines were incubated with different concentration of anti-SIGLEC1 mAb (clone 7-239) and infected with SARS-CoV-2-Nluc. Infection as a percentage of untreated cells remained near to exceeded 100% in 293T cells expressing DC-SIGN, L-SIGN, or ACE2, but dropped to below 50% (0.2 1.1g/mL anti-SIGLEC) to close to 0 (1 Kg/mL or 5 Kg/mL anti-SIGLEC) in 293T cells expressing SIGLEC- I .
Single cell expression levels of selected potential SARS-CoV-2 (co)receptor candidates were determined in different lung cell types derived from the Human Lung Cell Atlas (nature.com/articles/s41586-020-2922-4). DC-SIGN, L-SIGN and SIGLEC-1 are expressed in a variety of cell types in the lung at levels similar to or even higher than ACE2.
Binding of antibodies targeting DC-/L-SIGN, DC-SIGN, SIGLEC1 or ACE2 on HEK293T cells stably over-expressing the respective attachment receptor was analyzed by flow cytometry and immunofluorescence analysis. HEK 293T cells over-expressing the respective attachment receptors were infected with VSV pseudotyped with SARS-COV-2 wildtype spike or spike bearing mutations of the B1.1.7 lineage.
Luminescence was analyzed one day post infection. Infection was increased in cells expressing the attachment receptors. Infection by VSV pseudotyped with either spike was similar for each test group. Cells expressing ACE2 gave the highest luminescence signal.
Vero E6 cells, in vitro differentiated moDCs or PBMCs were infected with SARS-CoV-2 at MOI 0.01. At 24h post infection, cells were fixed, immunostained for viral nucleocapsid protein and infected cells were quantified. Only VeroE6 cells showed infection (approximately 7% of cells). Supernatant of the infected cells was taken at 24, 48 and 72h and infectious viral titer was quantified by FFU assay on Vero E6 cells.
Major cell types with detectable SARS-CoV-2 genome in bronchoalveolar lavage fluid (BALF) and sputum of severe COVID-19 patients were assessed. A t-SNE
plot was generated, and the count of each SARS-CoV-2+ cell type was determined (total n=3,085 cells from 8 subjects in Ken et al. Cell 2021). Cell types were T, NK, plasma, neutrophil, macrophage, ciliated, squamous, and secretory. Expression of ACE2, DC-SIGN, L-SIGN, SIGLEC-1, and combinations of these was assessed for each cell type.
ACE2, DC-SIGN (CD209), L-SIGN (CLEC4M), SIGLEC1 transcript counts were correlated with SARS-CoV-2 RNA counts in macrophages and in secretory cells.
Correlation was based on counts (before log transformation), from Ren et al.
Cell 2021.
Representative data showing expression of receptors in stable HEK293T cell lines are shown in Figure 40. Cell lines were generated to overexpress DC-SIGN, L-SIGN or ACE2 by transducing HEK293T cells with lentivirus encoding the transgene, and immunofluorescence assays were performed to assess transgene expression.
Representative data showing the ability of VSV pseudovirus expressing SARS-CoV-2 S protein with luciferase reporter to infect the HEK293T cells (using a luminescence assay) are shown in Figure 41; expression of DC-SIGN or L-SIGN
increased pseudovirus infection levels by over 10-fold compared to infection of WT
HEK293T cells, and expression of ACE2 increased pseudovirus infection levels by over 100-fold compared to infection of WT HEK293T cells.

