Human Anti-SARS-COV-2 spike (s) antibodies

Abstract

The present invention includes a monoclonal antibody or antigen-binding fragment thereof, methods of using, detection, recombinant vectors, host cells, kits, variants, and pharmaceutical compositions that include the antibody or antigen-binding fragment thereof that binds to the SARS-CoV-2 Spike protein.

Description

HUMAN ANTI-SARS-COV-2 SPIKE (S) ANTIBODIESCROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims benefit of and priority to U.S. Provisional Application No. 63 / 677,975, filed July 31, 2024, which is specifically incorporated by reference herein in its entirety.REFERENCE TO THE SEQUENCE LISTING

[0002] The Sequence Listing submitted as an XML file named “LJI_2024-109-02PCT_ST26.xml” created on July 31, 2025, and having a size of 57,626 bytes is hereby incorporated by reference pursuant to 37 C.F.R. § 1.834(c)(1).TECHNICAL FIELD OF THE INVENTION

[0003] The present invention relates in general to the field of antibodies against coronavirus, and more particularly, to human antibodies targeting the Spike protein of SARS CoV-2.STATEMENT OF GOVERNMENT SUPPORT

[0004] The inventions described in the present disclosure were made with government support under Contract No. CCHI AI142742, awarded by the National Institutes of Health. The government has certain rights in the invention.BACKGROUND OF THE INVENTION

[0005] Without limiting the scope of the invention, its background is described in connection with SARS-CoV-2.

[0006] The worldwide spread of SARS-CoV-2 in the human population resulted in the ongoing COVID- 19 pandemic that has already caused more than 700 million infections and more than 7 million deaths. To initiate infection, the SARS-CoV-2 spike (S) glycoprotein promotes binding to ACE2 located on the surface of the host cell, initiating a cascade of conformational changes in the protein that drives from a metastable pre-fusion conformation to a stable post-fusion conformation. That reorganization of the protein exposes the fusion peptide and a fusion between the viral and host membranes is driven by the S2 chain of the proprotein. Given its external location on the virus and its functionality, SARS-CoV-2 spike ‘S’ protein in its pre- fusion state is one target of neutralizing antibodies and therefore the main target of the design of safe and effective therapies.SUMMARY OF THE INVENTION

[0007] As embodied and broadly described herein, an aspect of the present disclosure relates to a monoclonal antibody or antigen-binding fragment thereof comprising heavy chain variable regioncomprising an amino acid sequence at least about 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, or 100% identical to an amino acid sequence of SEQ ID NOS: 1-29. In another aspect, the antibody light chain variable region comprises an amino acid sequence at least about 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, or 100% identical to an amino acid sequence of SEQ ID NOS:30-58. In alternative aspects, the antibody or antigen-binding fragment thereof comprises those heavy chain complementarity determining regions (CDR)l, CDR2, and CDR3 as set forth in any one of SEQ ID NOS 1-29 and / or those light chain CDR1, CDR2, and CDR3 as set forth in any one of SEQ ID NOS 30-58. In some forms there is 1, 2, or 3 mutations in in one, two or all three of the CDRs of any of SEQ ID NOS:1- 58. In some forms there are no mutations in the CDRs relative to SEQ ID NOS:l-58. In another aspect, the antigen-binding fragment is a recombinant single-chain fragment variable (scFV) antibody, Fab fragment, F(ab’)2 fragment, or Fv fragment. In another aspect, the antibody or antigenbinding fragment is chimeric, humanized, fully human, bispecific, or multimeric. In another aspect, the antibody or antigen-binding fragment comprises an Fc portion mutated to at least one of: eliminate or enhance Fc Receptor (FcR) interactions to change a half-life, increase or decrease antibodydependent cellular cytotoxicity, or increase or decrease complement activation. In another aspect, the antibody or antigen-binding fragment is adapted for administration or genetic delivery with an RNA or DNA sequence or vector encoding the antibody or antigen-binding fragment. In another aspect, the monoclonal antibody further comprises a peptide linker in a hinge region between the variable and constant domains of the heavy chain.

[0008] As embodied and broadly described herein, an aspect of the present disclosure relates to a pharmaceutical composition comprising a pharmaceutically acceptable excipient and a therapeutically effective amount of a monoclonal antibody or antigen-binding fragment thereof comprising a heavy chain variable region comprising an amino acid sequence at least about 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, or 100% identical to an amino acid sequence of SEQ ID NOS:l-29. In another aspect, the antibody light chain variable region comprises an amino acid sequence at least about 90, 91 , 92, 93, 94, 95, 96, 97, 98, 99, or 100% identical to an amino acid sequence of SEQ ID NOS:30-58. In alternative aspects, the antibody or antigen-binding fragment thereof comprises those heavy chain complementarity determining regions (CDR) 1 , CDR2, and CDR3 as set forth in any one of SEQ ID NOS 1-29 and / or those light chain CDR1, CDR2, and CDR3 as set forth in any one of SEQ ID NOS 30-58. In some forms there is 1, 2, or 3 mutations in in one, two or all three of the CDRs of any of SEQ ID NOS:l-58. In some forms there are no mutations in the CDRs relative to SEQ ID NOS:l-58. In one aspect, the antibody or antigen-binding fragment that binds to the SARS CoV-2 Spike protein. In another aspect, the antigen-binding fragment is a recombinant single-chain fragment variable (scFV) antibody, Fab fragment, F(ab’)2 fragment, or Fv fragment. In anotheraspect, the antibody or antigen-binding fragment is chimeric, humanized, fully human, bispecific, or multimeric. In another aspect, the antibody or antigen-binding fragment comprises an Fc portion mutated to at least one of: eliminate or enhance Fc Receptor (FcR) interactions to change a half-life, increase or decrease antibody-dependent cellular cytotoxicity, or increase or decrease complement activation. In another aspect, the antibody or binding fragment thereof of further comprising a peptide linker in a hinge region between the variable and constant domains of the heavy chain. In another aspect, the antibody or antigen-binding fragment is adapted for administration or genetic delivery with an RNA or DNA sequence or vector encoding the antibody or antigen-binding fragment. In another aspect, the antibody or antigen-binding fragment is formulated for nasal, pulmonary, alveolar, intravenous, administration.

[0009] As embodied and broadly described herein, an aspect of the present disclosure relates to a method of treating SARS-CoV-2 in a subject in need thereof, the method comprising administering to the subject a monoclonal antibody or antigen-binding fragment thereof comprising heavy chain variable region comprising an amino acid sequence at least about 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, or 100% identical to an amino acid sequence of SEQ ID NOS:l-29. In another aspect, the antibody light chain variable region comprises an amino acid sequence at least about 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, or 100% identical to an amino acid sequence of SEQ ID NOS:30-58. In alternative aspects, the antibody or antigen-binding fragment thereof comprises those heavy chain complementarity determining regions (CDR) 1 , CDR2, and CDR3 as set forth in any one of SEQ ID NOS 1-29 and / or those light chain CDR1, CDR2, and CDR3 as set forth in any one of SEQ ID NOS 30-58. In some forms there is 1, 2, or 3 mutations in in one, two or all three of the CDRs of any of SEQ ID NOS:l-58. In some forms there are no mutations in the CDRs relative to SEQ ID NOS: 1- 58. In one aspect, the antibody or antigen-binding fragment binds to a domain of SARS CoV-2 Spike protein. In another aspect, the antibody heavy chain In another aspect, the antigen-binding fragment is a recombinant single-chain fragment variable (scFV) antibody, Fab fragment, F(ab’)2 fragment, or Fv fragment. In another aspect, the antibody or antigen-binding fragment is chimeric, humanized, fully human, bispecific, or multimeric. In another aspect, the antibody or antigen-binding fragment comprises an Fc portion mutated to at least one of: eliminate or enhance Fc Receptor (FcR) interactions to change a half-life, increase or decrease antibody-dependent cellular cytotoxicity, or increase or decrease complement activation. In another aspect, the antibody or antigen-binding fragment is adapted for administration or genetic delivery with an RNA or DNA sequence or vector encoding the antibody or antigen-binding fragment.

[0010] As embodied and broadly described herein, an aspect of the present disclosure relates to a pharmaceutical device suitable for nasal or pulmonary delivery of monoclonal antibody or antigen-binding fragment thereof comprising heavy chain variable region comprising an amino acid sequence at least about 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, or 100% identical to an amino acid sequence of SEQ ID NOS: 1-29 and / or a light chain variable region comprising an amino acid sequence at least about 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, or 100% identical to an amino acid sequence of SEQ ID NOS:30-58, respectively, selected from an inhaler for liquids, a nebulizer, metered-dose inhaler, aerosol, and a dry powder inhaler. In one aspect, the antibody or antigen-binding fragment that binds to a domain of SARS CoV-2 Spike protein. In alternative aspects, the antibody or antigen-binding fragment thereof comprises those heavy chain complementarity determining regions (CDR)l, CDR2, and CDR3 as set forth in any one of SEQ ID NOS 1-29 and / or those light chain CDR1, CDR2, and CDR3 as set forth in any one of SEQ ID NOS 30-58. In some forms there is 1, 2, or 3 mutations in in one, two or all three of the CDRs of any of SEQ ID NOS:l-58. In some forms there are no mutations in the CDRs relative to SEQ ID NOS:l-58.

[0011] As embodied and broadly described herein, an aspect of the present disclosure relates to a method for detecting a Spike protein of SARS-CoV-2 (SARS2-S) comprising: obtaining or having obtained a biological sample suspected of comprising a SARS-CoV-2 virus; contacting the biological sample with a monoclonal antibody or antigen-binding fragment thereof comprising heavy chain heavy chain variable region comprising an amino acid sequence at least about 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, or 100% identical to an amino acid sequence of SEQ ID NOS:l-29 and / or an antibody light chain variable region comprising an amino acid sequence at least about 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, or 100% identical to an amino acid sequence of SEQ ID NOS:30-58, respectively; and detecting binding of the antibody or antigen-binding fragment to the SARS-CoV-2 virus. In alternative aspects, the antibody or antigen-binding fragment thereof comprises those heavy chain complementarity determining regions (CDR) 1 , CDR2, and CDR3 as set forth in any one of SEQ ID NOS 1-29 and / or those light chain CDR1, CDR2, and CDR3 as set forth in any one of SEQ ID NOS 30-58. In some forms there is 1, 2, or 3 mutations in in one, two or all three of the CDRs of any of SEQ ID NOS:l -58. In some forms there are no mutations in the CDRs relative to SEQ ID NOS: 1 - 58. In one aspect, the method further comprises the step of detecting the antibody or antigen -binding fragment is diagnostic for the detection of SARS-CoV-2 in the biological sample. In one aspect, the method further comprises detecting SARS-CoV-2 by performing an immunoassay on the biological sample from a subject; wherein the immunoassay uses an antibody or antigen-binding fragment comprising the monoclonal antibody or antigen-binding fragment thereof comprising heavy chain complementarity determining regions (CDR)l, CDR2, and CDR3, provided in In another aspect, the immunoassay is selected from radioimmunoassay, enzyme-linked immunosorbent assay (ELISA),sandwich assays, Western blot, immunoprecipitation, immunohistochemistry, immunofluorescence, antibody microarray, dot blotting, and fluorescence-activated cell sorting (FACS).

[0012] As embodied and broadly described herein, an aspect of the present disclosure relates to a kit for detection of a Spike protein of SARS-CoV-2 (SARS2-S) comprising a monoclonal antibody or antigen-binding fragment thereof comprising heavy chain complementarity determining regions (CDR)l , CDR2, and CDR3 as set forth in any one of SEQ ID NOS: 1-29, respectively; and light chain CDR1, CDR2, and CDR3 as set forth in any one of SEQ ID NOS:30-58, respectively, for isolation and / or detection of SARS-CoV-2 in one or more sample types of human or nonhuman origin, including: blood, plasma, serum, saliva, tears, cerebrospinal fluid, lymph, urine, feces, exhaled breath condensate, perspiration, amniotic fluid, exosomes, cell and tissue lysates. In alternative aspects, the antibody or antigen-binding fragment thereof comprises those heavy chain complementarity determining regions (CDR)l, CDR2, and CDR3 as set forth in any one of SEQ ID NOS 1-29 and / or those light chain CDR1 , CDR2, and CDR3 as set forth in any one of SEQ ID NOS 30-58. In some forms there is 1, 2, or 3 mutations in in one, two or all three of the CDRs of any of SEQ ID NOS:1- 58. In some forms there are no mutations in the CDRs relative to SEQ ID NOS:l-58.

[0013] As embodied and broadly described herein, an aspect of the present disclosure relates to a recombinant nucleic acid molecule encoding an antibody or antigen-binding fragment thereof encoding a monoclonal antibody or antigen-binding fragment thereof comprising heavy chain complementarity determining regions (CDR) 1 , CDR2, and CDR3 as set forth in any one of SEQ ID NOS: 1-29, respectively; and light chain CDR1, CDR2, and CDR3 as set forth in any one of SEQ ID NQS:30-58, respectively. In one aspect, the nucleic acid encoding the antigen-binding fragment thereof, wherein the antibody heavy chain variable region is encoded by a nucleic acid sequence having at least 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, or 100% identical to a nucleic acid of SEQ ID NOS: 1-29. In one aspect, the nucleic acid encoding the antigen-binding fragment thereof, wherein the antibody heavy chain variable region and light chain variable region is encoded by a nucleic acid sequence having at least 90, 91 , 92, 93, 94, 95, 96, 97, 98, 99, or 100% identical to a nucleic acid of SEQ ID NOS:30-58.

[0014] As embodied and broadly described herein, an aspect of the present disclosure relates to a recombinant expression vector comprising an expression control sequence operatively linked to the recombinant nucleic acid molecule encoding a monoclonal antibody or antigen-binding fragment thereof comprising heavy chain complementarity determining regions (CDR) 1 , CDR2, and CDR3 as set forth in any one of SEQ ID NOS:l-29; and light chain CDR1, CDR2, and CDR3 as set forth in any one of SEQ ID NOS:30-58, respectively.

[0015] As embodied and broadly described herein, an aspect of the present disclosure relates to a host cell comprising the recombinant nucleic acid molecule encoding a monoclonal antibody or antigen-binding fragment thereof comprising heavy chain complementarity determining regions (CDR)l, CDR2, and CDR3 as set forth in any one of SEQ ID NOS: 1-29; and light chain CDR1, CDR2, and CDR3 as set forth in any one of SEQ ID NOS:30-58, respectively.

[0016] As embodied and broadly described herein, an aspect of the present disclosure relates to a method of making an antibody or antigen-binding fragment of an antibody comprising culturing a recombinant host cell comprising a recombinant expression construct comprising an expression control sequence operatively linked to a recombinant nucleic acid molecule encoding a monoclonal antibody or binding fragment thereof comprising heavy chain complementarity determining regions (CDR)l, CDR2, and CDR3 as set forth in any one of SEQ ID NOS: l-29; and light chain CDR1, CDR2, and CDR3 as set forth in any one of SEQ ID NOS:3Q-58, respectively, wherein the host cell produces the antibody or antigen-binding fragment of an antibody, and isolating the antibody or antigen-binding fragment of an antibody from the recombinant host cell. In alternative aspects, the antibody or antigen-binding fragment thereof comprises those heavy chain complementarity determining regions (CDR)l, CDR2, and CDR3 as set forth in any one of SEQ ID NOS 1-29 and / or those light chain CDR1 , CDR2, and CDR3 as set forth in any one of SEQ ID NOS 30-58. In some forms there is 1, 2, or 3 mutations in in one, two or all three of the CDRs of any of SEQ ID NOS:1- 58. In some forms there are no mutations in the CDRs relative to SEQ ID NOS:l-58.BRIEF DESCRIPTION OF THE DRAWINGS

[0017] For a more complete understanding of the features and advantages of the present invention, reference is now made to the detailed description of the invention along with the accompanying figures and in which:

[0018] Figures 1A-1C show a table that summarizes the binding results of the antibodies of the present invention. 1A, Phylogenic trees representing Omicron- specific B cell clonal lineages from donor 4809. Tip sizes, number of cells with identical features; colors, cell type; labels, validated antibodies and ri by SPR; * Ari for BA.4 / 5 spike. Figure IB shows a summary of ELISA, PSV neutralization and SPR data for ancestral (D614G), Omicron BA.4 / 5 spike, RBD for antibodies in 1A. Figure IB shows binding of ancestral and Omicron; O, Omicron- specific. Grey, undetectable binding / neutralization. Figure 1C shows comparison of heavy chain mutations for donor 4809 RBD- and nucleocapsid-specific BGC. Student’s / -test,0.0001.

[0019] Figure 2 is a non-limiting example of ELISA IgG dilution curves for ancestral and Omicron BA.4 / 5 RBD for the antibodies disclosed herein (related to Figure IB). Mean and standard deviation (error bars) for three independent experiments are shown for controls and the mAbs in Figure IB.DETAILED DESCRIPTION OF THE INVENTIONI. DEFINITIONS

[0020] While the making and using of various embodiments of the present invention are discussed in detail below, it should be appreciated that the present invention provides many applicable inventive concepts that can be embodied in a wide variety of specific contexts. The specific embodiments discussed herein are merely illustrative of specific ways to make and use the invention and do not delimit the scope of the invention.

[0021] To facilitate the understanding of this invention, a number of terms are defined below. Terms defined herein have meanings as commonly understood by a person of ordinary skill in the areas relevant to the present invention. Terms such as “a”, “an” and “the” are not intended to refer to only a singular entity, but include the general class of which a specific example may be used for illustration. The terminology herein is used to describe specific embodiments of the invention, but their usage does not delimit the invention, except as outlined in the claims.

[0022] As used herein, the term “binds” in reference to the interaction of a binding protein and an antigen means that the interaction is dependent upon the presence of a particular structure (e.g., an antigenic determinant or epitope) on the antigen. For example, a binding protein recognizes and binds to a specific antigen structure rather than to antigens generally. For example, if a binding protein binds to epitope "A", the presence of a molecule containing epitope “A” (or free, unlabeled “A”), in a reaction containing labeled “A” and the binding protein, will reduce the amount of labeled “A” bound to the binding protein.

[0023] As used herein, a molecule is said to be able to “immunospecifically bind” a second molecule if such binding exhibits the specificity and affinity of an antibody to its cognate antigen. Antibodies are said to be capable of “immunospecifically binding” to a target region or conformation (“epitope”) of an antigen (and in particular, an antigen of SARS-CoV-2 such as the spike protein) if such binding involves the antigen recognition site of the immunoglobulin molecule. An antibody that immunospecifically binds to a particular antigen may bind to other antigens with lower affinity if the other antigen has some sequence or conformational similarity that is recognized by the antigen recognition site as determined by, e.g., immunoassays, BIACORE® assays, or other assays known in the art, but would not bind to a totally unrelated antigen. Preferably, however, antibodies (and their antigen binding fragments) will not cross-react with other antigens. Antibodies may also bind to other molecules in a way that is not immunospecific, such as to FcR receptors, by virtue of bindingdomains in other regions / domains of the molecule that do not involve the antigen recognition site, such as the Fc region.

[0024] The term “substantially,” as used in the context of binding or exhibited effect, is intended to denote that the observed effect is physiologically or therapeutically relevant. Similarly, a molecule is said to have substantially the same immunospecificity and / or characteristic as another molecule, if such immunospecificities and characteristics are greater than 60% identical, greater than 70% identical, greater than 75% identical, greater than 80% identical, greater than 85% identical, greater than 90% identical, greater than 95% identical, or greater than 97% identical).

[0025] The term “antibody” is used in the broadest sense unless clearly indicated otherwise. Therefore, an "antibody" can be naturally occurring or man-made such as monoclonal antibodies produced by conventional hybridoma technology. Antibodies include monoclonal and polyclonal antibodies as well as fragments and polymers containing the antigen binding domain and / or one or more complementarity determining regions of these antibodies. As used herein, the term “antibody” refers to an intact antibody or a binding fragment thereof that binds specifically to a target antigen. Binding fragments are produced by recombinant DNA techniques, or by enzymatic or chemical cleavage of intact antibodies. Binding fragments include Fab, Fab’, Fiab’ h, Fv, and single-chain variable fragment (scFv) antibodies. An antibody substantially inhibits adhesion of a receptor to a counterreceptor when an excess of antibody reduces the quantity of receptor bound to counterreceptor by at least about 20%, 40%, 60% or 80%, and more usually greater than about 85% (as measured in an in vitro competitive binding assay). The term “antibody” is used in the broadest sense, and encompasses monoclonal antibodies (including full-length antibodies or other bivalent, Fc-region containing antibodies such as bivalent scFv Fc-fusion antibodies), polyclonal antibodies, multispecific antibodies (e.g., bispecific antibodies), and antigen-binding fragments (e.g., Fab, Fab’, F(ab’)2, Fv, scFv) so long as they exhibit the desired biological activity. Antibodies (Abs) and immunoglobulins (Igs) are glycoproteins having the same structural characteristics. For example, antibodies described herein include monoclonal antibodies (and binding fragments thereof) that are recombinant, in other words, where the complementarity determining regions (CDRs) are genetically spliced into a human antibody backbone, often referred to as veneering an antibody. Thus, in some forms, the monoclonal antibody is a fully synthesized antibody. In some forms, the monoclonal antibodies (and binding fragments thereof) can be made in bacterial or eukaryotic cells, including plant cells. The term “antibody” also encompasses an immunoglobulin molecule that possesses a “variable region” antigen recognition site. Thus, the term "antibody" encompasses a molecule having at least one variable region from a light chain immunoglobulin molecule and at least one variable region from a heavy chain molecule that in combination form a specific binding site for the targetantigen. The term antibody includes monoclonal antibodies, multi- specific antibodies, human antibodies, humanized antibodies, synthetic antibodies, chimeric antibodies, camelized antibodies (See e.g., Muyldermans et al., 2001, Trends Biochem. Sci. 26:230; Nuttall et al., 2000, Cur. Pharm. Biotech. 1:253; Reichmann and Muyldermans, 1999, J. Immunol. Meth. 231:25; International Publication Nos. WO 94 / 04678 and WO 94 / 25591 ; U.S. Patent No. 6,005,079), single-chain Fvs (scFv) (see, e.g., see Pluckthun in The Pharmacology of Monoclonal Antibodies, vol. 113, Rosenburg and Moore eds. Springer-Verlag, New York, pp. 269-315 (1994)), single chain antibodies, disulfide- linked Fvs (sdFv), intrabodies, and anti-idiotypic (anti-Id) antibodies (including, e.g., anti-Id and anti- anti-Id antibodies to the disclosed antibodies). In particular, such antibodies include immunoglobulin molecules of any type (e.g., IgG, IgE, IgM, IgD, IgA and IgY), class (e.g., IgGl, IgG2, IgG3, IgG4, IgAl and IgA2) or subclass.

[0026] As used herein, the term “monoclonal antibody” refers to an antibody obtained from a population of substantially homogeneous antibodies, i.e., the individual antibodies comprising the population are identical except for possible naturally occurring mutations that may be present in minor amounts. Monoclonal antibodies are highly specific, being directed against a single antigenic site. Furthermore, in contrast to conventional (polyclonal) antibody preparations, which typically include different antibodies directed against different determinants (epitopes), each monoclonal antibody is directed against a single determinant on the antigen. In addition to their specificity, the monoclonal antibodies are advantageous in that they are synthesized by the hybridoma culture, uncontaminated by other immunoglobulins. The modifier “monoclonal” indicates the character of the antibody as being obtained from a substantially homogeneous population of antibodies, and is not to be construed as requiring production of the antibody by any particular method. For example, the monoclonal antibodies to be used in accordance with the presently disclosed and claimed invention may be made by the hybridoma method first described by Kohler and Milstein, Nature 256, 495 (1975), relevant portions incorporated herein by reference.

[0027] All monoclonal antibodies used in accordance with the presently disclosed and claimed invention will be either (1) the result of a deliberate immunization protocol, as described in more detail hereinbelow; or (2) the result of an immune response that results in the production of antibodies naturally in the course of a disease or infection.

[0028] The terms “full-length antibody”, “intact antibody” or “whole antibody” are used interchangeably to refer to an antibody in its substantially intact form, as opposed to an antigen binding fragment of an antibody. Specifically, whole antibodies include those with heavy and light chains including an Fc region. The constant domains may be wild-type sequence constant domains (e.g., human wild-type sequence constant domains) or amino acid sequence variants thereof.

[0029] A “chimeric antibody” is a molecule in which different portions of the antibody are derived from different immunoglobulin molecules such as antibodies having a variable region derived from a non-human antibody and a human immunoglobulin constant region. Methods for producing chimeric antibodies are known in the art. See e.g., Morrison, 1985, Science 229: 1202; Oi et al., 1986, BioTechniques 4:214; Gillies et al., 1989, J. Immunol. Methods 125:191-202; and U.S. Patent Nos. 6,311,415, 5,807,715, 4,816,567, and 4,816,397. Chimeric antibodies including one or more CDRs from a non-human species and framework regions from a human immunoglobulin molecule can be produced using a variety of techniques known in the art including, for example, CDR-grafting (EP 239,400; International Publication No. WO 91 / 09967; and U.S. Patent Nos. 5,225,539, 5,530,101, and 5,585,089), veneering or resurfacing (EP 592,106; EP 519,596; Padlan, 1991, Molecular Immunology 28(4 / 5):489-498; Studnicka et al., 1994, Protein Engineering 7:805; and Roguska et al., 1994, Proc. Natl. Acad. Sci. USA 91 :969), and chain shuffling (U.S. Patent No. 5,565,332).

[0030] As used herein, the term “humanized antibody” refers to an immunoglobulin including a human framework region and one or more CDR’s from a non-human (usually a mouse or rat) immunoglobulin. The non-human immunoglobulin providing the CDR’s is called the “donor” and the human immunoglobulin providing the framework is called the “acceptor.”

[0031] As used herein, the term “antigen binding fragment” of an antibody refers to one or more portions of an antibody that contain the antibody’s Complementarity Determining Regions (“CDRs”) and optionally the framework residues that include the antibody’s “variable region” antigen recognition site, and exhibit an ability to immunospecifically bind antigen. In some forms, the “antibody fragment” or “antigen-binding fragment” can encompass a portion of a full-length antibody, including the antigen-binding or variable region, and the Fab, Fab’, F(ab’)2, Fv, and scFv fragments. For example, Papain digestion of antibodies produces two identical antigen-binding fragments, called the Fab fragment, each with a single antigen-binding site, and a residual “Fc” fragment, so-called for its ability to crystallize readily. Pepsin treatment yields an F(ab’)2 fragment that has two antigen-binding fragments which are capable of cross-linking antigen, and a residual other fragment (which is termed pFc’). As used herein, “functional fragment” with respect to antibodies, includes Fv, Fab’, F(ab) and F(ab’)2 fragments, diabodies; linear antibodies; single-chain antibody molecules and multispecific antibodies formed from antibody fragments, and mutants thereof, naturally occurring variants, and fusion proteins including the antibody’s “variable region” antigen recognition site and a heterologous protein (e.g., a toxin, an antigen recognition site for a different antigen, an enzyme, a receptor or receptor ligand, etc.). For example, the term antigen binding fragment may be used to refer to recombinant single chain Fv fragments (scFv) as well asdivalent (di-scFv) and trivalent (tri-scFV) forms thereof. Such fragments can be produced via various methods known in the art.

[0032] As used herein, the “Fv” fragment is the minimum antigen-binding fragment that contains a complete antigen recognition and binding site. This region consists of a dimer of one heavy and one light chain variable domain in a tight, non-covalent association (VH-VL dimer). It is in this configuration that the three CDRs of each variable domain interact to define an antigen-binding site on the surface of the VH-VL dimer. Collectively, the six CDRs confer antigen-binding specificity to the antibody. However, even a single variable domain (or half of an Fv comprising only three CDRs specific for an antigen) has the ability to recognize and bind antigen, although at a lower affinity than the entire binding site.

[0033] The Fab fragment, also designated as F(ab), also contains the constant domain of the light chain and the first constant domain (CHI) of the heavy chain. Fab’ fragments differ from Fab fragments by the addition of a few residues at the carboxyl terminus of the heavy chain CHI 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 have a free thiol group. F(ab’) fragments are produced by cleavage of the disulfide bond at the hinge cysteines of the F(ab’ pepsin digestion product. Additional chemical couplings of antigen-binding fragment are known to those of ordinary skill in the art.

[0034] Native antibodies and immunoglobulins are usually heterotetrameric glycoproteins of about 150,000 daltons, composed of two identical light (L) chains and two identical heavy (H) chains. Each light chain is linked to a heavy chain by at least one covalent disulfide bond; however, the number of disulfide linkages varies between the heavy chains of different immunoglobulin isotypes. Each heavy and light chain also has regularly spaced intrachain disulfide bridges. Each heavy chain has at one end a variable domain (VH) followed by the constant domains. Each light chain has a variable domain at one end (VL) and a constant domain at its other end. The constant domain of the light chain is aligned with the first constant domain of the heavy chain, and the light chain variable domain is aligned with the variable domain of the heavy chain. Particular amino acid residues are believed to form an interface between the light and heavy chain variable domains (Clothia et al., J. Mol. Biol. 186, 651-66, 1985); Novotny and Haber, Proc. Natl. Acad. Sci. USA 82 4592-4596 (1985), relevant portions incorporated herein by reference.

[0035] As used herein, an “isolated” antibody is one that has been identified and separated and / or recovered from a component of the environment in which it was produced. Contaminant components of its production environment are materials, that would interfere with diagnostic or therapeutic uses for the antibody, and may include enzymes, hormones, and other proteinaceous or nonproteinaceoussolutes. In certain embodiments, the antibody will be purified as measurable by at least three different methods: 1) to greater than 50% by weight of antibody as determined by the Lowry method, such as more than 75% by weight, or more than 85% by weight, or more than 95% by weight, or more than 99% by weight; 2) to a degree sufficient to obtain at least 10 residues of N-terminal or internal amino acid sequence by use of a spinning cup sequenator, such as at least 15 residues of sequence; or 3) to homogeneity by SDS-PAGE under reducing or non-reducing conditions using Coomasie blue or, preferably, silver stain. Isolated antibody includes the antibody in situ within recombinant cells since at least one component of the antibody’s natural environment will not be present. Ordinarily, however, isolated antibody will be prepared by at least one purification step.

[0036] As used herein, the terms “antibody mutant” or “antibody variant” refer to an amino acid sequence variant of an antibody wherein one or more of the amino acid residues have been modified. Such mutants necessarily have less than 100% sequence identity or similarity with the amino acid sequence having at least 75% amino acid sequence identity or similarity with the amino acid sequence of either the heavy or light chain variable domain of the antibody, such as at least 80%, or at least 85%, or at least 90%, or at least 95, 96, 97, 98, or 99%.

[0037] The term “constant region” as used herein, refers to a portion of heavy chain or light chain of an antibody other than the variable region. In a heavy chain, the constant region generally includes a plurality of constant domains and a hinge region, e.g., an IgG constant region includes the following linked components, a constant heavy CHI, a linker, a CH2 and a CH3. In a heavy chain, a constant region includes a Fc. In a light chain, a constant region generally include one constant domain (a CL1).

[0038] The term “fragment crystalizable” or “Fc” or “Fc region” or “Fc portion” (which can be used interchangeably herein) refers to a region of an antibody including at least one constant domain and which is generally (though not necessarily) glycosylated and which is capable of binding to one or more Fc receptors and / or components of the complement cascade. The heavy chain constant region can be selected from any of the five isotypes: a, 5, £, y, or p. Exemplary heavy chain constant regions are gamma 1 (IgGl), gamma 2 (IgG2) and gamma 3 (IgG3), or hybrids thereof.

[0039] The term “variable region” is intended to distinguish such domain of the immunoglobulin from domains that are broadly shared by antibodies (such as an antibody Fc domain). The variable region refers to the portions of the light and / or heavy chains of an antibody as defined herein that specifically binds to an antigen and, for example, includes amino acid sequences of CDRs; i.e., CDR1, CDR2, and CDR3, and framework regions (FRs). For example, the variable region can include three or four FRs (e.g., FR1, FR2, FR3 and optionally FR4) together with three CDRs. VH refers to the variable region of the heavy chain. VL refers to the variable region of the light chain. The variableregion includes a “hypervariable region” whose residues are responsible for antigen binding. As used herein, the term “variable” in the context of the variable domain of antibodies, also encompasses certain portions of the variable domains differing extensively in sequence among antibodies and are used in the binding and specificity of each particular antibody for its particular antigen. However, the variability is not evenly distributed through the variable domains of antibodies. It is concentrated in three segments called complementarity determining regions (CDRs) also known as hypervariable regions both in the light chain and the heavy chain variable domains. The hypervariable region includes amino acid residues from a “Complementarity Determining Region” or “CDR” (e.g., typically at approximately residues 24-34 (LI), 50-56 (L2) and 89-97 (L3) in the light chain variable domain and at approximately residues 27-35 (Hl), 50-65 (H2) and 95-102 (H3) in the heavy chain variable domain according to Kabat; Kabat et al., Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, MD. (1991)) and / or those residues from a “hypervariable loop” (e.g., residues 26-32 (LI), 50-52 (L2) and 91-96 (L3) in the light chain variable domain and 26-32 (Hl), 53-55 (H2) and 96-101 (H3) in the heavy chain variable domain according to Chothia; Chothia and Lesk, 1987, 1. Mol. Biol. 196:901-917). Conventions that include corrections or alternate numbering systems for variable domains include not only Kabat and Chothia, but also IMGT (Lefranc, et al. (2003), Dev Comp Immunol 27: 55- 77), Chothia (Chothia C, Lesk AM (1987), J Mai Biol 196: 901-917; Chothia, et al. (1989), Nature 342: 877-883) and AHo (Honegger A, Pliickthun A (2001) J Mol Biol 309: 657-670). For convenience, examples of binding proteins of the present disclosure may also be labelled according to Kabat, Chothia, or IMGT. These examples are expressly indicated as such. There are at least two techniques for determining CDRs: (1) an approach based on cross-species sequence variability (i.e., Kabat et al., Sequences of Proteins of Immunological Interest (National Institute of Health, Bethesda, Md. 1987); and (2) an approach based on crystallographic studies of antigen-antibody complexes (Chothia, C. et al. (1989), Nature 342: 877), or both, that is Chothia plus Kabat. The more highly conserved portions of variable domains are called the framework (FR). The variable domains of native heavy and light chains each comprise four FR regions, largely adopting a P-sheet configuration, connected by three CDRs, which form loops connecting, and in some cases forming part of, the P-sheet structure. The CDRs in each chain are held together in close proximity by the FR regions and, with the CDRs from the other chain, contribute to the formation of the antigen-binding site of antibodies (see Kabat et al.). The constant domains are not involved directly in binding an antibody to its cognate antigen but exhibit various effector function, such as participation of the antibody in antibody-dependent cellular toxicity. A “constant domain” is a domain in an antibody the sequence of which is highly similar in antibodies / antibodies of the same type, e.g., IgG or IgM or IgE. A constant region of an antibodygenerally includes a plurality of constant domains, e.g., the constant region of y, a or 5 heavy chain include two constant domains.

[0040] A “constant domain” is a domain in an antibody the sequence of which is highly similar in antibodies / antibodies of the same type, e.g., IgG or IgM or IgE. A constant region of an antibody generally includes a plurality of constant domains, e.g., the constant region of y, a or 5 heavy chain include two constant domains.

[0041] The light chains of antibodies (immunoglobulin) from any vertebrate species can be assigned to one of two clearly distinct types, called kappa and lambda, based on the amino sequences of their constant domain. Depending on the amino acid sequences of the constant domain of their heavy chains, “immunoglobulins” can be assigned to different classes. There are at least five (5) major classes of immunoglobulins: IgA, IgD, IgE, IgG, and IgM, and several of these may be further divided into subclasses (isotypes), e.g., IgG-1, IgG-2, IgG-3, and IgG4; IgA-1 and IgA-2. The subunit structures and three-dimensional configurations of different classes of immunoglobulins are well known.

[0042] As used herein, the term “fusion protein” refers to a polypeptide formed by the joining of two or more polypeptides through a peptide bond formed between the amino terminus of one polypeptide and the carboxyl terminus of another polypeptide or through linking of one polypeptide to another through reactions between amino acid side chains (for example disulfide bonds between cysteine residues on each polypeptide). The fusion protein can be formed by the chemical coupling of the constituent polypeptides or it can be expressed as a single polypeptide from a nucleic acid sequence encoding the single contiguous fusion protein. Fusion proteins can be prepared using conventional techniques in molecular biology to join the two genes in frame into a single nucleic acid sequence, and then expressing the nucleic acid in an appropriate host cell under conditions in which the fusion protein is produced.

[0043] As used herein, the term “variant” refers to a polypeptide or polynucleotide that differs from a reference polypeptide or polynucleotide, but retains essential properties. A typical variant of a polypeptide differs in amino acid sequence from another, reference polypeptide. Generally, differences are limited so that the sequences of the reference polypeptide and the variant are closely similar overall and, in many regions, identical. A variant and reference polypeptide may differ in amino acid sequence by one or more modifications (e.g., substitutions, additions, and / or deletions). A substituted or inserted amino acid residue may or may not be one encoded by the genetic code. A variant of a polypeptide may be naturally occurring such as an allelic variant, or it may be a variant that is not known to occur naturally.

[0044] Modifications and changes can be made in the structure of the polypeptides of the in disclosure and still obtain a molecule having similar characteristics as the polypeptide (e.g., a conservative amino acid substitution). For example, certain amino acids can be substituted for other amino acids in a sequence without appreciable loss of activity. Because it is the interactive capacity and nature of a polypeptide that defines that polypeptide’s biological functional activity, certain amino acid sequence substitutions can be made in a polypeptide sequence and nevertheless obtain a polypeptide with like properties. In making such changes, the hydropathic index of amino acids can be considered. The importance of the hydropathic amino acid index in conferring interactive biologic function on a polypeptide is generally understood in the art. It is known that certain amino acids can be substituted for other amino acids having a similar hydropathic index or score and still result in a polypeptide with similar biological activity. Each amino acid has been assigned a hydropathic index on the basis of its hydrophobicity and charge characteristics. Those indices are: isoleucine (+4.5); valine (+4.2); leucine (+3.8); phenylalanine (+2.8); cysteine / cystine (+2.5); methionine (+1.9); alanine (+1.8); glycine (-0.4); threonine (-0.7); serine (-0.8); tryptophan (-0.9); tyrosine (-1.3); proline (-1.6); histidine (-3.2); glutamate (-3.5); glutamine (-3.5); aspartate (-3.5); asparagine (-3.5); lysine (- 3.9); and arginine (-4.5). It is believed that the relative hydropathic character of the amino acid determines the secondary structure of the resultant polypeptide, which in turn defines the interaction of the polypeptide with other molecules, such as enzymes, substrates, receptors, antibodies, antigens, and cofactors. It is known in the art that an amino acid can be substituted by another amino acid having a similar hydropathic index and still obtain a functionally equivalent polypeptide. In such changes, the substitution of amino acids whose hydropathic indices are within + 2 is preferred, those within + 1 are particularly preferred, and those within + 0.5 are even more particularly preferred.

[0045] Substitution of like amino acids can also be made on the basis of hydrophilicity, particularly where the biological functional equivalent polypeptide or peptide thereby created is intended for use in immunological embodiments. The following hydrophilicity values have been assigned to amino acid residues: arginine (+3.0); lysine (+3.0); aspartate (+3.0 + 1 ); glutamate (+3.0 + 1); serine (+0.3); asparagine (+0.2); glutamine (+0.2); glycine (0); proline (-0.5 + 1); threonine (-0.4); alanine (-0.5); histidine (-0.5); cysteine (-1.0); methionine (-1.3); valine (-1.5); leucine (-1.8); isoleucine (-1.8); tyrosine (-2.3); phenylalanine (-2.5); tryptophan (-3.4). It is understood that an amino acid can be substituted for another having a similar hydrophilicity value and still obtain a biologically equivalent, and in particular, an immunologically equivalent polypeptide. In such changes, the substitution of amino acids whose hydrophilicity values are within + 2 is preferred, those within + 1 are particularly preferred, and those within + 0.5 are even more particularly preferred.

[0046] As outlined above, amino acid substitutions are generally based on the relative similarity of the amino acid side-chain substituents, for example, their hydrophobicity, hydrophilicity, charge, size, and the like. Exemplary substitutions that take various of the foregoing characteristics into consideration are well known to those of skill in the art and include (original residue: exemplary substitution): (Ala: Gly, Ser), (Arg: Lys), (Asn: Gin, His), (Asp: Glu, Cys, Ser), (Gin: Asn), (Glu: Asp), (Gly: Ala), (His: Asn, Gin), (He: Leu, Vai), (Leu: He, Vai), (Lys: Arg), (Met: Leu, Tyr), (Ser: Thr), (Thr: Ser), (Tip: Tyr), (Tyr: Trp, Phe), and (Vai: He, Leu). Embodiments of this disclosure thus contemplate functional or biological equivalents of a polypeptide as set forth above. In particular, embodiments of the polypeptides can include variants having about 50%, 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98%, 99%, or more sequence identity to the polypeptide of interest.

[0047] " Percent (%) amino acid sequence identity" with respect to a reference polypeptide sequence is defined as the percentage of amino acid residues in a candidate sequence that are identical with the amino acid residues in the reference polypeptide sequence, after aligning the sequences and introducing gaps, if necessary, to achieve the maximum percent sequence identity, and not considering any conservative substitutions as part of the sequence identity. Alignment for purposes of determining percent amino acid sequence identity can be achieved in various ways that are within the skill of those practicing in the art, for instance, using publicly available computer software such as BLAST, BLAST-2, ALIGN or Megalign (DNASTAR) software. Those skilled in the art can determine appropriate parameters for aligning sequences, including any algorithms needed to achieve maximal alignment over the full length of the sequences being compared.

[0048] Disclosed are materials, compositions, and components that can be used for, can be used in conjunction with, can be used in preparation for, or are products of the disclosed method and compositions. These and other materials are disclosed herein, and it is understood that when combinations, subsets, interactions, groups, etc. of these materials are disclosed that while specific reference of each various individual and collective combinations and permutation of these compounds may not be explicitly disclosed, each is specifically contemplated and described herein. For example, if a ligand is disclosed and discussed and a number of modifications that can be made to a number of molecules including the ligand are discussed, each and every combination and permutation of ligand and the modifications that are possible are specifically contemplated unless specifically indicated to the contrary. Thus, if a class of molecules A, B, and C are disclosed as well as a class of molecules D, E, and F and an example of a combination molecule, A-D is disclosed, then even if each is not individually recited, each is individually and collectively contemplated. Thus, in this example, each of the combinations A-E, A-F, B-D, B-E, B-F, C-D, C-E, and C-F are specifically contemplated and should be considered disclosed from disclosure of A, B, and C; D, E, and F; and the examplecombination A-D. Likewise, any subset or combination of these is also specifically contemplated and disclosed. Thus, for example, the sub-group of A-E, B-F, and C-E are specifically contemplated and should be considered disclosed from disclosure of A, B, and C; D, E, and F; and the example combination A-D. Further, each of the materials, compositions, components, etc. contemplated and disclosed as above can also be specifically and independently included or excluded from any group, subgroup, list, set, etc. of such materials.

[0049] These concepts apply to all aspects of this application including, but not limited to, steps in methods of making and using the disclosed compositions. Thus, if there are a variety of additional steps that can be performed it is understood that each of these additional steps can be performed with any specific form or combination of forms of the disclosed methods, and that each such combination is specifically contemplated and should be considered disclosed.

[0050] All methods described herein can be performed in any suitable order unless otherwise indicated or otherwise clearly contradicted by context. The use of any and all examples, or exemplary language (e.g., “such as”) provided herein, is intended merely to better illuminate the forms and does not pose a limitation on the scope of the forms unless otherwise claimed. No language in the specification should be construed as indicating any non-claimed element as essential to the practice of the invention.

[0051] Recitation of ranges of values herein are merely intended to serve as a shorthand method of referring individually to each separate value falling within the range, unless otherwise indicated herein, and each separate value is incorporated into the specification as if it were individually recited herein.