Neutralizing activity of mAb S309 against the VSV pseudovirus was assessed in the engineered HEK293T cells. S309 fully neutralized infection via DC-SIGN and L-SIGN, and to a lesser extent, ACE2.
The ability of live SARS-CoV-2 with luciferase reporter to infect the HEK293T
cells was examined using a luminescence assay. expression of DC-SIGN or L-SIGN
increased live virus infection levels by over 3-fold compared to infection of WT
1-1EK2931 cells, and expression of ACE2 increased live virus infection levels by over 100-fold compared to infection of WT HEK293T cells.
Neutralizing activity of mAb S309 against the VSV pseudovirus was assessed in the engineered HEK293T cells. S309 fully neutralized infection via DC-SIGN and L-SIGN, and neutralized infection via ACE2 to a lesser extent.
Experiments were performed to investigate whether S309 antibody can neutralize entry of SARS-CoV-2 via SIGLEC-1. Briefly, stable cell HEK293T
lines were generated as described above to overexpress DC-SIGN/L-SIGN, DC-SIGN, SIGLEC-1, or ACE2. Expression of DC-SIGN, L-SIGN, or SIGLEC increased live virus infection levels by over 10-fold compared to infection of WT HEK293T
cells, and expression of ACE2 increased pseudovirus infection levels by over 100-fold compared to infection of WT HEK293T cells. S309 fully neutralized infection via DC-SIGN, L-SIGN, and SIGLEC-1.
Expression of DC-SIGN (CD209) and other cell surface receptor proteins including SIGLEC-1 and other SIGLECs was determined on a variety of cell types.
Data are summarized in Figure 45.
Further experiments were performed to investigate the function(s) of DC-SIGN, L-SIGN, and SIGLEC-1 in SARS-CoV-2 infection. In one set of experiments, HEK293T cells stably expressing DC-SIGN, L-SIGN, SIGLEC-1 or ACE2 were infected with live SARS-CoV-2 Nluc at three different multiplicities of infection (MOI): 0.01, 0.1, and 1). Infection was determined using relative luminescence units and compared to infection in HEK293T cells (parental) At the lowest MOT
tested, an increase of infection in cells expressing DC-SIGN, L-SIGN, or SIGLEC was observed.
At the highest MOI tested, infection was not further increased versus parental by expression of DC-SIGN, L-SIGN, or SIGLEC. These data indicate that the parental 293T cells are susceptible to infection by SARS-CoV-2 and L-SIGN, DC-SIGN, and SIGLEC-1 enhance infection levels but do not function as primary receptors for infection.
In another set of experiments, 293T cells, HeLa cells, and MRCS cells were transiently transduced with lentivirus encoding DC-SIGN, L-SIGN, SIGLEC-1 or ACE2 and infected with VSV pseudovirus three days after transduction. While the 293T cells showed a low level of susceptibility (compare uninfected with untransduced), HeLa and MRCS cells were completely refractory to the virus.
The low level of infection in 293T cells can be increased by expression of L-SIGN, DC-SIGN, or SIGLEC-1, consistent with a role for these proteins as as attachment factors. The HeLa and MRCS cells remained refractory to infection even after expression of L-SIGN, DC-SIGN, or SIGLEC-1, and only become susceptible after expression of ACE2. These data indicate that L-SIGN, DC-SIGN, and SIGLEC-1 are not primary receptors for SARS-CoV-2.
The various embodiments described above can be combined to provide further embodiments. All of the U.S. patents, U.S. patent application publications, U.S. patent applications, foreign patents, foreign patent applications and non-patent publications referred to in this specification and/or listed in the Application Data Sheet, including U.S. Provisional Application No. 63/084,501, filed September 28, 2020; U.S.
Provisional Application No. 63/111,435, filed November 9, 2020; U.S.
Provisional Application No. 63/112,505, filed November 11, 2020; U.S. Provisional Application No. 63/119,545, filed November 30, 2020; U.S. Provisional Application No.
63/137,112 filed January 13, 2021; and U.S. Provisional Application No.
63/170,356, filed April 2, 2021, are incorporated herein by reference, in their entirety.
Aspects of the embodiments can be modified, if necessary to employ concepts of the various patents, applications and publications to provide yet further embodiments.
These and other changes can be made to the embodiments in light of the above-detailed description. In general, in the following claims, the terms used should not be construed to limit the claims to the specific embodiments disclosed in the specification and the claims, but should be construed to include all possible embodiments along with the full scope of equivalents to which such claims are entitled. Accordingly, the claims are not limited by the disclosure.

Claims (61)

PCT/ITS2021/052481What is claimed is:

1. An antibody, or antigen-binding fragment thereof, comprising a heavy chain variable domain (VH) comprising a CDRH1, a CDRH2, and a CDRH3, and a light chain variable domain (VL) comprising a CDRL1, a CDRL2, and a CDRL3, wherein:
(i) the CDRH1 comprises or consists of the amino acid sequence according to any one of SEQ ID NOs.: 53, 33, 43, 73, 83, 93, 103, 113, 123, 133, 143, 153, 163, 173, 183, 193, 203, 213, 223, 233, 243, 253, 263, 273, 283, 293, 303, 313, 323, 333, 343, 353, 363, 373, 383, 393, 403, 413, 423, or 433, or a sequence variant thereof comprising one, two, or three acid substitutions, one or more of which substitutions is optionally a conservative substitution and/or is a substitution to a germline-encoded amino acid;
(ii) the CDRH2 comprises or consists of the amino acid sequence according to any one of SEQ ID NOs.: 54, 34, 44, 74, 84, 94, 104, 114, 124, 134, 144, 154, 164, 174, 184, 194, 204, 214, 224, 234, 244, 254, 264, 274, 284, 294, 304, 314, 324, 334, 344, 354, 364, 374, 384, 394, 404, 414, 424, or 434, or a sequence variant thereof comprising one, two, or three amino acid substitutions, one or more of which substitutions is optionally a conservative substitution and/or is a substitution to a germline-encoded amino acid;
(iii) the CDRH3 comprises or consists of the amino acid sequence according to any one of SEQ ID NOs.: 55, 35, 45, 75, 85, 95, 105, 115, 125, 135, 145, 155, 165, 175, 185, 195, 205, 215, 225, 235, 245, 255, 265, 275, 285, 295, 305, 315, 325, 335, 345, 355, 365, 375, 385, 395, 405, 415, 425, or 435, or a sequence variant thereof comprising one, two, or three amino acid substitutions, one or more of which substitutions is optionally a conservative substitution and/or is a substitution to a germline-encoded amino acid;
(iv) the CDRL1 comprises or consists of the amino acid sequence according to any one of SEQ ID NOs.: 57, 37, 47, 77, 87, 97, 107, 117, 127, 137, 147, 157, 167, 177, 187, 197, 207, 217, 227, 237, 247, 257, 267, 277, 287, 297, 307, 317, 327, 337, 347, 357, 367, 377, 387, 397, 407, 417, 427, or 437, or a sequence variant thereof comprising one, two, or three amino acid substitutions, one or more of which substitutions is optionally a conservative substitution and/or is a substitution to a germline-encoded amino acid;
(v) the CDRL2 comprises or consists of the amino acid sequence according to any one of SEQ ID NOs.: 58, 38, 48, 78, 88, 98, 108, 118, 128, 138, 148, 158, 168, 178, 188, 198, 208, 218, 228, 238, 248, 258, 268, 278, 288, 298, 308, 318, 328, 338, 348, 358, 368, 378, 388, 398, 408, 418, 428, or 438, or a sequence variant thereof comprising one, two, or three amino acid substitutions, one or more of which substitutions is optionally a conservative substitution and/or is a substitution to a germline-encoded amino acid; and/or (vi) the CDRL3 comprises or consists of the amino acid sequence according to any one of SEQ ID NOs.: 59, 39, 49, 79, 89, 99, 109, 119, 129, 139, 149, 159, 169, 179, 189, 199, 209, 219, 229, 239, 249, 259, 269, 279, 289, 299, 309, 319, 329, 339, 349, 359, 369, 379, 389, 399, 409, 419, 429, or 439, or a sequence variant thereof comprising having one, two, or three amino acid substitutions, one or more of which substitutions is optionally a conservative substitution and/or is a substitution to a germline-encoded amino acid, wherein the antibody or antigen binding fragment is capable of binding to a surface glycoprotein of a SARS-CoV-2, optionally when the surface glycoprotein is expressed on a cell surface of a host cell and/or on a virion.

2. The antibody or antigen-binding fragment of claim 1, which is capable of neutralizing a SARS-CoV-2 infection in an in vitro model of infection and/or in an in vivo animal model of infection and/or in a human.

3 The antibody or antigen-binding fragment of any one of claims 1-2, comprising CDRH1, CDRH2, CDRH3, CDRL1, CDRL2, and CDRL3 amino acid sequences according to SEQ ID NOs.:

(i) 53-55 and 57-59, respectively;
(ii) 33-35 and 37-39, respectively, (iii) 43-45 and 47-49, respectively;
(iv) 73-75 and 77-79, respectively, (v) 83-85 and 87-89, respectively;
(vi) 93-95 and 97-99, respectively;
(vii) 103-105 and 107-109, respectively (viii) 113-115 and 117-119, respectively, (ix) 123-125 and 127-129, respectively;
(x) 133-135 and 137-139, respectively;
(xi) 143-145 and 147-149, respectively;
(xii) 153-155 and 157-159, respectively, (xiii) 163-165 and 167-169, respectively, (xiv) 173-175 and 177-179, respectively;
(xv) 183-185 and 187-189, respectively;
(xvi) 193-195 and 197-199, respectively;
(xvii) 203-205 and 207-209, respectively, (xviii) 213-215 and 217-219, respectively;
(xix) 223-225 and 227-229, respectively;
(xx) 233-235 and 237-239, respectively;
(xxi) 243-245 and 247-249, respectively;
(xxiii) 253-255 and 257-259, respectively, (xxiii) 263-265 and 267-269, respectively;
(xxiv) 273-275 and 277-279, respectively;
(xxv) 283-285 and 287-289, respectively, (xxvi) 293-295 and 297-299, respectively;
(xxvii) 303-305 and 307-309, respectively;
(xxviii) 313-315 and 317-319, respectively;
(xxix) 323-325 and 327-329, respectively, (xxx) 333-335 and 337-339, respectively;

(xxxi) 343-345 and 347-349, respectively;
(xxxii) 353-355 and 357-359, respectively;
(xxxiii) 363-365 and 367-369, respectively;
(xxxiv) 373-375 and 377-379, respectively;
(xxxv) 383-385 and 387-389, respectively;
(xxxvi) 393-395 and 397-399, respectively;
(xxxvii) 403-405 and 407-409, respectively;
(xxxviii) 413-415 and 417-419, respectively;
(xxxix) 423-425 and 427-429, respectively; or (xxxxi) 433-435 and 437-439, respectively.

4. The antibody or antigen-binding fragment of any one of claims 1-3, wherein:
(i) the VH comprises or consists of an amino acid sequence having at least 85% identity to the amino acid sequence according to any one of SEQ ID NOs.:
52, 32, 42, 72, 82, 92, 102, 112, 122, 132, 142, 152, 162, 172, 182 192, 202, 212, 222, 232, 242, 252, 262, 272, 282, 292, 302, 312, 322, 332, 342, 352, 362, 372, 382, 392, 402, 412, 422, and 432, wherein the variation is optionally limited to one or more framework regions and/or the variation comprises one or more substitution to a germline-encoded amino acid; and/or (ii) the VL comprises or consists of an amino acid sequence having at least 85% identity to the amino acid sequence according to any one of SEQ ID NOs.:
56, 36, 46, 76, 86, 96, 106, 116, 126, 136, 146, 156, 166, 176, 186, 196, 206, 216, 226, 236, 246, 256, 266, 276, 286, 296, 306, 316, 326, 336, 346, 356, 366, 376, 386, 396, 406, 416, 426, and 436, wherein the variation is optionally limited to one or more framework regions and/or the variation comprises one or more substitution to a germline-encoded amino acid.

5. The antibody or antigen-binding fragment of any one of claims 1-4, wherein the VH and the VL comprise or consist of the amino acid sequences according to SEQ ID NOs.:
(i) 52 and 56, respectively, (ii) 32 and 36, respectively;
(iii) 42 and 46, respectively;
(iv) 72 and 76, respectively, (v) 82 and 86, respectively, (vi) 92 and 96, respectively;
(vii) 102 and 106, respectively;
(viii) 112 and 116, respectively;
(ix) 122 and 126, respectively, (x) 132 and 136, respectively;
(xi) 142 and 146, respectively;
(xii) 152 and 156, respectively;
(xiii) 162 and 166, respectively;
(xiv) 172 and 176, respectively, (xv) 182 and 186, respectively;
(xvi) 192 and 196, respectively;
(xvii) 202 and 206, respectively;
(xviii) 212 and 216, respectively;
(xix) 222 and 226, respectively, (xx) 232 and 236, respectively;
(xxi) 242 and 246, respectively;
(xxii) 252 and 256, respectively, (xxiii) 262 and 266, respectively;
(xxiv) 272 and 276, respectively;
(xxv) 282 and 286, respectively;
(xxvi) 292 and 296, respectively, (xxvii) 302 and 306, respectively;

(xxviii) 312 and 316, respectively;
(xxix) 322 and 326, respectively;
(xxix) 332 and 336, respectively;
(xxxii) 342 and 346, respectively;
(xxxiv) 352 and 356, respectively;
(xxxv) 362 and 366, respectively;
(xxxvi) 372 and 376, respectively;
(xxxvii) 382 and 386, respectively;
(xxxviii) 392 and 396, respectively;
(xxxix) 402 and 406, respectively;
(xxxx) 412 and 416, respectively;
(xxxxi) 422 and 426, respectively; or (xxxxii) 432 and 436, respectively.

6. The antibody or antigen-binding fragment of any one of claims 1-5, which: (i) recognizes an epitope in a Domain A of SARS-CoV-2; (ii) is capable of neutralizing a SARS CoV-2 infection, (iii) is capable of eliciting at least one immune effector function against SARS CoV-2; (iv) is capable of preventing shedding, from a cell infected with SARS CoV-2, of S1 protein; or (v) any combination of (i)-(iv).

7. The antibody or antigen-binding fragment of any one of claims 1-6, which is a IgG, IgA, IgM, IgE, or IgD isotype.

8. The antibody or antigen-binding fragment of any one of claims 1-7, which is an IgG isotype selected from IgGl, IgG2, IgG3, and IgG4.

9. The antibody or antigen-binding fragment of any one of claims 1-8, which is human, humanized, or chimeric

10. The antibody or antigen-binding fragment of any one of claims 1-9, wherein the antibody, or the antigen-binding fragment, comprises a human antibody, a monoclonal antibody, a purified antibody, a single chain antibody, a Fab, a Fab', a F(ab')2, a Fv, a scFv, or a scFab.

11. The antibody or antigen-binding fragment of claim 10, wherein the scFv comprises more than one VH domain and more than one VL domain.

12. The antibody or antigen-binding fragment of any one of claims 1-11, wherein the antibody or antigen-binding fragment is a multi-specific antibody or antigen binding fragment.

13. The antibody or antigen-binding fragment of claim 12, wherein the antibody or antigen binding fragment is a bispecific antibody or antigen-binding fragment.

14. The antibody or antigen-binding fragment of claim 12 or 13, comprising.
(i) a first VH and a first VL; and (ii) a second VH and a second VL, wherein the first VH and the second VH are different and each independently comprise an amino acid sequence having at least 85% identity to the amino acid sequence set forth in any one of SEQ ID NOs.: 52, 32, 42, 72, 82, 92, 102, 112, 122, 132, 142, 152, 162, 172, 182 192, 202, 212, 222, 232, 242, 252, 262, 272, 282, 292, 302, 312, 322, 332, 342, 352, 362, 372, 382, 392, 402, 412, 422, and 432, wherein the first VL and the second VL are different and each independently comprise an amino acid sequence having at least 85% identity to the amino acid sequence set forth in any one of SEQ ID NOs.: 56, 36, 46, 76, 86, 96, 106, 116, 126, 136, 146, 156, 166, 176, 186, 196, 206, 216, 226, 236, 246, 256, 266, 276, 286, 296, 306, 316, 326, 336, 346, 356, 366, 376, 386, 396, 406, 416, 426, and 436, and wherein the first VH and the first VL together form a first antigen-binding site, and wherein the second VH and the second VL together form a second antigen-binding site.

15. The antibody or antigen-binding fragment of any one of claims 1-14, wherein the antibody or antigen-binding fragment further comprises a Fc polypeptide or a fragment thereof

16. The antibody or antigen-binding fragment of claim 15, wherein the Fc polypeptide or fragment thereof comprises:
(i) a mutation that enhances binding to a FcRn as compared to a reference Fc polypeptide that does not comprise the mutation; and/or (ii) a mutation that enhances binding to a FcyR as compared to a reference Fc polypeptide that does not comprise the mutation.

17. The antibody or antigen-binding fragment of claim 16, wherein the mutation that enhances binding to a FcRn comprises. M428L, N434S, N434H, N434A, N434S; M252Y; S254T; T256E; T250Q; P257I; Q311I; D376V; T307A; or E380A; or any combination thereof.

18. The antibody or antigen-binding fragment of claim 16 or 17, wherein the mutation that enhances binding to FcRn comprises:
(i) M428L7N434S;
(ii) M252Y/S254T/T256E;
(iii) T250Q/M428L;
(iv) P257I/Q3111;
(v) P257I/N43 4H;
(vi) D376V/N434H;
(vii) T307A/E380A/N434A; or (viii) any combination of (i)-(vii).

19. The antibody or antigen-binding fragment of any one of claims 16-18, wherein the mutation that enhances binding to FcRn comprises M428L/N434S.

20. The antibody or antigen-binding fragment of any one of claims 16-19, wherein the mutation that enhances binding to a FcyR comprises 5239D; 1332E;
A330L; G236A; or any combination thereof.