[0052] Use of the term “about” is intended to describe values either above or below the stated value in a range of approx. + / - 10%; in other forms the values can range in value either above or below the stated value in a range of approx. + / - 5%; in other forms the values can range in value either above or below the stated value in a range of approx. + / - 2%; in other forms the values can range in value either above or below the stated value in a range of approx. + / - 1 %. The preceding ranges are intended to be made clear by context, and no further limitation is implied.II. COMPOSITIONS

[0053] Disclosed are antibodies and their antigen binding fragments and other molecules that are capable of immunospecifically binding to the receptor-binding domain (RBD) of the SARS-CoV-2 spike protein, including RBD variants from Omicron sublineages BA.4 / 5 (also referred to herein as SARs-COV-2 RBD-binding polypeptides). As discussed elsewhere herein in more details, in nonlimiting embodiments, such molecules can be, for example, a monoclonal antibody, a human antibody, a chimeric antibody or a humanized antibody, or a fragment thereof, and fusion proteinsformed from the heavy and light chain variable regions thereof. The antibodies and antigen binding fragments can be monospecific, bispecific, trispecific or multispecific.

[0054] Additionally provided are chimeric antigen receptors (CARs) including such antibodies and antigen binding fragments, and cells having the CARs.

[0055] Also provided are uses of such molecules and cells in the detection and treatment (e.g., prophylactic treatment and / or therapeutic treatment) of SARs-CoV-2.A. SARS-CoV-2 RBD-Binding Polypeptides

[0056] Polypeptides that selectively bind the receptor-binding domain (RBD) of the SARS-CoV-2 spike protein are provided. The SARS-CoV-2 RBD mediates attachment of the SARs-CoV-2 virus to the human ACE2 receptor, allowing viral entry into host cells. The RBD is located within the SI subunit of the ‘spike’ glycoprotein (SI -domain) and contains a receptor-binding motif responsible for direct interaction with human ACE2 receptor for viral entry. Typically, the S-domain is also the site of neutralizing antibodies. Variant forms of the RBD, including those found in Omicron sublineages BA.4 / 5, contain mutations that increase immune evasion and increase binding affinity to the ACE2 receptor.

[0057] Polypeptides, e.g., recombinant antibodies that bind specifically to SARS-CoV-2 RBD, including Omicron BA.4 / 5 variants were developed. The disclosed polypeptides, e.g., recombinant antibodies, are useful for detecting, neutralizing, or preventing SARS-CoV-2 infection. It was discovered that antigen- specific B cells isolated from the upper respiratory tract exhibit immune signatures that are distinct from those found in circulating immune compartments (Ramirez, et al., 2024, Nature, vol. 632, pgs. 630-636). Specifically, B cells from the nasopharyngeal mucosa, including tissue-resident memory B cells (BRM cells), were found to be enriched for class-switched isotypes (e.g., IgAl) and displayed elevated levels of somatic hypermutation, indicative of localized affinity maturation. Recombinant antibodies derived from clonal families that included BRM cells were synthesized based on B cell receptor (BCR) sequences obtained from these mucosal samples and demonstrated binding specificity to the receptor-binding domain (RBD) of SARS-CoV-2, including variant forms thereof. The experimental data and conclusions are described in detail in Ramirez, et al., 2024, “Immunological memory’ diversity in the human upper airway”, Nature, vol. 632, pgs. 630-636, which is specifically incorporated herein by reference in its entirety.

[0058] In some forms, the disclosed polypeptides e.g., recombinant monoclonal antibodies derived from B-cells, including BRM cells exhibit increased binding affinity and neutralizing activity against mutated SARS-CoV-2 variants, such as Omicron sublineages (e.g., BA.4 / 5), compared to earlier viral strains such as D614G. These polypeptides e.g., recombinant monoclonal antibodies, are disclosedfor use in a range of applications, including variant-sensitive diagnostic assays, therapeutic formulations, and prophylactic interventions targeting SARS-CoV-2 and its evolving variants.1. Exemplary SARS-CoV-2 RBD-Binding Polypeptides

[0059] As introduced above, the term antibody herein refers to natural or synthetic polypeptides that bind a target antigen. The term includes polyclonal and monoclonal antibodies, including intact antibodies and functional (e.g., antigen-binding) antibody fragments, including Fab fragments, F(ab')2 fragments, Fab' fragments, Fv fragments, recombinant IgG (rlgG) 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, and tetrabodies, tandem di-scFv, tandem tri-scFv. The term also encompasses intact or full-length antibodies, including antibodies of any class or subclass, including IgG and sub classes thereof, IgM, IgE, IgA, and IgD. Thus, although typically discussed in the context of IgG, the target antibody of the Ig-Fc-specific immunoglobulin variable domain can be IgM, IgE, IgA, or IgD.

[0060] Each of the 29 different IgG candidate clones were identified, namely B_01, B_02, B_03, B_04, B_05, B_06, B_07, B_08, B_09, B_10, O_01, O_02, O_03, O_04, O_05, O_06, O_07, O_08, O_09, O_10, O_l l, GL2_V01, GL2_V02, GL2_V03, GL2_V04, GL3_V01, GL3_V02, GL3_V03, and GL3_V04, respectively.

[0061] Amino acid sequences for variable region of the heavy and light chains are provided and can be used to prepare the antibodies disclosed herein, as well as the additional and alternative binder embodiments disclosed herein.

[0062] Provided are molecules or antibodies that bind to SARS-CoV-2 RBD, and wherein the antigen binding domain includes six CDRs, wherein the CDRs include at least one CDR of the CDRs of anti- SARS-CoV-2 RBD antibodies (referred to as “anti-SARS2 RBD antibodies): B_01 , B_02, B_03, B_04, B_05, B_06, B_07, B_08, B_09, B_10, O_01, O_02, O_03, O_04, O_05, O_06, O_07, O_08, O_09, O_10, O_l l, GL2_V01, GL2_V02, GL2_V03, GL2_V04, GL3_V01, GL3_V02, GL3_V03, and GL3_V04 with all remaining CDRs selected from:(i) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody B_01;(ii) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody B_02;(iii) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody B_03;(iv) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody B_04;(v) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody B_05;(vi) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody B_06;(vii) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody B_07;(viii) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody B_08;(ix) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody B_09;(x) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody B_10;(xi) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody O_01;(xii) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody O_02;(xiii) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody O_03;(xiv) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody O_04;(xv) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody O_05;(xvi) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody O_06;(xvii) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody O_07;(xviii) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody O_08;(xix) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody O_09;(xx) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody O_10;(xxi) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody O_11;(xxii) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody GL2_V01 ;(xxiii) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody GL2_V02;(xxiv) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody GL2_V03;(xxv) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody GL2_V04;(xxvi) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody GL3_V01 ;(xxvii) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody GL3_V02;(xxviii) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody GL3_V03; or(xxix) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody GL3_V04.

[0063] In some embodiments, the molecules or antibodies which binds to SARS-CoV-2 RBD, and wherein the antigen binding domain includes six CDRs, wherein the six CDRs are:(i) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody B_01;(ii) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody B_02;(iii) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody B_03;(iv) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody B_04;(v) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody B_05;(vi) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody B_06;(vii) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody B_07;(viii) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody B_08;(ix) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody B_09;(x) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody B_10;(xi) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody O_01;(xii) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody O_02;(xiii) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody O_03;(xiv) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody O_04;(xv) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody O_05;(xvi) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody O_06;(xvii) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody O_07; (xviii) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody O_08;(xix) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody O_09;(xx) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody O_10;(xxi) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody O_l l ;(xxii) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody GL2_V01 ; (xxiii) the three light chain and the three heavy chain CDRs of anti-S ARS2 RBD antibody GL2_V02;(xxiv) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody GL2_V03;(xxv) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody GL2_V04;(xxvi) the three light chain and the three heavy chain CDRs of anti-S ARS2 RBD antibody GL3_V01 ; (xxvii) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody GL3_V02; (xxviii) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody GL3_V03; or(xxix) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody GL3_V04.

[0064] Methods of determining CDR sequences are known in the art. To determine complementaritydetermining regions (CDRs) from heavy and light chain immunoglobulin sequences, one typically begins with the nucleotide or amino acid sequences of the variable regions (VH and VL) of the antibody. The CDRs are the hypervariable regions of these variable domains and are primarilyresponsible for antigen binding specificity. These regions are flanked by more conserved framework regions (FRs). The identification of CDRs can be performed using established numbering schemes such as Kabat, Chothia, IMGT, or AHO, which define CDR boundaries based on structural or sequence conservation principles. Each scheme may define slightly different positions for CDRs, particularly CDR-H3, which is the most variable in both length and sequence.

[0065] Several bioinformatics tools and software platforms can be used to annotate and extract CDRs from antibody sequences. For example, IMGT / V-QUEST (from the international ImMunoGeneTics information system®) is a widely used web-based tool that performs alignment of input VH and VL sequences against a curated database of germline immunoglobulin genes and outputs the positions of the CDRs according to the IMGT unique numbering system. Similarly, IgBlast (developed by NCBI) is another tool that aligns immunoglobulin sequences to germline V, D, and J genes and provides annotations including CDR definitions. For structural and modeling purposes, tools such as ABnum (part of the Abysis suite) or ANARCI (Antigen receptor Numbering And Receptor Classification) can assign canonical CDR positions using various numbering schemes and facilitate downstream analysis. These tools are suitable for high-throughput analysis or incorporation into antibody discovery and engineering pipelines.

[0066] In practical applications, once the CDRs are identified from input heavy and light chain sequences using one or more of these tools, researchers can isolate the amino acid residues that include CDR1, CDR2, and CDR3 from each chain. These sequences can then be analyzed for diversity, somatic hypermutation, affinity maturation, or grafted onto therapeutic scaffolds for antibody development.

[0067] In some embodiments, the CDRs in the light and heavy chains are in the same order and / or orientation as in B_01, B_02, B_03, B_04, B_05, B_06, B_07, B_08, B_09, B_10, O_01, O_02, O_03, O_04, O_05, O_06, O_07, O_08, O_09, O_10, O_l l, GL2_V01, GL2_V02, GL2_V03, GL2_V04, GL3_V01, GL3_V02, GL3_VO3, and GL3_V04.

[0068] In some embodiments, the antibody binds to SARS-CoV-2 RBD and includes one or both of the light and heavy chain variable domain(s) of clone B_01, B_02, B_03, B_04, B_05, B_06, B_07, B_08, B_09, B_10, O_01, O_02, O_03, O_04, O_05, O_06, O_07, O_08, O_09, O_10, O_l l, GL2_V01, GL2_V02, GL2_V03, GL2_V04, GL3_V01, GL3_V02, GL3_V03, and GL3_V04.

[0069] The amino acid sequences for the heavy chain variable regions of B_01, B_02, B_03, B_04, B_05, B_06, B_07, B_08, B_09, B_10, O_01, O_02, O_03, O_04, O_05, O_06, O_07, O_08, O_09, O_10, O_l l, GL2_V01, GL2_V02, GL2_V03, GL2_V04, GL3_V01, GL3_V02, GL3_V03, and GL3_V04 are provided in Table 1 as SEQ ID NOS: 1-29.

[0070] Table 1: Heavy Chain Amino Acid and Nucleic Acid Sequences:

[0071] The amino acid sequences for the heavy chain variable regions of B_01, B_02, B_03, B_04, B_05, B_06, B_07, B_08, B_09, B_10, O_01, O_02, O_03, O_04, O_05, O_06, O_07, O_08, O_09, O_10, O_ll, GL2_V01, GL2_V02, GL2_V03, GL2_V04, GL3_V01, GL3_V02, GL3_V03, and GL3_V04 are provided in Table 2 as SEQ ID NOS:30-58.

[0072] Table 2: Light Chain Amino Acid and Nucleic Acid Sequences

[0073] Variants of SEQ ID NOS:l-58 that bind SARS-CoV-2 RBD and have at least 70% and up to 99% sequence identity to any of SEQ ID NOS:l-58 are also provided, and can be used as the heavy (i.e., variants of SEQ ID NOS: 1-29) and / or light (i.e., variants of SEQ ID NOS:30-58) chain variable regions of the disclosed antigen binding domains (i.e., in place the correspondence sequence of SEQ ID NOS:l-58). For example, in some forms, the variant sequence has at least about 70%, 75%, 80%, 85%, 90%, or 95% identity to any one of SEQ ID NOS:l-58. Therefore, in some forms, the amino acid sequence has one or more amino acids different, such as one or more substitutions, deletions or additions at any one of the amino acid positions of its corresponding sequence of SEQ ID NOS:l-58. In some forms, variants of SEQ ID NOS: 1-58 include the CDRs of SEQ ID NOS: 1-58, respectively (i.e., without variation).

[0074] As embodied and broadly described herein, an aspect of the present disclosure relates to a monoclonal antibody or antigen-binding fragment thereof antibody contain: a heavy chain variable region including an amino acid sequence at least about 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, or 100% identical to an amino acid sequence of SEQ ID NOS: 1-29. In another aspect, the antibody light chain variable region includes an amino acid sequence at least about 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, or 100% identical to an amino acid sequence of SEQ ID NOS:30-58. In alternative aspects, the antibody or antigen-binding fragment thereof includes those heavy chain complementaritydetermining regions (CDR)l, CDR2, and CDR3 as set forth in any one of SEQ ID NOS 1-29 and / or those light chain CDR1, CDR2, and CDR3 as set forth in any one of SEQ ID NOS 30-58. Thus, in some forms, there is no variation in the CDRs. In some forms, the antibodies or antigen binding fragments thereof include one, two, three, four, or five mutations in one or more CDRs. In another aspect, the antigen-binding fragment is a recombinant single-chain fragment variable (scFV) antibody, Fab fragment, F(ab’)2 fragment, or Fv fragment. In another aspect, the e antibody or antigen-binding fragment is chimeric, humanized, fully human, bispecific, or multimeric. In another aspect, the antibody or antigen-binding fragment includes an Fc portion mutated to at least one of: eliminate or enhance Fc Receptor (FcR) interactions to change a half-life, increase or decrease antibody-dependent cellular cytotoxicity, or increase or decrease complement activation. In another aspect, the antibody or antigen-binding fragment is adapted for administration or genetic delivery with an RNA or DNA sequence or vector encoding the antibody or antigen-binding fragment. In another aspect, the monoclonal antibody further includes a peptide linker in a hinge region between the variable and constant domains of the heavy chain2. Chimeric and Humanized Antibodies

[0075] The disclosure particularly concerns chimeric and humanized antibodies. Constant regions need not be present, but if they are, are typically substantially identical to human immunoglobulin constant regions, i.e., at least about 85-90%, preferably about 95% or more identical. Hence, all parts of a humanized immunoglobulin, except possibly the CDR’s, are substantially identical to corresponding parts of natural human immunoglobulin sequences. A humanized antibody is an antibody having a humanized light chain and a humanized heavy chain immunoglobulin. For example, a humanized antibody would not encompass a typical chimeric antibody, because, e.g., the entire variable region of a chimeric antibody is non-human. One says that the donor antibody has been “humanized,” by the process of “humanization,” because the resultant humanized antibody is expected to bind to the same antigen as the donor antibody that provides the CDR’s.

[0076] The uses of the monoclonal antibodies of the presently disclosed may require administration of such or similar monoclonal antibody to a subject, such as a human. However, when the monoclonal antibodies are produced in a non-human animal, such as a rodent or chicken, administration of such antibodies to a human patient will normally elicit an immune response, wherein the immune response is directed towards the antibodies themselves. Such reactions limit the duration and effectiveness of such a therapy. In order to overcome such a problem, the monoclonal antibodies of the presently disclosed and claimed invention can be “humanized”, that is, the antibodies are engineered such that antigenic portions thereof are removed and like portions of a human antibody are substituted, while the antibodies’ affinity for the SARS CoV-2 Spike protein from different variants is retained. Thisengineering may only involve a few amino acids or may include entire framework regions of the antibody, leaving only the complementarity determining regions of the antibody intact. Several methods of humanizing antibodies are known in the art and are disclosed in U.S. Pat. No. 6,180,370, issued to Queen et al on Jan. 30, 2001; U.S. Pat. No. 6,054,927, issued to Brickell on Apr. 25, 2000; U.S. Pat. No. 5,869,619, issued to Studnicka on Feb. 9, 1999; U.S. Pat. No. 5,861,155, issued to Lin on Jan. 19, 1999; U.S. Pat. No. 5,712,120, issued to Rodriquez et al on Jan. 27, 1998; and U.S. Pat. No. 4,816,567, issued to Cabilly et al on Mar. 28, 1989, relevant portions incorporated herein by reference.

[0077] Humanized forms of antibodies are chimeric immunoglobulins, immunoglobulin chains or fragments thereof (such as Fab, Fab’, F(ab’)2, Fv, scFv or other antigen-binding subsequences of antibodies) that are principally comprised of the sequence of a human immunoglobulin, and contain minimal sequence derived from a non-human immunoglobulin. Humanization can be performed following the method of Winter and co-workers (Jones et al., 1986; Riechmann et al., 1988; Verhoeyen et al., 1988), by substituting nonhuman (i.e., rodent, chicken) CDRs or CDR sequences for the corresponding sequences of a human antibody, see, e.g., U.S. Pat. No. 5,225,539. In some instances, Fvframework residues of the human immunoglobulin are replaced by corresponding non- human residues from the donor antibody. Humanized antibodies can also comprise residues that are found neither in the recipient antibody nor in the imported CDR or framework sequences. In general, the humanized antibody will comprise substantially all of, at least one, and typically two, variable domains, in which all or substantially all of the CDR regions correspond to those of a non-human immunoglobulin and all or substantially all of the framework regions are those of a human immunoglobulin consensus sequence. The humanized antibody optionally also will include at least a portion of an immunoglobulin constant region (Fc), typically that of a human immunoglobulin that immunospecifically binds to an FcyRIIB polypeptide, that has been altered by the introduction of amino acid residue substitutions, deletions or additions (i.e., mutations). See also, e.g., European Patent Nos. EP 239,400, EP 592,106, and EP 519,596; International Publication Nos. WO 91 / 09967 and WO 93 / 17105; U.S. Patent Nos. 5,225,539, 5,530,101, 5,565,332, 5,585,089, 5,766,886, and 6,407,213; and Padlan, 1991, Molecular Immunology 28(4 / 5):489498; Studnicka et al., 1994, Protein Engineering 7(6):805-814; Roguska et al., 1994, PNAS 91:969 973; Tan et al., 2002, J. Immunol. 169:1119-1125; Caldas et al., 2000, Protein Eng. 13:353-360; Morea et al., 2000, Methods 20:267 79; Baca et al., 1997, J. Biol. Chem. 272:10678 10684; Roguska et al., 1996, Protein Eng. 9:895 904; Couto et al., 1995, Cancer Res. 55 (23 Supp):5973s 5977s; Couto et al., 1995, Cancer Res. 55:1717 22; Sandhu, 1994, Gene 150:409 10; Pedersen et al., 1994, J. Mol. Biol. 235:959-973; Jones et al.,1986, Nature 321 :522-525; Reichmann et al., 1988, Nature 332:323-329; and Presta, 1992, Curr. Op. Struct. Biol. 2:593-596).

[0078] DNA sequences coding for preferred human acceptor framework sequences include but are not limited to FR segments from the human germline VH segment VH1-18 and JH6 and the human germline VL segment VK-A26 and JK4. In a specific embodiment, one or more of the CDRs are inserted within framework regions using routine recombinant DNA techniques. The framework regions can be naturally occurring or consensus framework regions, and preferably human framework regions (see, e.g., Chothia et al., 1998, “Structural Determinants In The Sequences Of Immunoglobulin Variable Domain J. Mol. Biol. 278: 457-479 for a listing of human framework regions).

[0079] A humanized or chimeric antibodies can include substantially all of at least one, and typically two, variable domains in which all or substantially all of the CDR regions correspond to those of a non-human immunoglobulin (i.e., donor antibody) and all or substantially all of the framework regions are those of a human immunoglobulin consensus sequence. Preferably, the antibody also includes at least a portion of an immunoglobulin constant region (Fc), typically that of a human immunoglobulin. The constant domains of the antibodies can be selected with respect to the proposed function of the antibody, in particular the effector function which can be required. In some embodiments, the constant domains of the antibodies are (or include) human IgA, IgD, IgE, IgG or IgM domains. The disclosure encompasses Fc constant domains including one or more amino acid modifications which alter antibody effector functions such as those disclosed in U.S. Patent Application Publication Nos. 2005 / 0037000 and 2005 / 0064514.

[0080] In some embodiments, the antibody contains both the light chain as well as at least the variable domain of a heavy chain. In other embodiments, the antibody can further include one or more of the CHI, hinge, CH2, CH3, and CH4 regions of the heavy chain. The antibody can be selected from any class of immunoglobulins, including IgM, IgG, IgD, IgA and IgE, and any isotype, including IgGl, IgG2, IgG3 and IgG4. The antibody can include sequences from more than one class or isotype, and selecting particular constant domains to optimize desired effector functions is within the ordinary skill in the art. In some embodiments, the antibody is not a mouse IgGl or a mouse IgG2a.

[0081] The framework and CDR regions of a humanized antibody need not correspond precisely to the parental sequences, e.g., the donor CDR or the consensus framework can be mutagenized by substitution, insertion or deletion of at least one residue so that the CDR or framework residue at that site does not correspond to either the consensus or the donor antibody. Such mutations, however, are preferably not extensive. Usually, at least 75% of the humanized antibody residues will correspond to those of the parental framework region (FR) and CDR sequences, more often 90%, and mostpreferably greater than 95%. Humanized antibodies can be produced using variety of techniques known in the art, including, but not limited to, CDR-grafting (European Patent No. EP 239,400; International Publication No. WO 91 / 09967; and U.S. Patent Nos. 5,225,539, 5,530,101, and 5,585,089), veneering or resurfacing (European Patent Nos. EP 592,106 and EP 519,596; Padlan, 1991, Molecular Immunology 28(4 / 5):489-498; Studnicka et al., 1994, Protein Engineering 7(6):805-814; and Roguska et al., 1994, Proc. Natl. Acad. Sci. 91:969-973), chain shuffling (U.S. Patent No. 5,565,332), and techniques disclosed in, e.g., U.S. Patent Nos. 6,407,213, 5,766,886, 5,585,089, International Publication No. WO 9317105, Tan et al., 2002, J. Immunol. 169:11 19-25, Caldas et al., 2000, Protein Eng. 13:353-60, Morea et al., 2000, Methods 20:267-79, Baca et al., 1997, J. Biol. Chem. 272:10678-84, Roguska et al., 1996, Protein Eng. 9:895-904, Couto et al., 1995, Cancer Res. 55 (23 Supp):5973s-5977s, Couto et al., 1995, Cancer Res. 55: 1717-22, Sandhu, 1994, Gene 150:409-10, Pedersen etal., 1994, J. Mol. Biol. 235:959-73, Jones et al., 1986, Nature 321 :522- 525, Riechmann et al., 1988, Nature 332:323, and Presta, 1992, Curr. Op. Struct. Biol. 2:593-596. Often, framework residues in the framework regions will be substituted with the corresponding residue from the CDR donor antibody to alter, preferably improve, antigen binding. These framework substitutions are identified by methods well known in the art, e.g., by modeling of the interactions of the CDR and framework residues to identify framework residues important for antigen binding and sequence comparison to identify unusual framework residues at particular positions. (See, e.g. , Queen et al., U.S. Patent No. 5,585,089; U.S. Publication Nos. 2004 / 0049014 and 2003 / 0229208; U.S. Patent Nos. 6,350,861 ; 6,180,370; 5,693,762; 5,693,761; 5,585,089; and 5,530,101 and Riechmann et al., 1988, Nature 332:323).3. Bispecific and Multispecific Antibodies

[0082] The antibodies used in the methods of the present disclosure can be monospecific. Antibodies monospecific for SARS-CoV-2 RBD can have a targeting moiety conjugated or otherwise linked thereto. In some embodiments, the targeting moiety is an antibody or antigen binding fragment thereof. Thus, in some embodiments, the anti-SARS-CoV-2 RBD antibody or antigen binding fragment is a bispecific, trispecific or multispecific antibody that includes a second (or third or more) antigen binding fragment that binds to a cell specific antigen. Thus provided are bispecific, trispecific or multispecific antibodies having one or more antigen binding fragments that binds to anti-SARS- CoV-2 RBD and a second (third or more) antigen binding fragment that binds to a cell specific antigen. For example, such antibodies can bind to both SARS-CoV-2 RBD and to an antigen that is important for targeting the antibody to a particular cell type or tissue.

[0083] In some embodiments, the antibodies are heterodimeric bi- and tri- (or more) specific Ig antibodies and Fc fusion proteins. Exemplary structures include, but are not limited to, IgG, IgM,mono-, di-, tri-, or more scFv-Fcs. For example, bispecific, trispecific, and multispecific formats include, but are not limited to, bispecific and trispecific IgG, IgG-scFv, IgG-dAb, scFv-Fc-scFv, knob-in-hole (KIH)-IgG, K / .-body, KIHOFc-Fab / scFv, tandem scFv, KIH trispecific, bispecific Fc fusion (N- or C -terminal, with or without KIH).

[0084] In embodiments, multispecific antibody molecules can include more than one antigen-binding site, where different sites are specific for different antigens. In embodiments, multispecific antibody molecules can bind more than one (e.g., two or more) epitopes on the same antigen. In embodiments, multispecific antibody molecules include an antigen-binding site specific for a target cell and a different antigen-binding site specific for SARS-CoV-2 RBD. In some embodiments, the multispecific antibody molecule is a bispecific antibody molecule. Bispecific antibody molecules can be classified into five different structural groups: (i) bispecific immunoglobulin G (BsIgG); (ii) IgG appended with an additional antigen-binding moiety; (iii) bispecific antibody fragments; (iv) bispecific fusion proteins; and (v) bispecific antibody conjugates.

[0085] BsIgG is a format that is monovalent for each antigen. Exemplary BsIgG formats include but are not limited to crossMab, DAF (two-in-one), DAF (four-in-one), DutaMab, DT-IgG, knobs-in- holes common LC, knobs-in-holes assembly, charge pair, Fab-arm exchange, SEEDbody, triomab, LUZ-Y, Fcab, kappa-lamda-body, orthogonal Fab. See Spiess et al. Mol. Immunol. 67(2015):95- 106. Exemplary BsIgGs include catumaxomab (Fresenius Biotech, Trion Pharma, Neopharm), which contains an anti-CD3 arm and an anti-EpCAM arm; and ertumaxomab (Neovii Biotech, Fresenius Biotech), which targets CD3 and HER2.

[0086] In some embodiments, BsIgG includes heavy chains that are engineered for heterodimerization. For example, heavy chains can be engineered for heterodimerization using a "knobs-into-holes" strategy, a SEED platform, a common heavy chain (e.g., in Kk-bodies), and use of heterodimeric Fc regions. See Spiess et al., Mol. Immunol. 67(2015):95- 106. Strategies that have been used to avoid heavy chain pairing of homodimers in BsIgG include knobs-in-holes, duobody, azymetric, charge pair, HA-TF, SEEDbody, and differential protein A affinity. See Id. BsIgG can be produced by separate expression of the component antibodies in different host cells and subsequent purification / assembly into a BsIgG. BsIgG can also be produced by expression of the component antibodies in a single host cell. BsIgG can be purified using affinity chromatography, e.g., using protein A and sequential pH elution.

[0087] IgG appended with an additional antigen-binding moiety is another format of bispecific antibody molecules. For example, monospecific IgG can be engineered to have bispecificity by appending an additional antigen-binding unit onto the monospecific IgG, e.g., at the N- or C-terminus of either the heavy or light chain. Exemplary additional antigen-binding units include single domainantibodies (e.g., variable heavy chain or variable light chain), engineered protein scaffolds, and paired antibody variable regions (e.g., single chain variable fragments or variable fragments). See Id. Examples of appended IgG formats include dual variable domain (DVD) IgG (DVD-Ig), 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). See Spiess et al. Mol. Immunol. 67(2015):95-106. An example of an IgG-scFv is MM-141 (Merrimack Pharmaceuticals), which binds IGF-1R and HER3. Examples ofDVD-Ig include ABT-981 (AbbVie), which binds IL-lalpha and IL-lbeta; and ABT-122 (AbbVie), which binds TNF and IL-17A.

[0088] Bispecific antibody fragments (BsAb) are a format of bispecific antibody molecules that lack some or all of the antibody constant domains. For example, some BsAb lack an Fc region. In some embodiments, bispecific antibody fragments include heavy and light chain regions that are connected by a peptide linker that permits efficient expression of the BsAb in a single host cell. Exemplary bispecific antibody fragments include but are not limited to nanobody, nanobody-HAS, BiTE, Diabody, DART, TandAb, scDiabody, scDiabody-CH3, Diabody-CH3, triple body, miniantibody, minibody, TriBi minibody, scFv-CH3 KIH, Fab-scFv, scFv-CH-CL-scFv, F(ab’)2, F(ab')2-scFv2, scFv-KIH, Fab-scFv-Fc, tetravalent HCAb, scDiabody-Fc, Diabody-Fc, tandem scFv-Fc, and intrabody. See Id. For example, the BiTE format includes tandem scFvs, where the component scFvs bind to CD3 on T cells.

[0089] Bispecific fusion proteins include antibody fragments linked to other proteins, e.g., to add additional specificity and / or functionality. An example of a bispecific fusion protein is an immTAC, which includes an anti-CD3 scFv linked to an affinity-matured T-cell receptor that recognizes HLA- presented peptides. In embodiments, the dock-and-lock (DNL) method can be used to generate bispecific antibody molecules with higher valency. Also, fusions to albumin binding proteins or human serum albumin can be extend the serum half-life of antibody fragments. See Id.

[0090] In embodiments, chemical conjugation, e.g., chemical conjugation of antibodies and / or antibody fragments, can be used to create BsAb molecules. See Id. An exemplary bispecific antibody conjugate includes the CovX-body format, in which a low molecular weight drug is conjugated site- specifically to a single reactive lysine in each Fab arm or an antibody or fragment thereof. In embodiments, the conjugation improves the serum half-life of the low molecular weight drug. An exemplary CovX-body is CVX-241 (NCT01004822), which includes an antibody conjugated to two short peptides inhibiting either VEGF or Ang2. See Id.

[0091] In some embodiments the multispecific molecule further includes a heavy chain constant region (e.g., an Fc region) chosen from the heavy chain constant regions of IgGl, IgG2, and IgG4, more particularly, the heavy chain constant region of human IgGl, IgG2 or IgG4. In someembodiments, the heavy chain constant region (e.g., an Fc region) is linked to, e.g., covalently linked to, one or both of the SARS-CoV-2 RBD binding antibody molecule and the second antibody molecule.

[0092] In some embodiments, the heavy chain constant region (e.g., an Fc region) is altered, e.g., mutated, to increase or decrease one or more of: Fc receptor binding, antibody glycosylation, the number of cysteine residues, effector cell function, or complement function. In some embodiments, an interface of a first and second heavy chain constant regions (e.g., Fc region) is altered, e.g., mutated, to increase or decrease dimerization, e.g., relative to a non-engineered interface. In some embodiments, the dimerization of the heavy chain constant region (e.g., Fc region) is enhanced by providing an Fc interface of a first and a second Fc region with one or more of: a paired cavityprotuberance ("knob-in-a hole"), an electrostatic interaction, or a strand-exchange, such that a greater ratio of heteromultimerhomomultimer forms, e.g., relative to a non-engineered interface. In some embodiments, the heavy chain constant region (e.g., Fc region) includes an amino acid substitution at a position chosen from one or more of 347, 349, 350, 351, 366, 368, 370, 392, 394, 395, 397, 398, 399, 405, 407, or 409, e.g., of the Fc region of human IgGl, numbered based on the Eu numbering system. Unless otherwise specified herein, numbering of amino acid residues in the Fc region or constant region is according to the EU numbering system, also called the EU index, as described in Kabat et al., Sequences of Proteins of Immunological Interest, 5 th Ed. Public Health Service, National Institutes of Health, Bethesda, Md., 1991.

[0093] In some embodiments, the heavy chain constant region (e.g., Fc region) includes an amino acid substitution chosen from: T366S, L368A, or Y407V (e.g., corresponding to a cavity or hole), or T366W (e.g., corresponding to a protuberance or knob), or a combination thereof, numbered based on the Eu numbering system.

[0094] In some embodiments, the heavy chain constant region (e.g., an Fc region) includes one or more mutations that increase or decrease one or more of: Fc receptor binding, antibody glycosylation, the number of cysteine residues, effector cell function, or complement function, relative to a naturally-existing heavy chain constant region.

[0095] In some embodiments, the SARS-CoV-2 RBD binding molecule includes a first heavy chain constant region (e.g., a first Fc region) and the second antibody molecule includes a second heavy chain constant region (e.g., a second Fc region), wherein the first heavy chain constant region includes one or more mutations that increase heterodimerization of the first heavy chain constant region and the second heavy chain constant region, relative to a naturally-existing heavy chain constant region, and / or wherein the second heavy chain constant region includes one or more mutations that increase heterodimerization of the second heavy chain constant region and the first heavy chain constantregion, relative to a naturally-existing heavy chain constant region. In some embodiments, the first and the second heavy chain constant regions (e.g., first and second Fc regions) include one or more of: a paired cavity-protuberance ("knob-in-a hole"), an electrostatic interaction, or a strand-exchange, such that a greater ratio of heteromultimer:homomultimer forms, e.g., relative to naturally-existing heavy chain constant regions.

[0096] In some embodiments, the first and / or second heavy chain constant region (e.g., a first and / or second Fc region, e.g., a first and / or second IgGl Fc region) includes an amino acid substitution at a position chosen from one or more of 347, 349, 350, 351, 366, 368, 370, 392, 394, 395, 397, 398, 399, 405, 407, or 409, numbered based on the Eu numbering system. In some embodiments, the first and / or second heavy chain constant region (e.g., a first and / or second Fc region, e.g., a first and / or second IgGl Fc region) includes an amino acid substitution chosen from: T366S, L368A, Y407V, or Y349C (e.g., corresponding to a cavity or hole), or T366W or S354C (e.g., corresponding to a protuberance or knob), or a combination thereof, numbered based on the Eu numbering system.

[0097] In some embodiments, the multispecific molecule further includes a linker, e.g., a linker between one or more of: the SARS-CoV-2 RBD binding molecule and the second antibody molecule, the SARS-CoV-2 RBD binding antibody molecule and the heavy chain constant region (e.g., the Fc region), or the second antibody molecule and the heavy chain constant region. In some embodiments, the linker is chosen from: a cleavable linker, a non-cleavable linker, a peptide linker, a flexible linker, a rigid linker, a helical linker, or a non-helical linker. In some embodiments, the linker is a peptide linker. In some embodiments, the peptide linker includes Gly and Ser.4. Derivatives and Conjugates

[0098] The disclosure particularly contemplates the production and use of derivatives of any of the above-described antibodies and their antigen-binding fragments. The term derivative encompasses an antibody or antigen-binding fragment thereof that immunospecifically binds to an antigen but which includes, one, two, three, four, five or more amino acid substitutions, additions, deletions or modifications relative to a “parental” (or wild-type) molecule (also referred to as variants). Such amino acid substitutions or additions can introduce naturally occurring (i.e., DNA-encoded) or non- naturally occurring amino acid residues.

[0099] The term derivative also encompasses, for example, chimeric or humanized variants of any of the disclosed antibodies, as well as variants having altered CHI, hinge, CH2, CH3 or CH4 regions, so as to form, for example antibodies, etc., having variant Fc regions that exhibit enhanced or impaired effector or binding characteristics.

[0100] The term derivative additionally encompasses non-amino acid modifications, for example, amino acids that may be glycosylated (e.g., have altered mannose, 2-N-acetylglucosamine, galactose,fucose, glucose, sialic acid, 5 -N- acetylneuraminic acid, 5 -glycolneuraminic acid, etc. content), acetylated, pegylated, phosphorylated, amidated, derivatized by known protecting / blocking groups, proteolytic cleavage, linked to a cellular ligand or other protein, etc. In some embodiments, the altered carbohydrate modifications modulate one or more of the following: solubilization of the antibody, facilitation of subcellular transport and secretion of the antibody, promotion of antibody assembly, conformational integrity, and antibody -mediated effector function. In a specific embodiment the altered carbohydrate modifications enhance antibody mediated effector function relative to the antibody lacking the carbohydrate modification. Carbohydrate modifications that lead to altered antibody mediated effector function are well known in the art (for example, see Shields,R.L. et al. (2002) “Lack Of Fucose On Human IgG N-Linked Oligosaccharide Improves Binding To Human Fcgamma RIII And Antibody -Dependent Cellular Toxicity.,” J. Biol. Chem. 277(30): 26733- 26740; Davies J. et al. (2001) “Expression Of GnTIII In A Recombinant Anti-CD20 CHO Production Cell Line: Expression Of Antibodies With Altered Glycoforms Leads To An Increase In ADCC Through Higher Affinity For FC Gamma RIII,” Biotechnology & Bioengineering 74(4): 288-294). Methods of altering carbohydrate contents are known to those skilled in the art, see, e.g., Wallick,S.C. et al. (1988) “Glycosylation Of A VH Residue Of A Monoclonal Antibody Against Alpha (1 — -6) Dextran Increases Its Affinity For Antigen,” J. Exp. Med. 168(3): 1099-1109; Tao, M.H. et al. (1989) “Studies Of Aglycosylated Chimeric Mouse-Human IgG. Role Of Carbohydrate In The Structure And Effector Functions Mediated By The Human IgG Constant Region,” J. Immunol. 143(8): 2595-2601; Routledge, E.G. et al. (1995) “The Effect Of Aglycosylation On The Immunogenicity Of A Humanized Therapeutic CD3 Monoclonal Antibody,” Transplantation 60(8):847-53; Elliott, S. et al. (2003) “Enhancement Of Therapeutic Protein In Vivo Activities Through Glycoengineering,” Nature Biotechnol. 21:414-21; Shields, R.L. et al. (2002) “Lack Of Fucose On Human IgG N-Linked Oligosaccharide Improves Binding To Human Fcgamma RIII And Antibody-Dependent Cellular Toxicity.,” J. Biol. Chem. 277(30): 26733-26740).

[0101] In some embodiments, a humanized antibody is a derivative. Such a humanized antibody includes amino acid residue substitutions, deletions or additions in one or more non-human CDRs. The humanized antibody derivative can have substantially the same binding, better binding, or worse binding when compared to a non-derivative humanized antibody. In specific embodiments, one, two, three, four, or five amino acid residues of the CDRs have been substituted, deleted or added (i.e., mutated).

[0102] A derivative antibody or antibody fragment can be modified by chemical modifications using techniques known to those of skill in the art, including, but not limited to, specific chemical cleavage, acetylation, formulation, metabolic synthesis of tunicamycin, etc. In one embodiment, an antibodyderivative will possess a similar or identical function as the parental antibody. In another embodiment, an antibody derivative will exhibit an altered activity relative to the parental antibody. For example, a derivative antibody (or fragment thereof) can bind to its epitope more tightly or be more resistant to proteolysis than the parental antibody.

[0103] In some forms, derivative antibodies are engineered to increase neutralization potency, binding affinity, and breadth of recognition across spike glycoproteins from SARS-CoV-2 and related coronaviruses. In addition to SARS-CoV-2 ancestral strains (e.g., D614G), such viruses include, but are not limited to, emerging SARS-CoV-2 variants of concern and interest, such as Alpha (B. 1.1.7), Beta (B. 1.351), Gamma (P.l), Delta (B.1.617.2), Omicron (including sublineages BA.l, BA.2, BA.4, BA.5, BQ.l, XBB, XBB.1.5, and EG.5), as well as other sarbecoviruses with zoonotic potential, including SARS-CoV (responsible for the 2002-2003 SARS outbreak), bat-derived coronaviruses (e.g., RaTG13, BANAL-52, WIV 1), and pangolin-derived coronaviruses.

[0104] Derivatized antibodies can be used to alter the half-lives (e.g., serum half-lives) of parental antibodies in a mammal, preferably a human. Preferably such alteration will result in a half-life of greater than 15 days, preferably greater than 20 days, greater than 25 days, greater than 30 days, greater than 35 days, greater than 40 days, greater than 45 days, greater than 2 months, greater than 3 months, greater than 4 months, or greater than 5 months. The increased half-lives of the humanized antibodies of the present disclosure or fragments thereof in a mammal, preferably a human, results in a higher serum titer of said antibodies or antibody fragments in the mammal, and thus, reduces the frequency of the administration of said antibodies or antibody fragments and / or reduces the concentration of said antibodies or antibody fragments to be administered. Antibodies or fragments thereof having increased in vivo half-lives can be generated by techniques known to those of skill in the art. For example, antibodies or fragments thereof with increased in vivo half-lives can be generated by modifying (e.g., substituting, deleting or adding) amino acid residues identified as involved in the interaction between the Fc domain and the FcRn receptor. The humanized antibodies can be engineered to increase biological half-lives (see, e.g. U.S. Patent No. 6,277,375). For example, humanized antibodies can be engineered in the Fc-hinge domain to have increased in vivo or serum half- lives.

[0105] Antibodies or fragments thereof with increased in vivo half-lives can be generated by attaching to said antibodies or antibody fragments polymer molecules such as high molecular weight polyethyleneglycol (PEG). PEG can be attached to said antibodies or antibody fragments with or without a multifunctional linker either through site-specific conjugation of the PEG to the N- or C- terminus of said antibodies or antibody fragments or via epsilon-amino groups present on lysine residues. Linear or branched polymer deriv arization that results in minimal loss of biological activitywill be used. The degree of conjugation will be closely monitored by SDS-PAGE and mass spectrometry to ensure proper conjugation of PEG molecules to the antibodies. Unreacted PEG can be separated from antibody-PEG conjugates by, e.g., size exclusion or ion-exchange chromatography.

[0106] The antibodies can also be modified by the methods and coupling agents described by Davis et al. (See U.S. Patent No. 4,179,337) in order to provide compositions that can be injected into the mammalian circulatory system with substantially no immunogenic response.

[0107] One embodiment encompasses modification of framework residues of the humanized SARS- CoV-2 RBD antibodies. Framework residues in the framework regions can be substituted with the corresponding residue from the CDR donor antibody to alter, preferably improve, antigen binding. These framework substitutions are identified by methods well known in the art, e.g., by modeling of the interactions of the CDR and framework residues to identify framework residues important for antigen binding and sequence comparison to identify unusual framework residues at particular positions. (See, e.g., U.S. Patent No. 5,585,089; and Riechmann, L. et al. (1988) “Reshaping Human Antibodies For Therapy,” Nature 332:323-327).

[0108] Yet another embodiment encompasses anti- SARS-CoV-2 RBD antibodies (and more preferably, humanized antibodies) and antigen-binding fragments thereof that are recombinantly fused or chemically conjugated (including both covalently and non-covalently conjugations) to a heterologous molecule (i.e., an unrelated molecule). The fusion does not necessarily need to be direct, but can occur through linker sequences.

[0109] In one embodiment, such heterologous molecules are polypeptides having at least 10, at least 20, at least 30, at least 40, at least 50, at least 60, at least 70, at least 80, at least 90 or at least 100 amino acids. Such heterologous molecules can alternatively be viral or immunomodulatory proteins, including SARS-CoV-2 antigens or fragments thereof (such as spike protein, receptor-binding domain (RBD), nucleocapsid, membrane protein, or envelope protein), or therapeutic and diagnostic agents, such as: neutralizing antibodies (e.g., anti-SARS2 RBD antibodies), engineered binding proteins, fusion inhibitors, viral protease inhibitors, or other antiviral agents (e.g., remdesivir, nirmatrelvir, or molnupiravir), cytokines (such as interferons a, , or k), immune checkpoint inhibitors, or immune stimulatory agents (e.g., interleukins such as IL-2, IL-6, IL-7, or GM-CSF). In some embodiments, such molecules may further include cytotoxic agents or immunomodulators for targeting SARS-CoV-2-infected cells, such as toxins (e.g., pseudomonas exotoxin, diphtheria toxin, or ricin A), or conjugated drug moieties (e.g., paclitaxel, doxorubicin, or vincristine), antimetabolites (e.g., 5 -fluorouracil, methotrexate), or other pharmacologic agents useful for modulating host immune response or controlling viral replication.