21. The antibody or antigen-binding fragment of any one of claims 16-20, wherein the mutation that enhances binding to a FcyR comprises:
(i) 5239D/I332E;
(ii) S239D/A330L/1332E;
(iii) G236A/5239D/I332E; or (iv) G236A/A330L/I332E.

22. The antibody or antigen-binding fragment of any one of claims 16-21, wherein the Fc polypeptide comprises a L234A mutation and a L235A mutation.

23. The antibody or antigen-binding fragment of any one of claims 1-22, which comprises a mutation that alters glycosylation, wherein the mutation that alters glycosylation comprises N297A, N297Q, or N297G, and/or which is aglycosylated and/or afucosylated.

24. An isolated polynucleotide encoding the antibody or antigen-binding fragment of any one of claims 1-23, or encoding a VH, a heavy chain, a VL, and/or a light chain of the antibody or the antigen-binding fragment.

25. The polynucleotide of claim 24, wherein the polynucleotide comprises deoxyribonucleic acid (DNA) or ribonucleic acid (RNA), wherein the RNA
optionally comprises messenger RNA (mRNA).

26. The polynucleotide of claim 24 or 25, which is codon-optimized for expression in a host cell.

27. The polynucleotide of any one of claims 24-26, comprising a polynucleotide having at least 50% identity to the polynucleotide sequence according to any one or more of SEQ ID NOs.: 60, 61, 30, 31, 40, 41, 50, 51, 70, 71, 80, 81, 90, 91, 100, 101, 110, 111, 120, 121, 130, 131, 140, 141, 150, 151, 160, 161, 170, 171, 180, 181, 190, 191, 200, 201, 210, 211, 220, 221, 230, 231, 240, 241, 250, 251, 260, 261, 270, 271, 280, 281, 290, 291, 300, 301, 310, 311, 320, 321, 330, 331, 340, 341, 350, 351, 360, 361, 370, 371, 380, 381, 390, 391, 400, 401, 410, 411, 420, 421, 430, 431, 440, and 441, or any combination thereof.

28. A recombinant vector comprising the polynucleotide of any one of claims 24-27.

29. A host cell comprising the polynucleotide of any one of claims 24-27 and/or the vector of claim 28, wherein the polynucleotide is heterologous to the host cell.

30. A human B cell comprising the polynucleotide of any one of claims 24-28, wherein polynucleotide is heterologous to the human B cell and/or wherein the human B cell is immortalized.

31. A composition or combination comprising:
(i) the antibody or antigen-binding fragment of any one of claims 1-23;
(ii) the polynucleotide of any one of claims 24-27;
(iii) the recombinant vector of claim 28;
(iv) the host cell of claim 29; and/or (v) the human B cell of claim 30, and an optional pharmaceutically acceptable excipient, carrier, or diluent.

32. The composition or combination of claim 31, comprising two or more antibodies or antigen-binding fragments of any one of claims 1-23, and/or comprising one or more antibody according to any one of claims 1-23 and an antibody or antigen-binding fragment that binds to a SARS CoV-2 surface glycoprotein RBD.

33. A composition comprising the polynucleotide of any one of claims 24-27 encapsulated in a carrier molecule, wherein the carrier molecule optionally comprises a lipid, a lipid-derived delivery vehicle, such as a liposome, a solid lipid nanoparticle, an oily suspension, a submicron lipid emulsion, a lipid microbubble, an inverse lipid micelle, a cochlear liposome, a lipid microtubule, a lipid microcylinder, lipid nanoparticle (LNP), or a nanoscale platform.

34. A method of treating a SARS-CoV-2 infection in a subject, the method comprising administering to the subject an effective amount of the antibody or antigen-binding fragment of any one of claims 1-23;
(ii) the polynucleotide of any one of claims 24-27;
(iii) the recombinant vector of claim 28, (iv) the host cell of claim 29;
(v) the human B cell of claim 30; and/or (vi) the composition or combination of any one of claims 31-33

35. The antibody or antigen-binding fragment of any one of claims 1-23, the polynucleotide of any one of claims 24-27, the recombinant vector of claim 28, the host cell of claim 29, the human B cell of claim 30, and/or the composition or combination of any one of claims 31-33 for use in a method of treating a SARS-CoV-2 infection in a subject.

36 The antibody or antigen-binding fragment of any one of claims 1-23, the polynucleotide of any one of claims 24-27, the recombinant vector of claim 28, the host cell of claim 29, the human B cell of claim 30, and/or the composition or combination of any one of claims 31-33 for use in the preparation of a medicament for the treatment of a SARS-CoV-2 infection in a subject.