[0110] Techniques for conjugating such therapeutic moieties to antibodies are well known; see, e.g., Arnon et al., “Monoclonal Antibodies For Immunotargeting Of Drugs In Cancer Therapy”, in MONOCLONAL ANTIBODIES AND CANCER THERAPY, Reisfeld et al. (eds.), 1985, pp. 243- 56, Alan R. Liss, Inc.); Hellstrom et al., “Antibodies For Drug Delivery”, in CONTROLLED DRUG DELIVERY (2nd Ed.), Robinson et al. (eds.), 1987, pp. 623-53, Marcel Dekker, Inc. ); Thorpe, “Antibody Carriers Of Cytotoxic Agents In Cancer Therapy: A Review”, in MONOCLONAL ANTIBODIES ‘84: BIOLOGICAL AND CLINICAL APPLICATIONS, Pinchera et al. (eds.), 1985, pp. 475-506); “Analysis, Results, And Future Prospective Of The Therapeutic Use Of Radiolabeled Antibody In Cancer Therapy”, in MONOCLONAL ANTIBODIES FOR CANCER DETECTION AND THERAPY, Baldwin et al. (eds.), 1985, pp. 303-16, Academic Press; and Thorpe et al. (1982) “The Preparation And Cytotoxic Properties Of Antibody-Toxin Conjugates,” Immunol. Rev. 62:119- 158.

[0111] In one embodiment, the SARS-CoV-2 antibodies or fusion molecules include an Fc portion. The Fc portion of such molecules can be varied by isotype or subclass, can be a chimeric or hybrid, and / or can be modified, for example to improve effector functions, control of half-life, tissue accessibility, augment biophysical characteristics such as stability, and improve efficiency of production (and less costly). Many modifications useful in construction of disclosed fusion proteins and methods for making them are known in the art, see for example Mueller, J.P. et al. (1997) “Humanized Porcine VCAM-Specific Monoclonal Antibodies With Chimeric IgG2 / G4 Constant Regions Block Human Leukocyte Binding To Porcine Endothelial Cells,” Mol. Immun. 34(6):441 - 452, Swann, P.G. (2008) “Considerations For The Development Of Therapeutic Monoclonal Antibodies,” Curr. Opin. Immun. 20:493-499 (2008), and Presta, L.G. (2008) “Molecular Engineering And Design Of Therapeutic Antibodies,” Curr. Opin. Immun. 20:460-470. In some embodiments the Fc region is the native IgG 1 , IgG2, or IgG4 Fc region. In some embodiments the Fc region is a hybrid, for example a chimeric having IgG2 / IgG4 Fc constant regions. Modifications to the Fc region include, but are not limited to, IgG4 modified to prevent binding to Fc gamma receptors and complement, IgGl modified to improve binding to one or more Fc gamma receptors, IgGl modified to minimize effector function (amino acid changes), IgGl with altered / no glycan (typically by changing expression host), and IgGl with altered pH-dependent binding to FcRn, and IgG4 with serine at amino acid resident #228 in the hinge region changed to proline (S228P) to enhance stability. The Fc region can include the entire hinge region, or less than the entire hinge region.

[0112] Substitutions, additions or deletions in the derivatized antibodies can be in the Fc region of the antibody and can thereby serve to modify the binding affinity of the antibody to one or moreFcyR. Methods for modifying antibodies with modified binding to one or more FcyR are known in the art, see, e.g., PCT Publication Nos. WO 04 / 029207, WO 04 / 029092, WO 04 / 028564, WO 99 / 58572, WO 99 / 51642, WO 98 / 23289, WO 89 / 07142, WO 88 / 07089, and U.S. Patent Nos. 5,843,597 and 5,642,821.

[0113] Modifications that affect Fc-mediated effector function are well known in the art (see U.S. Patent No. 6,194,551, and WO 00 / 42072; Stavenhagen, J.B. et al. (2007) “Fc Optimization Of Therapeutic Antibodies Enhances Their Ability To Kill Tumor Cells In Vitro And Controls Tumor Expansion In Vivo Via Low- Affinity Activating Fcgamma Receptors,” Cancer Res. 57(18):8882- 8890; Shields, R.L. et al. (2001) “High Resolution Mapping of the Binding Site on Human IgGl for FcyR I, FcyRII, FcyRIII, and FcRn and Design of IgGl Variants with Improved Binding to the FcyR,” J. Biol. Chem. 276(9):6591-6604). Exemplary variants of human IgGl Fc domains with reduced binding to FcyRIIA or FcyRIIIA, but unchanged or enhanced binding to FcyRIIB, include S239A, H268A, S267G, E269A, E293A, E293D, Y296F, R301A, V3O3A, A327G, K322A, E333A, K334A, K338A, A339A, D376A.

[0114] Any of the molecules of the present disclosure can be fused to marker sequences, such as a peptide, to facilitate purification. In preferred embodiments, the marker amino acid sequence is a hexa-histidine peptide, the hemagglutinin “HA” tag, which corresponds to an epitope derived from the influenza hemagglutinin protein (Wilson, LA. et al. (1984) “The Structure Of An Antigenic Determinant In A Protein,” Cell, 37:767-778) and the “flag” tag (Knappik, A. et al. (1994) “An Improved Affinity Tag Based On The FLAG Peptide For The Detection And Purification Of Recombinant Antibody Fragments,” Biotechniques 17(4):754-761).

[0115] The present disclosure also encompasses antibodies or their antigen-binding fragments that are conjugated to a diagnostic or therapeutic agent or any other molecule for which serum half-life is desired to be increased. The antibodies can be used diagnostically (in vivo, in situ or in vitro) to, for example, monitor the development or progression of a disease, disorder or infection as part of a clinical testing procedure to, e.g., determine the efficacy of a given treatment regimen. Detection can be facilitated by coupling the antibody to a detectable substance. Examples of detectable substances include various enzymes, prosthetic groups, fluorescent materials, luminescent materials, bioluminescent materials, radioactive materials, positron emitting metals, and nonradioactive paramagnetic metal ions. The detectable substance can be coupled or conjugated either directly to the antibody or indirectly, through an intermediate (such as, for example, a linker known in the art) using techniques known in the art. See, for example, U.S. Patent No. 4,741,900 for metal ions which can be conjugated to antibodies for use as diagnostics according to the present disclosure. Such diagnosis and detection can be accomplished by coupling the antibody to detectable substancesincluding, but not limited to, various enzymes, enzymes including, but not limited to, horseradish peroxidase, alkaline phosphatase, beta-galactosidase, or acetylcholinesterase; prosthetic group complexes such as, but not limited to, streptavidin / biotin and avidin / biotin; fluorescent materials such as, but not limited to, umbelliferone, fluorescein, fluorescein isothiocyanate, rhodamine, dichlorotriazinylamine fluorescein, dansyl chloride or phycoerythrin; luminescent material such as, but not limited to, luminol; bioluminescent materials such as, but not limited to, luciferase, luciferin, and aequorin; radioactive material such as, but not limited to, bismuth (213Bi), carbon (14C), chromium (51 Cr), cobalt (57Co), fluorine (18F), gadolinium (153Gd, 159Gd), gallium (68Ga, 67Ga), germanium (68Ge), holmium (166Ho), indium (115In, 113In, 112In, 11 Un), iodine (1311, 1251, 1231, 1211), lanthanium (140La), lutetium (177Lu), manganese (54Mn), molybdenum (99Mo), palladium (103Pd), phosphorous (32P), praseodymium (142Pr), promethium (149Pm), rhenium (186Re, 188Re), rhodium (105Rh), ruthemium (97Ru), samarium (153Sm), scandium (47Sc), selenium (75Se), strontium (85Sr), sulfur (35S), technetium (99Tc), thallium (201Ti), tin (113Sn, 117Sn), tritium (3H), xenon (133Xe), ytterbium (169Yb, 175Yb), yttrium (90Y), zinc (65Zn); positron emitting metals using various positron emission tomographies, and nonradioactive paramagnetic metal ions.

[0116] The molecules of the present disclosure can be conjugated to a second antibody to form an antibody heteroconjugate as described by Segal in U.S. Patent No. 4,676,980. Such heteroconjugate antibodies may additionally bind to haptens (such as fluorescein, etc.), or to cellular markers, or to cytokines, or chemokines (e.g., CCL21), etc.

[0117] The molecules of the present disclosed can be attached to solid supports, which are particularly useful for immunoassays or purification of the target antigen or of other molecules that are capable of binding to target antigen that has been immobilized to the support via binding to an antibody or antigen-binding fragment of the present disclosure. Such solid supports include, but are not limited to, glass, cellulose, polyacrylamide, nylon, polystyrene, polyvinyl chloride or polypropylene.5. Methods of Making Antibodies and Antigen Binding Fragments

[0118] The disclosed antibodies can be produced by any method known in the art useful for the production of polypeptides, e.g., in vitro synthesis, recombinant DNA production, and the like. Preferably, the antibodies are produced by recombinant DNA technology. The antibodies can be produced using recombinant immunoglobulin expression technology. The recombinant production of immunoglobulin molecules, including humanized antibodies are described in U.S. Patent No. 4,816,397 (Boss et al.), U.S. Patent Nos. 6,331,415 and 4,816,567 (both to Cabilly et al.), U.K. patent GB 2,188,638 (Winter et al.), and U.K. patent GB 2,209,757. Techniques for the recombinantexpression of immunoglobulins, including humanized immunoglobulins, can also be found, in Goeddel et al., Gene Expression Technology Methods in Enzymology Vol. 185 Academic Press (1991), and Borreback, Antibody Engineering, W. H. Freeman (1992). Additional information concerning the generation, design and expression of recombinant antibodies can be found in Mayforth, Designing Antibodies, Academic Press, San Diego (1993).

[0119] An exemplary process for the production of the recombinant chimeric antibodies can include the following: a) constructing, by conventional molecular biology methods, an expression vector that encodes and expresses an antibody heavy chain in which the CDRs and variable region of a murine anti-SARS-CoV-2 RBD monoclonal antibody are fused to an Fc region derived from a human immunoglobulin, thereby producing a vector for the expression of a chimeric antibody heavy chain; b) constructing, by conventional molecular biology methods, an expression vector that encodes and expresses an antibody light chain of the murine anti-SARS-CoV-2 RBD monoclonal antibody, thereby producing a vector for the expression of chimeric antibody light chain; c) transferring the expression vectors to a host cell by conventional molecular biology methods to produce a transfected host cell for the expression of chimeric antibodies; and d) culturing the transfected cell by conventional cell culture techniques so as to produce chimeric antibodies.

[0120] An exemplary process for the production of the recombinant humanized antibodies can include the following: a) constructing, by conventional molecular biology methods, an expression vector that encodes and expresses an anti-SARS-CoV-2 RBD heavy chain in which the CDRs and a minimal portion of the variable region framework that are required to retain donor antibody binding specificity are derived from a non-human immunoglobulin, such as a murine anti-SARS-CoV-2 RBD monoclonal antibody, and the remainder of the antibody is derived from a human immunoglobulin, thereby producing a vector for the expression of a humanized antibody heavy chain; b) constructing, by conventional molecular biology methods, an expression vector that encodes and expresses an antibody light chain in which the CDRs and a minimal portion of the variable region framework that are required to retain donor antibody binding specificity are derived from a non-human immunoglobulin, such as a murine anti-SARS-CoV-2 RBD monoclonal antibody, and the remainder of the antibody is derived from a human immunoglobulin, thereby producing a vector for the expression of humanized antibody light chain; c) transferring the expression vectors to a host cell by conventional molecular biology methods to produce a transfected host cell for the expression of humanized antibodies; and d) culturing the transfected cell by conventional cell culture techniques so as to produce humanized antibodies.

[0121] With respect to either exemplary method, host cells can be co-transfected with such expression vectors, which can contain different selectable markers but, with the exception of theheavy and light chain coding sequences, are preferably identical. This procedure provides for equal expression of heavy and light chain polypeptides. Alternatively, a single vector can be used which encodes both heavy and light chain polypeptides. The coding sequences for the heavy and light chains can include cDNA or genomic DNA or both. The host cell used to express the recombinant antibody can be either a bacterial cell such as Escherichia coli, or more preferably a eukaryotic cell (e.g., a Chinese hamster ovary (CHO) cell or a HEK-293 cell). The choice of expression vector is dependent upon the choice of host cell, and can be selected so as to have the desired expression and regulatory characteristics in the selected host cell. Other cell lines that can be used include, but are not limited to, CH0-K1, NSO, and PER.C6 (Crucell, Leiden, Netherlands).

[0122] Any of the above-described antibodies can be used to generate anti-idiotype antibodies using techniques well known to those skilled in the art (see, e.g., Greenspan, N.S. et al. (1989) “Idiotypes: Structure And Immunogenicity," FASEB J. 7:437-444; and Nisinoff, A. (1991) “Idiotypes: Concepts And Applications ," J • Immunol. 147(8):2429-2438).

[0123] The binding properties of any of the above antibodies can, if desired, be further improved by screening for variants that exhibit such desired characteristics. For example, such antibodies can be generated using various phage display methods known in the art. In phage display methods, functional antibody domains are displayed on the surface of phage particles which carry the polynucleotide sequences encoding them. In a particular embodiment, such phage can be utilized to display antigen binding domains, such as Fab and Fv or disulfide-bond stabilized Fv, expressed from a repertoire or combinatorial antibody library (e.g., human or murine). Phage expressing an antigen binding domain that binds the antigen of interest can be selected or identified with antigen, e.g., using labeled antigen or antigen bound or captured to a solid surface or bead. Phage used in these methods are typically filamentous phage, including fd and M13. The antigen binding domains are expressed as a recombinantly fused protein to either the phage gene III or gene VIII protein. Examples of phage display methods that can be used to make the immunoglobulins, or fragments thereof, of the present disclosure include those disclosed in Brinkman, U. et al. (1995) “Phage Display Of Disulfide - Stabilized Fv Fragments ," J. Immunol. Methods, 182:41-50, 1995; Ames, R.S. et al. (1995) “Conversion Of Murine Fabs Isolated From A Combinatorial Phage Display Library To Full Length Immunoglobulins," J. Immunol. Methods, 184: 177-186; Kettleborough, C.A. et al. (1994) “Isolation Of Tumor Cell-Specific Single-Chain Fv From Immunized Mice Using Phage -Antibody Libraries And The Re-Construction Of Whole Antibodies From These Antibody Fragments," Eur. J. Immunol., 24:952-958, 1994; Persic, L. et al. (1997) “An Integrated Vector System For The Eukaryotic Expression Of Antibodies Or Their Fragments After Selection From Phage Display Libraries," Gene, 187:9-18; Burton, D.R. et al. (1994) “Human Antibodies From Combinatorial Libraries," Adv.Immunol. 57:191-280; PCT Publications WO 92 / 001047; WO 90 / 02809; WO 91 / 10737; WO 92 / 01047; WO 92 / 18619; WO 93 / 11236; WO 95 / 15982; WO 95 / 20401; and U.S. Patents Nos. 5,698,426; 5,223,409; 5,403,484; 5,580,717; 5,427,908; 5,750,753; 5,821,047; 5,571,698; 5,427,908; 5,516,637; 5,780,225; 5,658,727; 5,733,743 and 5,969,108.

[0124] As described in the above references, after phage selection, the antibody coding regions from the phage can be isolated and used to generate whole antibodies, including humanized antibodies, or any other desired fragments, and expressed in any desired host, including mammalian cells, insect cells, plant cells, yeast, and bacteria, e.g., as described in detail below. For example, techniques to recombinantly produce Fab, Fab’ and F(ab’)2 fragments can also be employed using methods known in the art (such as those disclosed in PCT Publication WO 92 / 22324; Mullinax, R.L. et al. (1992) “Expression Of A Heterodimeric Fab Antibody Protein In One Cloning Step,” BioTechniques, 12(6):864-869; and Sawai et al. (1995) “Direct Production Of The Fab Fragment Derived From The Sperm Immobilizing Antibody Using Polymerase Chain Reaction And cDNA Expression Vectors,” Am. J. Reprod. Immunol. 34:26-34; and Better, M. et al. (1988) “Escherichia coli Secretion Of An Active Chimeric Antibody Fragment,” Science 240:1041-1043). Examples of techniques which can be used to produce single-chain Fvs and antibodies include those described in U.S. Patent Nos. 4,946,778 and 5,258,498; Huston, J.S. et al. (1991) “Protein Engineering Of Single-Chain Fv Analogs And Fusion Proteins,” Methods in Enzymology 203:46-88; Shu, L. et al. , “Secretion Of A Single-Gene-Encoded Immunoglobulin From Myeloma Cells,” Proc. Natl. Acad. Sci. (USA) 90:7995-7999; and Skerra. A. etal. (1988) “Assembly Of A Functional Immunoglobulin Fv Fragment In Escherichia coli,” Science 240:1038-1040.

[0125] Phage display technology can be used to increase the affinity of an antibody for anti-SARS- CoV-2 RBD. This technique would be useful in obtaining high affinity antibodies that could be used in the disclosed methods. This technology, referred to as affinity maturation, employs mutagenesis or CDR walking and re-selection using such receptors or ligands (or their extracellular domains) or an antigenic fragment thereof to identify antibodies that bind with higher affinity to the antigen when compared with the initial or parental antibody (See, e.g., Glaser, S.M. et al. (1992) “Antibody Engineering By Codon-Based Mutagenesis In A Filamentous Phage Vector System,” J. Immunol. 149:3903-3913). Mutagenizing entire codons rather than single nucleotides results in a semirandomized repertoire of amino acid mutations. Libraries can be constructed including of a pool of variant clones each of which differs by a single amino acid alteration in a single CDR and which contain variants representing each possible amino acid substitution for each CDR residue. Mutants with increased binding affinity for the antigen can be screened by contacting the immobilized mutants with labeled antigen. Any screening method known in the art can be used to identify mutantantibodies with increased avidity to the antigen e.g., ELISA) (see, e.g., Wu, H. et al. (1998) “Stepwise In Vitro Affinity Maturation OfVitaxin, AnAlphav Beta3-Specific Humanized Mab ,” Proc. Natl. Acad. Sci. (USA) 95(l l):6037-6042; Yelton, D.E. et al. (1995) “Affinity Maturation Of The BR96 Anti-Carcinoma Antibody By Codon-Based Mutagenesis ,” J. Immunol. 155:1994-2004). CDR walking which randomizes the light chain can be used (see, Schier et al. (1996) “Isolation Of Picomolar Affinity Anti-C-Erbb-2 Single-Chain Fv By Molecular Evolution Of The Complementarity Determining Regions In The Center Of The Antibody Binding Site,” J. Mol. Biol. 263:551-567).

[0126] The disclosure thus contemplates the use of random mutagenesis to identify improved CDRs. Phage display technology can alternatively be used to increase (or decrease) CDR affinity. This technology, referred to as affinity maturation, employs mutagenesis or “CDR walking” and reselection uses the target antigen or an antigenic fragment thereof to identify antibodies having CDRs that bind with higher (or lower) affinity to the antigen when compared with the initial or parental antibody (see, e.g., Glaser, S.M. et al. (1992) “Antibody Engineering By Codon-Based Mutagenesis In A Filamentous Phage Vector System,” J. Immunol. 149:3903-3913). Mutagenizing entire codons rather than single nucleotides results in a semi-randomized repertoire of amino acid mutations. Libraries can be constructed including of a pool of variant clones each of which differs by a single amino acid alteration in a single CDR and which contain variants representing each possible amino acid substitution for each CDR residue. Mutants with increased (or decreased) binding affinity for the antigen can be screened by contacting the immobilized mutants with labeled antigen. Any screening method known in the art can be used to identify mutant antibodies with increased (or decreased) avidity to the antigen (e.g., ELISA) (see, Wu, H. et al. (1998) “Stepwise In Vitro Affinity Maturation Of Vitaxin, An Alphav Beta3 -Specific Humanized Mab,” Proc. Natl. Acad. Sci. (USA) 95(ll):6037-6042; Yelton, D.E. et al. (1995) “Affinity Maturation Of The BR96 Anti-Carcinoma Antibody By Codon-Based Mutagenesis,” J. Immunol. 155:1994-2004). CDR walking which randomizes the light chain can be used (see, Schier et al. (1996) “Isolation Of Picomolar Affinity Anti-C-Erbb-2 Single-Chain Fv By Molecular Evolution Of The Complementarity Determining Regions In The Center Of The Antibody Binding Site,” J. Mol. Biol. 263:551-567).

[0127] Methods for accomplishing such affinity maturation are described for example in: Krause, J.C. et al. (2011) “An Insertion Mutation That Distorts Antibody Binding Site Architecture Enhances Function Of A Human Antibody ,” MBio. 2(1) pii: e00345-10. doi: 10.1 128 / mBio.00345-10; Kuan, C.T. et al. (2010) “Affinity -Matured Anti-Glycoprotein NMB Recombinant Immunotoxins Targeting Malignant Gliomas And Melanomas,” Int. J. Cancer 10.1002 / ijc.25645; Hackel, B.J. et al. (2010) “Stability And CDR Composition Biases Enrich Binder Functionality Landscapes,” J. Mol. Biol. 401(l):84-96; Montgomery, D.L. et al. (2009) “Affinity Maturation And Characterization Of AHuman Monoclonal Antibody Against HIV- 1 gp41 ,” MAbs l(5):462-474; Gustchina, E. et al. (2009) “Affinity Maturation By Targeted Diversification Of The CDR-H2 Loop Of A Monoclonal Fab Derived From A Synthetic Naive Human Antibody Library And Directed Against The Internal Trimeric Coiled-Coil Of Gp41 Yields A Set Of Fabs With Improved HIV-1 Neutralization Potency And Breadth,” Virology 393(1):112-119; Finlay, W.J. et al. (2009) “Affinity Maturation Of A Humanized Rat Antibody For Anti-RAGE Therapy: Comprehensive Mutagenesis Reveals A High Level Of Mutational Plasticity Both Inside And Outside The Complementarity-Determining Regions,” J. Mol. Biol. 388(3):541 -558; Bostrom, J. et al. (2009) “Improving Antibody Binding Affinity And Specificity For Therapeutic Development,” Methods Mol. Biol. 525:353-376; Steidl, S. et al. (2008) “In Vitro Affinity Maturation Of Human GM-CSF Antibodies By Targeted CDR-Diversification,” Mol. Immunol. 46(1):135-144; and Barderas, R. et al. (2008) “Affinity maturation of antibodies assisted by in silico modeling,” Proc. Natl. Acad. Sci. (USA) 105(26):9029-9034.

[0128] In some forms, the SARS-CoV-2 polypeptides include a fully human monoclonal antibody variant, binding fragment, and uses thereof, wherein the antigen-biding domain binds specifically to a domain of SARS CoV-2 Spike protein. Fully human antibodies essentially relate to antibody molecules in which the entire sequence of both the light chain and the heavy chain, including the CDRs, arise from human genes. Such antibodies are termed “human antibodies” or “fully human antibodies” herein. Human monoclonal antibodies can be prepared by, e.g., the trioma technique; the human B-cell hybridoma technique (see Kozbor, et al., Hybridoma, 2:7 (1983)) and the EBV hybridoma technique to produce human monoclonal antibodies (see Cole, et al., PNAS 82:859 (1985)), or as taught herein. Human monoclonal antibodies may be utilized in the practice of the presently disclosed and claimed invention and may be produced by using human hybridomas (see Cote, et al., PNAS 80:2026 (1983)) or by transforming human B-cells with Epstein Barr Virus in vitro (see Cole, et al., 1985), relevant portions incorporated herein by reference.

[0129] In addition, human antibodies can be made by introducing human immunoglobulin loci into transgenic animals, e.g., mice in which the endogenous immunoglobulin genes have been partially or completely inactivated. Upon challenge, human antibody production is observed, which closely resembles that seen in humans in all respects, including gene rearrangement, assembly, and antibody repertoire. This approach is described, for example but not by way of limitation, in U.S. Pat. Nos. 5,545,807; 5,545,806; 5,569,825; 5,625,126; 5,633,425; 5,661,016, and in Marks et al., J Biol. Chem. 267: 16007, (1992); Lonberg et al., Nature, 368:856 (1994); Morrison, 1994; Fishwild et al., Nature Biotechnol. 14:845 (1996); Neuberger, Nat. Biotechnol. 14:826 (1996); and Lonberg and Huszar, Int Rev Immunol. 13:65 (1995), relevant portions incorporated herein by reference.

[0130] A method for producing an antibody of interest, such as a human antibody, is disclosed in U.S. Pat. No. 5,916,771, issued to Hori et al. on Jun. 29, 1999, and incorporated herein by reference. It includes introducing an expression vector that contains a nucleotide sequence encoding a heavy chain into one mammalian host cell in culture, introducing an expression vector containing a nucleotide sequence encoding a light chain into another mammalian host cell, and fusing the two cells to form a hybrid cell. The hybrid cell expresses an antibody containing the heavy chain and the light chain.

[0131] An antibody or antigen-binding fragment can be generated with an engineered sequence or glycosylation state to confer preferred levels of activity in antibody-dependent cellular cytotoxicity (ADCC), antibody -dependent cellular phagocytosis (ADCP), antibody-dependent neutrophil phagocytosis (ADNP), or antibody-dependent complement deposition (ADCD) functions as measured by bead-based or cell-based assays or in vivo studies in animal models.

[0132] Alternatively, or additionally, it may be useful to combine amino acid modifications with one or more further amino acid modifications that alter complement component Clq binding and / or the complement-dependent cytotoxicity (CDC) function of the Fc region of an IL-23pl9 binding molecule. The binding polypeptide of particular interest may be one that binds to Clq and displays complement-dependent cytotoxicity. Polypeptides with pre-existing Clq binding activity, optionally further having the ability to mediate CDC may be modified such that one or both of these activities are enhanced. Amino acid modifications that alter Clq and / or modify its complement-dependent cytotoxicity function are described, for example, in W0 / 0042072, which is hereby incorporated by reference.

[0133] An Fc region of an antibody can be designed to alter the effector function, e.g., by modifying Clq binding and / or FcyR binding and thereby changing complement-dependent cytotoxicity (CDC) activity and / or antibody-dependent cell-mediated cytotoxicity (ADCC) activity. These "effector functions" are responsible for activating or diminishing a biological activity (e.g., in a subject). Examples of effector functions include, but are not limited to: Clq binding; CDC; Fc receptor binding; ADCC; phagocytosis; down-regulation of cell surface receptors (e.g., B cell receptor; BCR), etc. Such effector functions may require the Fc region to be combined with a binding domain (e.g., an antibody variable domain) and can be assessed using various assays (e.g., Fc binding assays, ADCC assays, CDC assays, etc.).

[0134] For example, one can generate a variant Fc region of an antibody with improved Clq binding and improved FcyRIII binding (e.g., having both improved ADCC activity and improved CDC activity). Alternatively, if it is desired that effector function be reduced or ablated, a variant Fc region can be engineered with reduced CDC activity and / or reduced ADCC activity. In other embodiments,only one of these activities may be increased, and, optionally, also the other activity reduced (e.g., to generate an Fc region variant with improved ADCC activity, but reduced CDC activity and vice versa).

[0135] A single chain variable fragment (scFv) is a fusion of the variable regions of the heavy and light chains of immunoglobulins, linked together with a short (usually serine, glycine) linker. This chimeric molecule retains the specificity of the original immunoglobulin, despite removal of the constant regions and the introduction of a linker peptide. This modification usually leaves the specificity unaltered. These molecules were created historically to facilitate phage display where it is highly convenient to express the antigen-binding domain as a single peptide. Alternatively, scFv can be created directly from subcloned heavy and light chains derived from a hybridoma or B cell. Single chain variable fragments lack the constant Fc region found in complete antibody molecules, and thus, the common binding sites (e.g., protein A / G) used to purify antibodies. These fragments can often be purified / immobilized using Protein L since Protein L interacts with the variable region of kappa light chains.

[0136] Flexible linkers generally are comprised of helix- and turn-promoting amino acid residues such as alanine, serine, and glycine. However, other residues can function as well. A random linker library was constructed in which the genes for the heavy and light chain variable domains were linked by a segment encoding an 18-amino acid polypeptide of variable composition. The scFv repertoire (approx. 5 x 106different members) is displayed on filamentous phage and subjected to affinity selection with hapten. The population of selected variants exhibited significant increases in binding activity but retained considerable sequence diversity. In certain embodiments, the antigen-binding fragments are further modified to increase their serum half-life by using modified Fc regions or mutations to the various constant regions, as are known in the art.B. Anti-SARS-CoV-2 RBD Chimeric Antigen Receptors (CAR)

[0137] Chimeric Antigen Receptor (CAR) proteins including the disclosed SARS-CoV-2 RBD- hinding proteins as an antigen binding domain, and cells expressing the same are also provided. Typically, CARs also include a transmembrane domain and one or more intracellular / cytoplasmic domains.

[0138] CARs are engineered receptors that possess both antigen-binding and T cell-activating functions. Immunotherapy using T cells genetically engineered to express a CAR is rapidly emerging as a promising new treatment for hematological and non- hematological malignancies. Based on the location of the CAR in the membrane of the cell, the CAR can be divided into three main distinct domains, including an extracellular antigen-binding domain, followed by a space region, a transmembrane domain, and the intracellular signaling domain. The antigen-binding domain, mostcommonly derived from variable regions of immunoglobulins, typically contains VH and VL chains that are joined up by a linker to form the so called “scFv.” The segment interposing between the antigen-binding domain (e.g., scFv) and the transmembrane domain is a “spacer domain.” The spacer domain can include the constant IgGl hinge CH2-CH3 Fc domain. In some cases, the spacer domain and the transmembrane domain are derived from CD8. The intracellular signaling domains mediating T cell activation can include a CD3^ co receptor signaling domain derived from C region of the TCR a and 0 chains and one or more costimulatory domains.

[0139] In the disclosed CARs, the antigen binding domain is typically a disclosed anti- SARS-CoV- 2 binding protein. In some forms, the antigen-binding domain is derived from an antibody, e.g., a disclosed SARS-CoV-2 antibody. As introduced above, the term antibody herein refers to natural or synthetic polypeptides that bind a target antigen, and such antibodies can form part or all of the antigen binding domain of the CAR. The term includes polyclonal and monoclonal antibodies, including intact antibodies and functional (e.g., antigen-binding) antibody fragments, including Fab fragments, F(ab')2 fragments, Fab' fragments, Fv fragments, recombinant IgG (rlgG) 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, and tetrabodies, tandem di scFv, tandem tri scFv. The term also encompasses intact or full-length antibodies, including antibodies of any class or subclass, including IgG and sub classes thereof, IgM, IgE, IgA, and IgD. The antigenbinding domain of a CAR can contain complementary determining regions (CDR) of an antibody, variable regions of an antibody, and / or antigen binding fragments thereof. For example, the antigenbinding domain for an SARS-CoV-2 RBD CAR can be derived from a disclosed SARS-CoV-2 antibody as described above. In some forms, the antigen-binding domain can include an F(ab')2, Fab’, Fab, Fv or scFv.

[0140] In some forms, the CAR includes one or more spacer domain(s) (also referred to as hinge domain) that is located between the extracellular antigen-binding domain and the transmembrane domain. A spacer domain is an amino acid segment that is generally found between two domains of a protein and may allow for flexibility of the protein and movement of one or both of the domains relative to one another. Any amino acid sequence that provides such flexibility and movement of the extracellular antigen-binding domain relative to the transmembrane domain can be used. The spacer domain can be a spacer or hinge domain of a naturally occurring protein. In some forms, the hinge domain is derived from CD8a, such as, a portion of the hinge domain of CD8a, e.g., a fragmentcontaining at least 5 (e.g., 5, 10, 15, 20, 25, 30, 35, or 40) consecutive amino acids of the hinge domain of CD8a. Hinge domains of antibodies, such as an IgG, IgA, IgM, IgE, or IgD antibodies can also be used. In some forms, the hinge domain is the hinge domain that joins the constant CHI and CH2 domains of an antibody. Non naturally occurring peptides may also be used as spacer domains. For example, the spacer domain can be a peptide linker, such as a (GxS)n linker, wherein x and n, independently can be an integer of 3 or more, including 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, or more.

[0141] In some forms, the CAR includes a transmembrane domain that can be directly or indirectly fused to the antigen-binding domain. The transmembrane domain may be derived either from a natural or a synthetic source. In some forms, the transmembrane domain of the CAR includes a transmembrane domain of an alpha, beta or zeta chain of a T cell receptor, CD8, CD4, CD28, CD137, CD80, CD86, CD 152 (CTLA-4) or PD1, or a portion thereof. Transmembrane domains can also contain at least a portion of a synthetic, non naturally occurring protein segment. In some forms, the transmembrane domain is a synthetic, non naturally occurring alpha helix or beta sheet. In some forms, the protein segment is at least about 15 amino acids, e.g., at least 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, or more amino acids. Examples of synthetic transmembrane domains are known in the art, for example in U.S. Patent No. 7,052,906 and PCT Publication No. WO 2000 / 032776.

[0142] The intracellular signaling domain is responsible for activation of at least one of the normal effector functions of the immune effector cell expressing the CAR. The term effector function refers to a specialized function of a cell. Effector function of a T cell, for example, may be cytolytic activity or helper activity including the secretion of cytokines. In some forms, an intracellular signaling domain includes the zeta chain of the T cell receptor or any of its homologs (e.g., eta, delta, gamma or epsilon), MB1 chain, B29, Fc RIII, Fc RI and combinations of signaling molecules such as CD3 and CD28, 4 IBB, 0X40 and combination thereof, as well as other similar molecules and fragments. Intracellular signaling portions of other members of the families of activating proteins can be used, such as FcyRIII and FcsRI.

[0143] Many immune effector cells require co stimulation, in addition to stimulation of an antigenspecific signal, to promote cell proliferation, differentiation and survival, as well as to activate effector functions of the cell. Therefore, in some forms, the CAR includes at least one co stimulatory signaling domain. The term co stimulatory signaling domain, refers to at least a portion of a protein that mediates signal transduction within a cell to induce an immune response such as an effector function. The co stimulatory signaling domain can be a cytoplasmic signaling domain from a co stimulatory protein, which transduces a signal and modulates responses mediated by immune cells, such as T cells, NK cells, macrophages, neutrophils, or eosinophils. In some forms, the co stimulatorysignaling domain is derived from a co stimulatory molecule selected from CD27, CD28, CD137, 0X40, CD30, CD40, CD3, LFA 1, ICOS, CD2, CD7, LIGHT, NKG2C, B7 H3, ligands of CD83 and combinations thereof.

[0144] CARs can be used in order to generate immuno-responsive cells, such as T cells, specific for selected targets, such as malignant cells, with a wide variety of receptor chimera constructs having been described (see U.S. Patent Nos. 5,843,728; 5,851,828; 5,912,170; 6,004,811 ; 6,284,240; 6,392,013; 6,410,014; 6,753,162; 8,211,422; and PCT Publication WO 9215322, each of which is specifically incorporated by reference herein in its entirety). Alternative CAR constructs can be characterized as belonging to successive generations. First-generation CARs typically include a single-chain variable fragment of an antibody specific for an antigen, for example including a VL linked to a VH of a specific antibody, linked by a flexible linker, for example by a CD8a hinge domain and a CD8a transmembrane domain, to the transmembrane and intracellular signaling domains of either CD3^ or FcRy (scFv CD3^ or scFv FcRy; see U.S. Patent No. 7,741,465; U.S. Patent No. 5,912,172; U.S. Patent No. 5,906,936, each of which is specifically incorporated by reference herein in its entirety). Second-generation CARs incorporate the intracellular domains of one or more costimulatory molecules, such as CD28, 0X40 (CD134), or 4 IBB (CD137) within the endodomain (for example scFv CD28 / OX40 / 4 IBB CD3 ; see U.S. Patent Nos.8, 911,993; 8,916,381; 8,975,071 ; 9,101,584; 9,102,760; 9,102,761, each of which is specifically incorporated by reference herein in its entirety). Third- generation CARs include a combination of costimulatory endodomains, such a CD3^ chain, CD97, GDI la CD18, CD2, ICOS, CD27, CD154, CDS, 0X40, 4 IBB, or CD28 signaling domains (for example scFv CD28 4 IBB CD3^ or scFv CD28 OX40-CD3 ; see U.S. Patent No.8,906,682; U.S. Patent No.8,399,645; U.S. Pat. No. 5,686,281 ; PCT Publication No. WO2014134165; PCT Publication No. W02012079000, each of which is specifically incorporated by reference herein in its entirety). Alternatively, co stimulation can be orchestrated by expressing CARs in antigen- specific T cells, chosen so as to be activated and expanded following engagement of their native aPTCR, for example by antigen on professional antigen-presenting cells, with attendant co stimulation. Any of the first, second, or third generation CARs described above can be used in accordance with the disclosed compositions and methods.

[0145] In some forms, the CAR cells target and / or is used for reducing SARS-CoV-2 symptoms, complete elimination or neutralization of the virus, or other infections / disease related to SARS-CoV- 2 proteins (e.g., e.g., spike protein, receptor-binding domain (RBD), nucleocapsid protein, membrane protein, or envelope protein). Such CAR cells may be engineered to express chimeric antigen receptors specific for one or more SARS-CoV-2 antigens, enabling targeted recognition and clearance of infected cells or viral particles. In certain embodiments, the CAR cells may further be employedto modulate inflammatory or immune responses associated with COVID- 19, including cytokine storm or post-acute sequelae of SARS-CoV-2 infection (e.g., long COVID).C. Nucleic Acids

[0146] Isolated nucleic acids and vectors encoding or expressing SARS-CoV-2 RBD binding molecules are also provided. As used herein, “isolated nucleic acid” refers to a nucleic acid that is separated from other nucleic acid molecules that are present in a mammalian genome, including nucleic acids that normally flank one or both sides of the nucleic acid in a mammalian genome.

[0147] An isolated nucleic acid can be, for example, a DNA molecule, provided one of the nucleic acid sequences normally found immediately flanking that DNA molecule in a naturally-occurring genome is removed or absent. Thus, an isolated nucleic acid includes, without limitation, a DNA molecule that exists as a separate molecule independent of other sequences (e.g., a chemically synthesized nucleic acid, or a cDNA or genomic DNA fragment produced by PCR or restriction endonuclease treatment), as well as recombinant DNA that is incorporated into a vector, an autonomously replicating plasmid, a virus (e.g., a retrovirus, lentivirus, adenovirus, or herpes virus), or into the genomic DNA of a prokaryote or eukaryote. In addition, an isolated nucleic acid can include an engineered nucleic acid such as a recombinant DNA molecule that is part of a hybrid or fusion nucleic acid. A nucleic acid existing among hundreds to millions of other nucleic acids within, for example, a cDNA library or a genomic library, or a gel slice containing a genomic DNA restriction digest, is not to be considered an isolated nucleic acid.

[0148] Nucleic acid encoding the disclosed polypeptide sequences are expressly provided. Nucleic acids can be single strand or double stranded, and can be in sense or antisense orientation, or can be complementary to a reference sequence.

[0149] Nucleic acids can be DNA, RNA, or nucleic acid analogs. Nucleic acid analogs can be modified at the base moiety, sugar moiety, or phosphate backbone. Such modification can improve, for example, stability, hybridization, or solubility of the nucleic acid. Modifications at the base moiety can include deoxyuridine for deoxythymidine, and 5-methyl-2’-deoxycytidine or 5-bromo- 2 ’-deoxy cytidine for deoxy cytidine. Modifications of the sugar moiety can include modification of the 2’ hydroxyl of the ribose sugar to form 2’-O-methyl or 2’-O-allyl sugars. The deoxyribose phosphate backbone can be modified to produce morpholino nucleic acids, in which each base moiety is linked to a six membered, morpholino ring, or peptide nucleic acids, in which the deoxyphosphate backbone is replaced by a pseudopeptide backbone and the four bases are retained. See, for example, Summerton and Weller (1997) Antisense Nucleic Acid Drug Dev. 7:187-195; and Hyrup et al. (1996) Bioorgan. Med. Chem. 4:5-23. In addition, the deoxyphosphate backbone can be replaced with, forexample, a phosphorothioate or phosphorodithioate backbone, a phosphoroamidite, or an alkyl phosphotriester backbone.

[0150] Nucleic acids, such as those described above, can be inserted into vectors for expression in cells. As used herein, a “vector” is a replicon, such as a plasmid, phage, or cosmid, into which another DNA segment may be inserted so as to bring about the replication of the inserted segment. Vectors can be expression vectors. An “expression vector” is a vector that includes one or more expression control sequences, and an “expression control sequence” is a DNA sequence that controls and regulates the transcription and / or translation of another DNA sequence.

[0151] Nucleic acids in vectors can be operably linked to one or more expression control sequences. As used herein, “operably linked” means incorporated into a genetic construct so that expression control sequences effectively control expression of a coding sequence of interest. Examples of expression control sequences include promoters, enhancers, and transcription terminating regions. A promoter is an expression control sequence composed of a region of a DNA molecule, typically within 100 nucleotides upstream of the point at which transcription starts (generally near the initiation site for RNA polymerase II). To bring a coding sequence under the control of a promoter, it is necessary to position the translation initiation site of the translational reading frame of the polypeptide between one and about fifty nucleotides downstream of the promoter. Enhancers provide expression specificity in terms of time, location, and level. Unlike promoters, enhancers can function when located at various distances from the transcription site. An enhancer also can be located downstream from the transcription initiation site. A coding sequence is “operably linked” and “under the control” of expression control sequences in a cell when RNA polymerase is able to transcribe the coding sequence into mRNA, which then can be translated into the protein encoded by the coding sequence.

[0152] Suitable expression vectors include, without limitation, plasmids and viral vectors derived from, for example, bacteriophage, baculoviruses, tobacco mosaic virus, herpes viruses, cytomegalovirus, retroviruses, vaccinia viruses, adenoviruses, and adeno-associated viruses. Numerous vectors and expression systems are commercially available from such corporations as Novagen (Madison, WI), Clontech (Palo Alto, CA), Stratagene (La Jolla, CA), and Invitrogen Life Technologies (Carlsbad, CA).

[0153] In expression vector can include a tag sequence. Tag sequences are typically expressed as a fusion with the encoded polypeptide. Such tags can be inserted anywhere within the polypeptide including at either the carboxyl or amino terminus. Examples of useful tags include, but are not limited to, green fluorescent protein (GFP), glutathione S-transferase (GST), polyhistidine, c-myc, hemagglutinin, Flag™ tag (Kodak, New Haven, CT), maltose E binding protein and protein A.

[0154] Vectors containing nucleic acids to be expressed can be transferred into host cells. The term “host cell” is intended to include prokaryotic and eukaryotic cells into which a recombinant expression vector can be introduced. As used herein, “transformed” and “transfected” encompass the introduction of a nucleic acid molecule (e.g., a vector) into a cell by one of a number of techniques. Although not limited to a particular technique, a number of these techniques are well established within the art. Prokaryotic cells can be transformed with nucleic acids by, for example, electroporation or calcium chloride mediated transformation. Nucleic acids can be transfected into mammalian cells by techniques including, for example, calcium phosphate co-precipitation, DEAE- dextran- mediated transfection, lipofection, electroporation, or microinjection. Host cells (e.g., a prokaryotic cell or a eukaryotic cell such as a CHO cell) can be used to, for example, produce the PD- 1 antagonist polypeptides described herein.

[0155] As used herein, the term “substantially purified” refers to isolation of the antibodies or antigen-binding portions thereof of the present invention against SARS-CoV-2 such that the antibodies or antigen-binding portions includes the majority percent of the sample in which it resides. Typically, in a sample, a substantially purified component includes 50%, preferably 80%-85%, more preferably 90, 91, 92, 93, 94, 95, 96, 97, 98, or 99% of the sample. Techniques for purifying polynucleotides and polypeptides of interest are well-known in the art and include, for example, ionexchange chromatography, affinity chromatography and sedimentation according to density.

[0156] As used herein, the term a “coding sequence” or a sequence which “encodes” the antibodies or antigen-binding portions thereof of the present invention against SARS-CoV-2, refers to a nucleic acid molecule that is transcribed (in the case of DNA) and translated (in the case of mRNA) into a polypeptide when placed under the control of appropriate regulatory sequences (or “control elements”) and in vitro or in vivo. The boundaries of the coding sequence are determined by a start codon at the 5’ (amino) terminus and a translation stop codon at the 3' (carboxy) terminus. A coding sequence can include, but is not limited to, cDNA from viral, prokaryotic or eukaryotic mRNA, genomic DNA sequences from viral or prokaryotic DNA, and even synthetic DNA sequences. A transcription termination sequence may be located 3' to the coding sequence.