37. A method for in vitro or ex vivo diagnosis of a SARS-CoV-2 infection, the method comprising:
contacting a sample from a subject with an antibody or antigen-binding fragment of any one of claims 1-23; and (ii) detecting a complex comprising an antigen and the antibody, or comprising an antigen and the antigen binding fragment.

38. The method of claim 37, wherein the sample comprises blood isolated from the subject.

39. An antibody, or an antigen-binding fragment thereof, that competes for binding to a SARS-CoV-2 surface glycoprotein with the antibody or antigen-binding fragment of any one of claims 1-23.

40. A method of preventing or treating or neutralizing a coronavirus infection in a subject, the method comprising administering to a subject an effective amount of (i) an antibody or antigen-binding fragment of any one of claims 1-23 or 39 and (ii) an antibody or antigen-binding fragment that is capable of specifically binding to a SARS CoV-2 S protein RBD.

41. A method of detecting a SARS-CoV-2 protein or polypeptide in a sample, comprising contacting the sample with the antibody or antigen-binding fragment of any one of claims 1-23 or 39 and detecting binding of the antibody or antigen-binding fragment to the SARS-CoV-2 protein or polypeptide.

42. The method of claim 41, wherein detecting binding of the antibody or antigen-binding fragment to the SARS-CoV-2 protein or polypeptide comprises immunohistochemistry, ELISA, agglutination, immuno-dot, immuno-chromatography, and/or immuno-filtration.

43. The antibody or antigen-binding fragment thereof of any one of claims 1-23 for use in a method of detecting a SARS-CoV-2 protein or polypeptide in a sample, the method comprising contacting the sample with the antibody or antigen-binding fragment and detecting binding of the antibody or antigen-binding fragment to the SARS-CoV-2 protein or polypeptide, wherein, optionally, detecting binding of the antibody or antigen-binding fragment to the SARS-CoV-2 protein or polypeptide comprises immunohistochemistry, ELISA, agglutination, immuno-dot, immuno-chromatography, and/or immuno-filtration.

44. A method of diagnosing a SARS-CoV-2 infection in a subject, comprising testing a biological sample from the subject for the presence of a SARS-CoV-2 protein or polypeptide, wherein the testing comprises contacting the sample with the antibody or antigen-binding fragment of any one of claims 1-23 and detecting binding of the antibody or antigen-binding fragment to the SARS-CoV-2 protein or polypeptide, wherein, optionally, detecting binding of the antibody or antigen-binding fragment to the SARS-CoV-2 protein or polypeptide comprises immunohistochemistry, ELIS A, agglutination, immuno-dot, immuno-chromatography, and/or immuno-filtration.

45 The method of claim 44, wherein the SARS-CoV-2 protein or polypeptide is detected by immunohistochemistry.

46. The method of any one of claims 41-45, wherein the sample comprises a nasal secretion, sputum, a bronchial lavage, urine, stool, saliva, sweat, or any combination thereof

47. An antibody or antigen-binding fragment thereof for use in a method of diagnosing a SARS-CoV-2 infection in a subject, the method comprising testing a biological sample from the subject for the presence of a SARS-CoV-2 protein or polypeptide, wherein the testing comprises contacting the sample with the antibody or antigen-binding fragment and detecting binding of the antibody or antigen-binding fragment to the SARS-CoV-2 protein or polypeptide, wherein, optionally, detecting binding of the antibody or antigen-binding fragment to the SARS-CoV-2 protein or polypeptide comprises immunohistochemistry, ELISA, agglutination, immuno-dot, immuno-chromatography, and/or immuno-filtration, wherein, optionally, the antibody or antigen-binding fragment is the antibody or antigen-binding fragment thereof of any one of claims 1-23.

48. The antibody or antigen-binding fragment of any one of claims 1-23 or the antibody or antigen-binding fragment for use of claim 43 or 47, or the method of any one of claims 41, 42, or 44-46, wherein the antibody or antigen-binding fragment comprises a detectable agent.

49. A kit comprising the antibody or antigen-binding fragment thereof of any one of claims 1-23, and optional instructions for using the antibody or antigen-binding fragment to detect the presence of a SARS-CoV-2 protein or polypeptide in a biological sample.

50. The kit according to claim 49 for use in a method of detecting the presence of a SARS-CoV-2 protein or polypeptide in a biological sample.