[0157] As used herein, the term “control elements”, includes, but is not limited to, transcription promoters, transcription enhancer elements, transcription termination signals, poly adenylation sequences (located 3' to the translation stop codon), sequences for optimization of initiation of translation (located 5' to the coding sequence), and translation termination sequences, and / or sequence elements controlling an open chromatin structure.

[0158] As used herein, the term “nucleic acid” includes, but is not limited to, DNA or RNA that encodes the antibodies or antigen- binding portions thereof of the present invention against SARS-CoV-2 of the present invention, whether expressed or optimized for prokaryotic or eukaryotic expression. The term also captures sequences that include any of the known base analogs of DNA and RNA.

[0159] As used herein, the term “operably linked” refers to an arrangement of elements wherein the components so described are configured so as to perform their usual function. Thus, a given promoter operably linked to a coding sequence is capable of effecting the expression of the coding sequence when active. The promoter need not be contiguous with the coding sequence, so long as it functions to direct the expression thereof. Thus, for example, intervening untranslated yet transcribed sequences can be present between the promoter sequence and the coding sequence and the promoter sequence can still be considered “operably linked” to the coding sequence.

[0160] As used herein, the term “recombinant” refers to a polynucleotide that encodes the mutant SARS-CoV-2 spike antibody whether from the viral genome, cDNA, semisynthetic, or synthetic origin which, by virtue of its origin or manipulation: (1) is not associated with all or a portion of the polynucleotide with which it is associated in nature; and / or (2) is linked to a polynucleotide other than that to which it is linked in nature. The term “recombinant” as used with respect to a protein or polypeptide means a polypeptide produced by expression of a recombinant polynucleotide. “Recombinant host cells,” “host cells,” “cells,” “cell lines,” “cell cultures,” and other such terms denoting prokaryotic microorganisms or eukaryotic cell lines cultured as unicellular entities, are used interchangeably, and refer to cells which can be, or have been, used as recipients for recombinant vectors or other transfer DNA, and include the progeny of the original cell which has been transfected. It is understood that the progeny of a single parental cell may not necessarily be completely identical in morphology or in genomic or total DNA complement to the original parent, due to accidental or deliberate mutation. Progeny of the parental cell which are sufficiently similar to the parent to be characterized by the relevant property, such as the presence of a nucleotide sequence encoding a desired peptide, are included in the progeny intended by this definition, and are covered by the above terms.

[0161] Techniques for determining amino acid sequence “similarity” are well known in the art. In general, “similarity” means the exact amino acid to amino acid comparison of two or more polypeptides at the appropriate place, where amino acids are identical or possess similar chemical and / or physical properties such as charge or hydrophobicity. A so-termed “percent similarity” then can be determined between the compared polypeptide sequences. Techniques for determining nucleic acid and amino acid sequence identity also are well known in the art and include determining the nucleotide sequence of the mRNA for that gene (usually via a cDNA intermediate) and determining the amino acid sequence encoded thereby and comparing this to a second amino acid sequence. Ingeneral, “identity” refers to an exact nucleotide to nucleotide or amino acid to amino acid correspondence of two polynucleotides or polypeptide sequences, respectively.

[0162] Two or more polynucleotide sequences can be compared by determining their “percent identity.” Two or more amino acid sequences likewise can be compared by determining their “percent identity.” The percent identity of two sequences, whether nucleic acid or peptide sequences, is generally described as the number of exact matches between two aligned sequences divided by the length of the shorter sequence and multiplied by 100. An approximate alignment for nucleic acid sequences is provided by the local homology algorithm of Smith and Waterman, Advances in Applied Mathematics 2:482-489 (1981). This algorithm can be extended to use with peptide sequences using the scoring matrix developed by Dayhoff, Atlas of Protein Sequences and Structure, M. O. Dayhoff ed., 5 suppl. 3:353-358, National Biomedical Research Foundation, Washington, D.C., USA, and normalized by Gribskov, Nucl. Acids Res. 14(6):6745-6763 (1986), relevant portion incorporated herein by reference. Suitable programs for calculating the percent identity or similarity between sequences are generally known in the art.

[0163] As used herein, the term a “vector” refers to a nucleic acid capable of transferring gene sequences to target cells (e.g., bacterial plasmid vectors, viral vectors, non-viral vectors, particulate carriers, and liposomes). Typically, “vector construct,” “expression vector,” and “gene transfer vector,” mean any nucleic acid construct capable of directing the expression of one or more sequences of interest in a host cell. Thus, the term includes cloning and expression vehicles, as well as viral vectors. The term is used interchangeably with the terms “nucleic acid expression vector” and “expression cassette.”

[0164] Many suitable expression systems are commercially available, including, for example, the following: baculovirus expression (Reilly, P. R„ et al., BACULO VIRUS EXPRESSION VECTORS: A LABORATORY MANUAL (1992); Beames, et al., Biotechniques 11:378 (1991); Pharmingen; Clontech, Palo Alto, Calif.)), vaccinia expression systems (Earl, P. L., et al., “Expression of proteins in mammalian cells using vaccinia” In Current Protocols in Molecular Biology (F. M. Ausubel, et al. Eds.), Greene Publishing Associates & Wiley Interscience, New York (1991); Moss, B., et al., U.S. Pat. No. 5,135,855, issued Aug. 4, 1992), expression in bacteria (Ausubel, F. M., et al., CURRENT PROTOCOLS IN MOLECULAR BIOLOGY, John Wiley and Sons, Inc., Media Pa.; Clontech), expression in yeast (Rosenberg, S. and Tekamp-Olson, P., U.S. Pat. No. RE35,749, issued, Mar. 17, 1998, herein incorporated by reference; Shuster, J. R., U.S. Pat. No. 5,629,203, issued May 13, 1997, herein incorporated by reference; Gellissen, G., et al., Antonie Van Leeuwenhoek, 62(l-2):79-93 (1992); Romanos, M. A., et al., Yeast 8(6):423-488 (1992); Goeddel, D. V., Methods in Enzymology 185 (1990); Guthrie, C., and G. R. Fink, Methods in Enzymology 194 (1991)), expression inmammalian cells (Clontech; Gibco-BRL, Ground Island, N.Y.; e.g., Chinese hamster ovary (CHO) cell lines (Haynes, J., et al., Nuc. Acid. Res. 11 :687-706 (1983); 1983, Lau, Y. F., et al., Mol. Cell. Biol. 4:1469-1475 (1984); Kaufman, R. J., “Selection and coamplification of heterologous genes in mammalian cells,” in Methods in Enzymology, vol. 185, pp 537-566. Academic Press, Inc., San Diego Calif. (1991)), and expression in plant cells (plant cloning vectors, Clontech Laboratories, Inc., Palo-Alto, Calif., and Pharmacia LKB Biotechnology, Inc., Pistcataway, N.J.; Hood, E., et al., J. Bacteriol. 168: 1291-1301 (1986); Nagel, R„ et al., FEMS Microbiol. Lett. 67:325 (1990); An, et al., “Binary Vectors”, and others in Plant Molecular Biology Manual A3:l-19 (1988); Miki, B. L. A., et al., pp. 249-265, and others in Plant DNA Infectious Agents (Hohn, T., et al., eds.) Springer-Verlag, Wien, Austria, (1987); Plant Molecular Biology: Essential Techniques, P. G. Jones and J. M. Sutton, New York, J. Wiley, 1997; Miglani, Gurbachan Dictionary of Plant Genetics and Molecular Biology, New York, Food Products Press, 1998; Henry, R. J., Practical Applications of Plant Molecular Biology, New York, Chapman & Hall, 1997), relevant portions of any of the above are incorporated herein by reference.D. Host Cells

[0165] some embodiments, polypeptides, nucleic acids, or vectors provided herein are present within a host cells. The term “host cell” is intended to include prokaryotic and eukaryotic cells into which a nucleic acid such as a recombinant expression vector can be introduced. As used herein, “transformed” and “transfected” encompass the introduction of a nucleic acid molecule (e.g., an mRNA, or a vector, etc.) into a cell by one of a number of techniques. Although not limited to a particular technique, a number of these techniques are well established within the art. Prokaryotic cells can be transformed with nucleic acids by, for example, electroporation or calcium chloride mediated transformation. Nucleic acids can be transfected into mammalian cells by techniques including, for example, calcium phosphate co-precipitation, DEAE-dextran-mediated transfection, lipofection, electroporation, or microinjection. Host cells (e.g., a prokaryotic cell or a eukaryotic cell) can be used to produce polypeptides described herein.

[0166] In some forms, the cell is from an established cell line, or a primary cell. The term “primary cell,” refers to cells and cell cultures derived from a subject and allowed to grow in vitro for a limited number of passages, i.e. splitting, of the culture.

[0167] In some embodiments, particularly CAR embodiments, the cells are obtained from a human subject. Therefore, human cells expressing and / or including the disclosed polypeptides are provided. In preferred embodiments, the human cells include or express a SARS-CoV-2 RBD-CAR. For example, in some forms, the cells are autologous cells, i.e., cells obtained from a subject prior to introduction of the SARS-CoV-2 RBD-CAR, and / or nucleic acids, or vectors encoding the same, andre-introduction to the same subject following modification. In other forms, the cells are heterologous cells, i.e., cells obtained from a different subject than the intended recipient. In some forms, the cells are frozen prior to or after introduction of the SARS-CoV-2 RBD-CAR. Methods and compositions for freezing and thawing viable eukaryotic cells are known in the art. In some forms, the cells are autologous immune cells, such as T cells or progenitor cells / stem cells.

[0168] In some forms, cells are obtained from a healthy subject. In other forms, cells are obtained from a subject identified as having or at risk of having a disease or disorder or other disease or condition such as those mentioned elsewhere herein.

[0169] In some embodiments, the introduction of the polypeptides to the cells occurs through genetic modification of the cells. In some embodiments, genetic modification of the cell includes introduction of nucleic acids, or vectors encoding the polypeptides to the cell for expression of the polypeptides within the cell. Therefore, genetically modified (transgenic) cells including the disclosed proteins, e.g., SARS-CoV-2 RBD-CAR fusion proteins, are described.

[0170] In some forms, the cells are human immune cells, such as T cells, Natural Killers (NK) cells, macrophages, dendritic cells etc. Therefore, human immune cells that include or express the disclosed polypeptides including the SARS-CoV-2 RBD-CAR polypeptides are described. In some forms, prior to expansion and genetic modification, immune cells such as T cells are obtained from a diseased or healthy subject. The cells can be obtained from a number of samples, including peripheral blood mononuclear cells, bone marrow, lymph node tissue, cord blood, thymus tissue, tissue from a site of infection, ascites, pleural effusion, spleen tissue, and tumors. In some forms, the immune cells are obtained from a unit of blood collected from a subject using any number of techniques known to the skilled artisan, such as FICOLL™ separation. In one preferred form, cells from the circulating blood of an individual are obtained by apheresis. The apheresis product typically contains lymphocytes, including T cells, monocytes, granulocytes, B cells, other nucleated white blood cells, red blood cells, and platelets. The cells collected by apheresis can be washed to remove the plasma fraction and to place the cells in an appropriate buffer or media for subsequent processing steps. In some forms, the cells are washed with phosphate buffered saline (PBS). In some forms, the wash solution lacks calcium and can lack magnesium or can lack many if not all divalent cations. After washing, the cells can be resuspended in a variety of biocompatible buffers, such as, for example, Ca2+ free, Mg2+ free PBS, PLASM ALYTE A, or other saline solution with or without buffer. Alternatively, the undesirable components of the apheresis sample are removed and the cells directly resuspended in culture media.

[0171] In some forms, T cells are isolated from peripheral blood lymphocytes by lysing the red blood cells and depleting the monocytes, for example, by centrifugation through a PERCOLL™ gradientor by counterflow centrifugal elutriation. In specific forms, a specific subpopulation of T cells, such as CD3+, CD28+, CD4+, CD8+, CD45RA+, and CD45RO+ T cells, is further isolated by positive or negative selection techniques. For example, in some forms, T cells are isolated by incubation with anti CD3 / anti CD28 (i.e., 3x28) conjugated beads, such as DYNABEADS® M 450 CD3 / CD28 T, for a time period sufficient for positive selection of the desired T cells.E. Pharmaceutical Compositions

[0172] The compositions can be formulated with appropriate pharmaceutically acceptable carriers into pharmaceutical compositions for administration to an individual in need thereof. The formulations can be administered enterally (e.g., oral) or parenterally (e.g., by injection or infusion).

[0173] The compositions can be formulated for parenteral administration. “Parenteral administration’’, as used herein, means administration by any method other than through the digestive tract or non-invasive topical or regional routes. For example, parenteral administration may include administration to a patient intravenously, intradermally, intraarterially, intraperitoneally, intralesionally, intracranially, intraarticularly, intraprostatically, intrapleurally, intratracheally, intravitreally, intratumorally, intramuscularly, subcutaneously, subconjunctivally, intravesicularly, intrapericardially, intraumbilically, or transmucosal (nasal, vaginal, pulmonary, or rectal), e.g., by injection, and by infusion.

[0174] In some embodiments, the compositions are administered systemically by, for example, injection or infusion. In some embodiments, the compositions are administered locally by injection or infusion.

[0175] Parenteral formulations can be prepared as aqueous compositions using techniques known in the art. Typically, such compositions can be prepared as injectable formulations, for example, solutions or suspensions; solid forms suitable for using to prepare solutions or suspensions upon the addition of a reconstitution medium prior to injection; emulsions, such as water-in-oil (w / o) emulsions, oil-in-water (o / w) emulsions, and microemulsions thereof, liposomes, or emulsomes.

[0176] The carrier can be a solvent or dispersion medium containing, for example, water, ethanol, one or more polyols (e.g., glycerol, propylene glycol, and liquid polyethylene glycol), oils, such as vegetable oils (e.g., peanut oil, corn oil, sesame oil, etc.), and combinations thereof. The proper fluidity can be maintained, for example, by the use of a coating, such as lecithin, by the maintenance of the required nanocarrier size in the case of dispersion and / or by the use of surfactants. In many cases, isotonic agents, for example, sugars or sodium chloride, are included.

[0177] Solutions and dispersions of the active compounds as the free acid or base or pharmacologically acceptable salts thereof can be prepared in water or another solvent or dispersing medium suitably mixed with one or more pharmaceutically acceptable excipients including, but notlimited to, surfactants, dispersants, emulsifiers, pH modifying agents, viscosity modifying agents, and combination thereof.

[0178] Suitable surfactants may be anionic, cationic, amphoteric or nonionic surface active agents. Suitable anionic surfactants include, but are not limited to, those containing carboxylate, sulfonate and sulfate ions. Examples of anionic surfactants include sodium, potassium, ammonium of long chain alkyl sulfonates and alkyl aryl sulfonates such as sodium dodecylbenzene sulfonate; dialkyl sodium sulfosuccinates, such as sodium dodecylbenzene sulfonate; dialkyl sodium sulfosuccinates, such as sodium bis-(2-ethylthioxyl)-sulfosuccinate; and alkyl sulfates such as sodium lauryl sulfate. Cationic surfactants include, but are not limited to, quaternary ammonium compounds such as benzalkonium chloride, benzethonium chloride, cetrimonium bromide, stearyl dimethylbenzyl ammonium chloride, polyoxyethylene and coconut amine. Examples of nonionic surfactants include ethylene glycol monostearate, propylene glycol myristate, glyceryl monostearate, glyceryl stearate, polyglyceryl-4-oleate, sorbitan acylate, sucrose acylate, PEG- 150 laurate, PEG-400 monolaurate, polyoxyethylene monolaurate, polysorbates, polyoxyethylene octylphenylether, PEG- 1000 cetyl ether, polyoxyethylene tridecyl ether, polypropylene glycol butyl ether, Poloxamer® 401, stearoyl monoisopropanolamide, and polyoxyethylene hydrogenated tallow amide. Examples of amphoteric surfactants include sodium N-dodecyl-beta-alanine, sodium N-lauryl-P-iminodipropionate, myristoamphoacetate, lauryl betaine and lauryl sulfobetaine.

[0179] The formulation can contain a preservative to prevent the growth of microorganisms. Suitable preservatives include, but are not limited to, parabens, chlorobutanol, phenol, sorbic acid, and thimerosal. The formulation may also contain an antioxidant to prevent degradation of the active agent(s).

[0180] The formulation is typically buffered to a pH of 3-8 for parenteral administration upon reconstitution. Suitable buffers include, but are not limited to, phosphate buffers, acetate buffers, and citrate buffers.

[0181] Water soluble polymers are often used in formulations for parenteral administration. Suitable water-soluble polymers include, but are not limited to, polyvinylpyrrolidone, dextran, carboxymethylcellulose, and polyethylene glycol.

[0182] Sterile injectable solutions can be prepared by incorporating the active compounds in the required amount in the appropriate solvent or dispersion medium with one or more of the excipients listed above, as required, followed by filtered sterilization. Generally, dispersions are prepared by incorporating the various sterilized active ingredients into a sterile vehicle which contains the basic dispersion medium and the required other ingredients from those listed above. In the case of sterile powders for the preparation of sterile injectable solutions, exemplary methods of preparation includevacuum-drying and freeze-drying techniques which yield a powder of the active ingredient plus any additional desired ingredient from a previously sterile-filtered solution thereof.

[0183] Enteral formulations are prepared using pharmaceutically acceptable carriers. As generally used herein “carrier” includes, but is not limited to, diluents, preservatives, binders, lubricants, disintegrators, swelling agents, fillers, stabilizers, and combinations thereof. Polymers used in the dosage form include hydrophobic or hydrophilic polymers and pH dependent or independent polymers. Hydrophobic and hydrophilic polymers include, but are not limited to, hydroxypropyl methylcellulose, hydroxypropyl cellulose, hydroxyethyl cellulose, carboxy methylcellulose, polyethylene glycol, ethylcellulose, microcrystalline cellulose, polyvinyl pyrrolidone, polyvinyl alcohol, polyvinyl acetate, and ion exchange resins.

[0184] Carrier also includes all components of the coating composition, which may include plasticizers, pigments, colorants, stabilizing agents, and glidants. Formulations can be prepared using one or more pharmaceutically acceptable excipients, including diluents, preservatives, binders, lubricants, disintegrators, swelling agents, fillers, stabilizers, and combinations thereof.

[0185] Controlled release dosage formulations can be prepared as described in standard references such as “Pharmaceutical dosage form tablets”, eds. Liberman et. al. (New York, Marcel Dekker, Inc., 1989), “Remington - The science and practice of pharmacy”, 20th ed., Lippincott Williams & Wilkins, Baltimore, MD, 2000, and “Pharmaceutical dosage forms and drug delivery systems”, 6th Edition, Ansel et al., (Media, PA: Williams and Wilkins, 1995). These references provide information on excipients, materials, equipment and process for preparing tablets and capsules and delayed release dosage forms of tablets, capsules, and granules. These references provide information on carriers, materials, equipment and process for preparing tablets and capsules and delayed release dosage forms of tablets, capsules, and granules.

[0186] Stabilizers are used to inhibit or retard drug decomposition reactions which include, by way of example, oxidative reactions. Suitable stabilizers include, but are not limited to, antioxidants, butylated hydroxytoluene (BHT); ascorbic acid, its salts and esters; Vitamin E, tocopherol and its salts; sulfites such as sodium metabisulphite; cysteine and its derivatives; citric acid; propyl gallate, and butylated hydroxyanisole (BHA).

[0187] In some embodiments, the compositions are formulated for mucosal administration, such as through nasal, pulmonary, or buccal delivery.

[0188] Mucosal formulations may include one or more agents for enhancing delivery through the nasal mucosa. Agents for enhancing mucosal delivery are known in the art, see for example U.S. Patent Application No. 20090252672 to Eddington, and U.S. Patent Application No. 20090047234 to Touitou. Acceptable agents include, but are not limited to, chelators of calcium (EDTA), inhibitorsof nasal enzymes (boro-leucin, aprotinin), inhibitors of muco-ciliar clearance (preservatives), solubilizers of nasal membrane (cyclodextrin, fatty acids, surfactants) and formation of micelles (surfactants such as bile acids, Laureth 9 and taurodehydrofusidate (STDHF)). Compositions may include one or more absorption enhancers, including surfactants, fatty acids, and chitosan derivatives, which can enhance delivery by modulation of the tight junctions (TJ) (B. J. Aungst, et al., J. Pharm. Sci. 89(4):429-442 (2000)). In general, the optimal absorption enhancer should possess the following qualities: its effect should be reversible, it should provide a rapid permeation enhancing effect on the cellular membrane of the mucosa, and it should be non-cytotoxic at the effective concentration level and without deleterious and / or irreversible effects on the cellular membrane or cytoskeleton of the TJ.

[0189] Any of the disclosed compositions including, but not limited to the provided SARS-CoV-2 RBD binder polypeptides and CAR fusion proteins formed therefrom and / or nucleic acids encoding the same, can be delivered to target cells using a delivery vehicle. The delivery vehicles can be, for example, polymeric particles, inorganic particles, silica particles, liposomes, micelles, multilamellar vesicles, etc.

[0190] Delivery vehicles may be microparticles or nanoparticles. Nanoparticles are often utilized for intertissue application, penetration of cells, and certain routes of administration. The nanoparticles may have any desired size for the intended use. The nanoparticles may have any diameter from 10 nm up to about 1,000 nm. The nanoparticle can have a diameter from 10 nm to 900 nm, from 10 nm to 800 nm, from 10 nm to 700 nm, from 10 nm to 600 nm, from 10 nm to 500 nm, from 20 nm from 500 nm, from 30 nm to 500 nm, from 40 nm to 500 nm, from 50 nm to 500 nm, from 50 nm to 400 nm, from 50 nm to 350 nm, from 50 nm to 300 nm, or from 50 nm to 200 nm. In some embodiments the nanoparticles can have a diameter less than 400 nm, less than 300 nm, or less than 200 nm. The range can be between 50 nm and 300 nm.