51. The kit of for use of claim 50, wherein the method comprises detecting the presence of a SARS-CoV-2 protein or polypeptide by immunohistochemistry, ELISA, agglutination, immuno-dot, immuno-chromatography, and/or immuno-filtration.

52. The kit of claim 49 or the kit for use of any one of claims 50 or 51, further comprising a detectably labeled secondary antibody.

53. The kit of claim 49 or the kit for use of any one of claims 50-52, further comprising one or more of a sample buffer, a wash buffer, an immunodetection buffer, a substrate, detection means, a control sample, a reference sample, and instructions for use.

54. The kit of claim 49 or the kit for use of any one of claims 50-53, wherein the sample comprises a nasal secretion, sputum, bronchial lavage, urine, stool, saliva, and/or sweat.

55. The composition or combination of claim 32, comprising (a) antibody S2X333 (or an antigen-binding fragment thereof) or an antibody or antigen-binding fragment thereof that competes with antibody S2X333 for SARS-CoV-2 S protein binding and (b) antibody S309 (or an antigen-binding fragment thereof) or an antibody or antigen-binding fragment thereof that competes with antibody S309 for SARS-CoV-2 S protein binding.

56. The composition of claim 32, comprising a) antibody S2X333 (or an antigen-binding fragment thereof) or an antibody or an antigen-binding fragment thereof that competes with antibody S2X333 for SARS-CoV-2 S protein binding and b) antibody 52EI2 (or an antigen-binding fragment thereof) or an antibody or an antigen-binding fragment thereof that competes with antibody S2E12 for SARS-CoV-2 S
protein binding.

57. The composition of claim 32, comprising (a) antibody 52X333 (or an antigen-binding fragment thereof) or an antibody or an antigen-binding fragment thereof that competes with antibody 52X333 for SARS-CoV-2 S protein binding and (b) antibody S2M11 (or an antigen-binding fragment thereof) or an antibody or an antigen-binding fragment thereof that competes with antibody S2M11 for SARS-CoV-2 S protein binding.

58. The antibody or antigen-binding fragment of claim 12 or 13, comprising (i) a first VH and a first VL; and (ii) a second VH and a second VL, wherein the first VH comprises an amino acid sequence having at least 85% identity to the amino acid sequence set forth in SEQ ID NO: 52 and the first VL comprises an amino acid sequence having at least 85% identity to the amino acid sequence set forth in SEQ ID
NO: 56; and a) the second VH comprises an amino acid sequence having at least 85%
identity to the amino acid sequence set forth in SEQ ID NO: 442 and the second VL
comprises an amino acid sequence having at least 85% identity to the amino acid sequence set forth in SEQ ID NO: 446;
b) the second VH comprises an amino acid sequence having at least 85%
identity to the amino acid sequence set forth in SEQ ID NO: 450 and the second VL
comprises an amino acid sequence having at least 85% identity to the amino acid sequence set forth in SEQ ID NO. 454, or c) the second VH comprises an amino acid sequence having at least 85%
identity to the amino acid sequence set forth in SEQ ID NO: 458 and the second VL
comprises an amino acid sequence having at least 85% identity to the amino acid sequence set forth in SEQ ID NO: 462; and wherein the first VH and the first VL together form a first antigen-binding site, and wherein the second VH and the second VL together form a second antigen-binding site.

59. A method of treating or preventing SARS-CoV-2 infection comprising administering a composition or combination of any one of claims 55-57 or the antibody or antigen-binding fragment of claim 58

60. The composition or combination of any one of claims 55-57, wherein, optionally the antibody or antigen-binding fragment of a) and/or b) comprises (i) a Fc polypeptide comprising a mutation that enhances binding to a FcRn as compared to a reference Fc polypeptide that does not comprise the mutation; and/or (ii) a Fc polypeptide comprising a mutation that enhances binding to a Fcylt as compared to a reference Fc polypeptide that does not comprise the mutation.

61 The antibody or antigen-binding fragment of claim 58, or the method of claim 59, wherein, optionally, the antibody or antigen-binding fragment comprises (i) a Fc polypeptide comprising a mutation that enhances binding to a FcRn as compared to a reference Fc polypeptide that does not comprise the mutation; and/or (ii) a Fc polypeptide comprising a mutation that enhances binding to a Fc7R as compared to a reference Fc polypeptide that does not comprise the mutation