[0191] Thus, in some embodiments, the delivery vehicles are nanoscale compositions, for example, 10 nm up to, but not including, about 1 micron. However, it will be appreciated that in some embodiments, and for some uses, the particles can be smaller, or larger (e.g., microparticles, etc.). Although many of the compositions disclosed herein are referred to as nanoparticle or nanocarrier compositions, it will be appreciated that in some embodiments and for some uses the carrier can be somewhat larger than nanoparticles. Such compositions can be referred to as microparticulate compositions. For example, a nanocarriers according to the present disclosure may be a microparticle. Microparticles can a diameter between, for example, 0.1 and 100 pm in size.

[0192] As provided are pharmaceutical packs and kits including one or more containers filled with antibody or fusion protein or nucleic acid. Additionally, one or more other prophylactic or therapeuticagents useful for the treatment of a disease can also be included in the pharmaceutical pack or kit. One embodiment provides a pharmaceutical pack or kit including one or more containers filled with one or more of the ingredients of the pharmaceutical compositions. Optionally associated with such container(s) can be a notice in the form prescribed by a governmental agency regulating the manufacture, use or sale of pharmaceuticals or biological products, which notice reflects approval by the agency of manufacture, use or sale for human administration.

[0193] Also provided are kits that can be used in the below methods. In one embodiment, a kit includes one or more antibodies or fusion proteins or nucleic acids. In another embodiment, a kit further includes one or more other prophylactic or therapeutic agents useful for the treatment or prevention of a disease or infection, in one or more containers.

[0194] For example, the antibodies of the present invention are formulated for administration to humans. For example, the antibodies of the present invention can be included in a pharmaceutical composition formulated for an administration that is: intranasal, intrapulmonary, intrabronchial, intravenous, oral, intraadiposal, intraarterial, intraarticular, intracranial, intradermal, intralesional, intramuscular, intrapericardial, intraperitoneal, intrapleural, intravesicular, local, mucosal, parenteral, enteral, subcutaneous, sublingual, topical, transbuccal, transdermal, via inhalation, via injection, in creams, in lipid compositions, via a catheter, via a lavage, via continuous infusion, via infusion, via local delivery, or via localized perfusion, and wherein the composition is a serum, drop, gel, ointment, spray, reservoir, or mist.III. Methods of UseA. Methods of Detecting SARS-CoV-2

[0195] The disclosed SARS-CoV-2 spike protein binders can be used to detect SARS-CoV-2 spike protein in various research, diagnostic, and prognostic applications. Typically, the binders are used to detect the spike protein or a fragment thereof (e.g., the receptor-binding domain) in a biological sample. Likewise, the disclosed binders can be used to isolate or purify SARS-CoV-2 spike protein, e.g., by immunoprecipitation, affinity purification, or related techniques.

[0196] Exemplary biological sources for detection of SARS-CoV-2 spike protein and fragments thereof are samples that include, but are not limited to, saliva, nasal swabs, bronchoalveolar lavage fluid, urine, serum, and blood, which may or may not include blood cells such as white blood cells, including immune cells or plasma cells. A sample may be obtained and processed using well-known and routine clinical methods. In some aspects, the biological sample includes a plurality of cells. The cells can be intact and / or permeabilized. In certain aspects, the biological sample includes fresh or frozen tissue. In specific aspects, the biological sample includes formalin-fixed, paraffin-embedded tissue.In some embodiments, the cells are permeabilized. Thus, in some embodiments, a cell lysate or homogenate is subjected to SARS-CoV-2 spike protein or fragment detection. The SARS-CoV-2 spike protein or fragment thereof is typically detected using one or more of the antibodies or other antigen binding molecules provided herein. The spike protein or fragment thereof can be detected by any suitable method utilizing the provided antibodies. In preferred embodiments, the spike protein or fragment thereof is detected and / or measured by an immunoassay. Immunoassays utilize biospecific capture reagents, such as antibodies, to capture or locate the SARS-CoV-2 spike protein or fragment.

[0197] It has been established that the SARS-CoV-2 polypeptides (e.g., recombinant antibodies) can be administered to a subject with a coronavirus infection to neutralize pathogenic coronaviruses in a subject. Therefore, methods of treating a subject with a coronavirus infection with a pharmaceutical composition containing the SARS-CoV-2 polypeptides (e.g., recombinant antibodies) are provided.

[0198] The steps of various useful immunodetection methods have been described in the scientific literature. In general, the immunobinding methods include obtaining a sample, and contacting the sample with an antibody specific for the protein to be detected, as the case may be, under conditions effective to allow the formation of immunocomplexes. In general, the detection of immunocomplex formation is well known in the art and may be achieved through the application of numerous approaches. These methods are generally based upon the detection of a label or marker, such as any of those radioactive, fluorescent, biological and enzymatic tags. Of course, one may find additional advantages through the use of a secondary binding ligand such as a second antibody and / or a biotin / avidin ligand binding arrangement, as is known in the art.

[0199] The antibody employed in the detection may itself be linked to a detectable label (also referred to as a detectable substance or reporter), wherein one would then simply detect this label, thereby allowing the amount of the primary immune complexes in the composition to be determined. Alternatively, the first antibody that becomes bound within the primary immune complexes may be detected by means of a second binding ligand that has binding affinity for the antibody. In these cases, the second binding ligand may be linked to a detectable label. The second binding ligand is itself often an antibody, which may thus be termed a “secondary” antibody. The primary immune complexes are contacted with the labeled, secondary binding ligand, or antibody, under effective conditions and for a period of time sufficient to allow the formation of secondary immune complexes. The secondary immune complexes are then generally washed to remove any non-specifically bound labeled secondary antibodies or ligands, and the remaining label in the secondary immune complexes is then detected.

[0200] Traditional immunoassays including, for example, sandwich immunoassays such as ELISA or fluorescence-based immunoassays, as well as other enzyme immunoassays, can be used fordetecting the SARS-CoV-2 spike protein or fragment thereof. In other embodiments, the detection of the SARS-CoV-2 spike protein or fragment thereof is carried out on slides of test material (e.g., immunohistochemistry), by immunoblotting (e.g., Western blotting), surface plasmon resonance (e.g., Biacore), or by flow cytometry (FACS) analysis.

[0201] Other specific examples include, but are not limited to, enzyme immunoassay (EIA), radioimmunoassay (RIA), fluoroimmunoassay (FIA), chemiluminescent immunoassay (CLIA) and counting immunoassay (CIA), homogeneous enzyme-multiplied immunoassays (“EMIT”), apoenzyme reactivation immunoassay (“ARIS”), dipstick immunoassays, and immunochromatography assays. Most assays now use nonradioactive labels. Enzyme immunoassays (enzyme- linked immunosorbent assays, or ELISA; immunometric assays) can use enzymes as labels, such as, for example, horseradish peroxidase or alkaline phosphatase. Chemiluminescent immunoassays (CIA) can use luminol. Fluorimetric immunoassays (FIA) use fluorescent compounds (e.g., fluorescein) as labels.

[0202] The assays can be homogenous or heterogeneous assays, competitive and non-competitive assays. There are four main kinds of ELISA: sandwich, competitive, direct, and indirect assays. These methods differ in how the antibody or antigen is attached to the solid plate, and how the signal is detected. For example, in a sandwich ELISA, for example, an antibody is immobilized on a plate. The sample containing the target antigen is added, which binds to the antibody and so is immobilized on the plate. Next, a second type of antibody is added, which also binds to the target antigen on the plate, forming a ‘sandwich’ with the target antigen in the middle. The second antibody is linked to an enzyme, called a reporter enzyme, which allows the binding reaction to be measured by creating a color signal. To create this signal, first any unbound antibody is washed away, and a colorimetric substrate is added. The enzyme catalyzes a reaction of the substrate, creating a color change. A stronger color signal indicates more target antigen is present. An example of this is a home pregnancy test. In some embodiments, the first or second antibody is one of the disclosed antibodies, and the other antibody is one that detects SARS-CoV-2 RBD protein or fragment thereof, but may or may not detect its phosphorylated state, and thus may target a different antigen of the protein (e.g., a nonphosphorylated antigen).

[0203] In some embodiments, the assay is in the form of a sandwich assay, which is a noncompetitive immunoassay, wherein the molecule to be detected and / or quantified is bound to a first antibody and to a second antibody. The first antibody may be bound to a solid phase, e.g., a bead, a surface of a well or other container, a chip or a strip, and the second antibody is an antibody which is labeled, e.g. with a dye, with a radioisotope, or a reactive or catalytically active moiety. The amount of labeled antibody bound to the analyte is then measured by an appropriate method. The general compositionand procedures involved with “sandwich assays” are well-established and known to the skilled person.

[0204] Immunohistochemistry (IHC) is a process of localizing antigens (e.g., proteins) in tissue utilizing antigen- specific antibodies. The antigen-binding antibody can be conjugated or fused to a tag that allows its detection, e.g., via visualization. In some embodiments, the tag is an enzyme that can catalyze a color-producing reaction, such as alkaline phosphatase or horseradish peroxidase. The enzyme can be fused to the antibody or non-covalently bound, e.g., using a biotin-avidin system. Alternatively, the antibody can be tagged with a fluorophore, such as fluorescein, rhodamine, Dy Light Fluor or Alexa Fluor. The antigen-binding antibody can be directly tagged or it can itself be recognized by a detection antibody that carries the tag.

[0205] Quantitative immunochemical techniques can also be used. For example, the Quantitative Tissue Biomarker Platform from HistoRx and / or measuring immunofluorescence level(s) can be used to quantify levels of SARS-CoV-2 spike protein or a fragment thereof.

[0206] Western blotting can be used and can be quantitative or qualitative. A typical Western blotting procedure includes the steps of immunoprecipitating a target protein from a lysate of cells expressing the protein, performing an SDS-PAGE with said protein, transferring the protein to a nitrocellulose membrane, incubating the nitrocellulose membrane with said antibody, detecting said antibody with a secondary antibody conjugated to a fluorescent or chromogenic compound (e.g. peroxidases such as horseradish peroxidase (HRP), alkaline phosphatase (AP), IRDye near-infrared (NIR) fluorescent dyes), and quantifying the respective signal of said compound (e.g. fluorescence, luminescence, chromogenic enzyme substrate).

[0207] The ratio of two signals generated by Western blotting employing the same antibody but two different samples can be calculated, thereby determining how much more / less (fold-change) of the SARS-CoV-2 spike protein or fragment thereof is present in one sample compared to another.

[0208] In some embodiments, the methods provided herein involve determining the presence, absence, and / or concentration of SARS-CoV-2 spike protein or a fragment thereof in a cell, and / or the number of cells positive for SARS-CoV-2 spike protein or fragment thereof in a sample by fluorescence-activated cell sorting (FACS) using a flow cytometry device (e.g., Beckman Coulter Z2 Coulter Counter, Beckman Coulter Inc.). In some embodiments, a FACS-based method includes the step of preparing the output composition for detection by flow cytometry before the SARS-CoV-2 spike protein or fragment thereof can be detected. For example, the output composition can be incubated with a fluorescently labeled SARS-CoV-2 spike protein binder, and then the sample can be analyzed using a flow cytometer.

[0209] In some embodiments, the cells are permeabilized to facilitate antibody access to intracellular SARS-CoV-2 spike protein or fragment thereof. In flow cytometry, cells bound by fluorescently labeled affinity reagents are carried in a fluidic stream, are separated based on size and / or fluorescent signal, and are subsequently analyzed and counted using a FACS software program (e.g., FlowJo software). The number or approximate number of cells can be determined by detection of the fluorescent signal, which optionally can be determined or processed by the FACS software program to provide the total or approximate number of particles in the output composition.

[0210] In some embodiments, a sample is analyzed by means of a biochip. Biochips generally include solid substrates and have a generally planar surface, to which a capture reagent (also called an adsorbent or affinity reagent) is attached. Frequently, the surface of a biochip includes a plurality of addressable locations, each of which has the capture reagent bound there.

[0211] Protein biochips are biochips adapted for the capture of polypeptides. Many protein biochips are described in the art. These include; for example, protein biochips produced by Ciphergen Biosystems, Inc. (Fremont, Calif.), Packard BioScience Company (Meriden Conn.), Zyomyx (Hayward, Calif.), Phylos (Lexington, Mass.) and Biacore (Uppsala, Sweden).

[0212] Photonic biosensors can be used in label-free assays. For example, photonic biosensors combine photonic sensing with bio recognition technology to create label-free testing on-chip. Instead of moving electrons around on silicon chips, light is moved around on silicon chips via waveguides. This technology has allowed the development of miniature lab-on-a-chip label-free immunoassay (LFIA) devices. These devices are functionalized with capture antibodies and have a resonance condition of light. This resonance wavelength will be shifted by a reaction between the capture antibody and the target antigen due to the change in refractive index. Measuring the shift in resonance wavelength provides a readout of a binding event. Label-free assays therefore enable the detection of antigen-antibody binding without the use of an additional label, resulting in increased assay sensitivity and decreased working time.B. Methods of Diagnosis

[0213] The SARS-CoV-2 spike protein and fragments thereof can be used in diagnostic tests to assess SARS-CoV-2-related infections, diseases, and disorder status in a subject, e.g., to distinguish between normal cells and infected cells, and to evaluate disease status. For example, disease status includes, without limitation, the presence or absence of infection (e.g., SARS-CoV-2 infection versus noninfection), the risk of developing COVID-19, the stage of the disease, the progression of disease (e.g., worsening or remission over time), and the effectiveness or response to treatment. Based on this status, further procedures may be indicated, including additional diagnostic tests or therapeutic interventions.

[0214] The SARS-CoV-2 spike protein can be present and / or expressed in certain cells of a subject, including but not limited to those described herein, and therefore, detection thereof is useful in aiding in the determination of infection and / or other SARS-CoV-2-related conditions. An exemplary method involves, first, measuring SARS-CoV-2 spike protein or a fragment thereof in a subject sample using the methods described herein, and, second, comparing the measurement with a diagnostic amount or cut-off that distinguishes a positive SARS-CoV-2 infection or related disease status from a negative status. The diagnostic amount represents a measured amount of spike protein or a fragment thereof above which a subject is classified as having the infection or related disease. For example, if the spike protein level or viral load is elevated compared to non-infected individuals, then a measured amount above the diagnostic threshold provides a diagnosis or disease status. As is well understood in the art, by adjusting the particular diagnostic cut-off used in an assay, one can increase the sensitivity or specificity of the diagnostic test depending on clinical needs. The diagnostic cut-off can be determined, for example, by measuring the amount of SARS-CoV-2 spike protein or fragment thereof in a statistically significant number of samples from subjects and selecting a cut-off to meet the desired diagnostic performance.1. Determining Risk of Developing Disease

[0215] Methods for determining the risk of developing disease in a subject are also provided. S ARS- CoV-2 spike protein or fragment thereof amounts or expression patterns can be characteristic of various risk states, e.g., high, medium, or low. The risk of developing a disease (e.g., COVID-19 or post-acute sequelae) is determined by measuring the spike protein or fragment thereof and then either submitting the measurement to a classification algorithm or comparing it with a reference amount and / or pattern of spike protein or fragment that is associated with a particular risk level.2. Determining Stage of Disease

[0216] Another embodiment provides methods for determining the stage of SARS-CoV-2-related disease in a subject. Each stage of infection or disease may be characterized by a distinct amount or pattern of SARS-CoV-2 spike protein or fragment thereof. The stage of the disease is determined by measuring the spike protein or fragment and then either submitting the data to a classification algorithm or comparing it to a reference amount and / or pattern associated with particular stages (e.g., acute, subacute, recovery).3. Determining Course (Progression / Remission) of Infection / Disease

[0217] Still another embodiment provides methods for determining the course of SARS-CoV-2- related disease in a subject. Disease course refers to changes in disease status over time, including disease progression (worsening) and disease regression (improvement). Over time, the amounts or relative amounts (e.g., the pattern) of the SARS-CoV-2 spike protein or fragment thereof may change.This method involves measuring spike protein or fragment thereof in a subject at two or more different time points (e.g., a first time and a second time) and comparing the changes in amounts. The course of the disease is determined based on these comparisons. Similarly, this method is useful for assessing the response to treatment. If treatment is effective, the spike protein or fragment thereof will trend toward normal levels, whereas if treatment is ineffective, the levels will trend toward those associated with active or worsening disease.4. Subject Management

[0218] In certain embodiments, the method includes detection and / or analysis of SARS-CoV-2 spike protein or a fragment thereof, and further includes managing subject treatment based on the determined status. Such management includes the actions of a physician or clinician following a diagnosis of SARS-CoV-2 infection or other spike protein-related condition. For example, if a physician determines that a subject has a SARS-CoV-2-related disease, then an appropriate treatment regimen may follow, such as administration of antiviral agents, monoclonal antibody therapy, immunomodulators, or other relevant compositions, including those discussed in further detail below. Alternatively, if the diagnosis rules out SARS-CoV-2-related disease, additional tests may be conducted to determine the presence of other infections or conditions. In the event of inconclusive results, follow-up testing may be warranted.

[0219] One embodiment provides a method for selecting a subject for treatment of SARS-CoV-2- related disease or condition by detecting the presence or amount of spike protein or a fragment thereof in a biological sample. The level of spike protein is compared to a predetermined threshold or diagnostic cutoff, and the subject is selected for treatment if the spike protein or fragment thereof meets or exceeds that threshold. Suitable treatments may include those known to be effective for subjects with elevated or abnormal spike protein expression, including variant-specific therapeutics. Additional embodiments provide methods for excluding a SARS-CoV-2-related condition when the test result is negative, allowing for alternative treatment decisions and helping to avoid therapies not effective for spike-positive conditions.

[0220] Further embodiments relate to communication of diagnostic or assay results to clinicians, technicians, or patients. In some cases, computers may be used to deliver diagnostic results to relevant parties such as physicians and their patients. Assays may be conducted or analyzed in one country or jurisdiction, and results may be communicated across different jurisdictions.

[0221] In preferred embodiments, a diagnosis based on the presence or absence of SARS-CoV-2 spike protein or fragment thereof is communicated to the subject as soon as available. The result may be delivered by the subject’s treating physician, or via automated electronic communication (e.g., email, phone). In certain embodiments, a computer-based system may generate and transmitdiagnostic messages automatically using software and hardware systems familiar to those skilled in the art of telecommunications. In some embodiments, various method steps — including sample testing, disease diagnosis, and result reporting — may be performed across different national or regional jurisdictions.5. Screening Assays

[0222] The SARS-CoV-2 spike protein binders can be used to screen for compounds that modulate the expression or presence of the SARS-CoV-2 spike protein or fragments thereof, in vitro or in vivo. Such compounds may be useful in treating or preventing SARS-CoV-2-related diseases or disorders in patients. Candidate compounds for therapeutic use may initially be screened by identifying those that reduce the presence of spike protein or fragments thereof in cells associated with SARS-CoV-2 infection or related conditions.

[0223] Test compounds capable of modulating the presence and / or expression of the SARS-CoV-2 spike protein or fragments thereof may be administered to patients who are either experiencing symptoms of SARS-CoV-2 infection or are at risk of developing such symptoms. For example, administering a compound that reduces the expression or activity of the spike protein or fragments thereof may help decrease the likelihood or severity of SARS-CoV-2-related disease if elevated spike protein levels are associated with disease progression or pathology.

[0224] At the clinical level, screening a test compound may involve collecting biological samples from test subjects before and after administration of the compound. The levels of SARS-CoV-2 spike protein or fragments thereof in these samples can be measured and compared to assess whether the compound affects expression or presence of the protein. These measurements can be conducted using methods known in the art, including those described herein. In further embodiments, in vitro assays may be used to assess compound effects on spike protein expression, such as by exposing human cell lines expressing SARS-CoV-2 spike protein to test compounds. Subjects receiving such compounds may also be monitored for physiological or clinical outcomes associated with SARS-CoV-2. Test compounds may be evaluated for their ability to prevent infection, reduce viral load, or limit disease progression in subjects diagnosed with SARS-CoV-2-related disease.6. Assessing the Effectiveness of Treatment or Risk for Developing Symptoms of SARS-CoV-2 Infection and / or SARS-CoV-2 Spike Protein-Related Diseases and Disorders

[0225] Methods for determining the course of SARS-CoV-2 infection and / or other SARS-CoV-2 spike protein-related diseases and disorders in a subject are also provided. Disease course refers to changes in disease status over time, including disease progression (worsening) and disease regression (improvement). Over time, the amounts or relative amounts (e.g., the pattern) of SARS-CoV-2 spike protein or a fragment thereof may change. Accordingly, this method involves measuring SARS-CoV-2 spike protein or a fragment thereof in a subject at least two different time points, e.g., a first time and a second time, and comparing the change in amounts, if any. The course of disease is determined based on these comparisons. Similarly, this method is useful for determining the response to treatment. If a treatment is effective, then the SARS-CoV-2 spike protein or fragment thereof will trend toward normal levels, while if treatment is ineffective, the spike protein or fragment thereof will trend toward levels associated with ongoing or worsening disease.

[0226] In yet another example, the binders can be used in studies to determine if the subject is at risk for developing symptoms of SARS-CoV-2 infection and / or other spike protein-related diseases or disorders.C. Methods of Treatment

[0227] Methods of treatment are also provided and can be used alone or in combination with other methods disclosed herein, such as the disclosed methods of detection, diagnosis, prognosis, and treatment monitoring. The methods typically include administering to a subject in need thereof an effective amount of a disclosed composition, e.g., an anti-SARS2 spike protein binder or a chimeric antigen receptor (CAR) T cell formed therefrom, to treat the subject. This is particularly applicable where a disease or condition is characterized by increased expression or presence of the SARS-CoV- 2 spike protein. Additionally or alternatively, particularly where increased live SARS-CoV-2 viral load has been detected, antiviral agents can be employed.

[0228] Cell-mediated immune responses can be directed against SARS-CoV-2 spike protein antigens to treat viral infection. Similarly, spike protein antigens can be targeted by antibody conjugates, such as antibody-drug conjugates.

[0229] In cases where viral assembly occurs, the use of antiviral drugs to inhibit productive viral replication can also be considered for use alone or in combination with an immunotherapeutic approach.

[0230] An exemplary method involves treating a subject (e.g., a human) having a disease, disorder, or condition by administering to the subject an effective amount of a pharmaceutical composition including genetically modified cells including an anti-SARS2 spike protein binder, optionally in a bispecific format and / or conjugated to an active agent such as a drug (e.g., chemotherapeutic agent).

[0231] In some embodiments, the methods include administering cells (e.g., T cells) engineered to express recombinant SARS2 spike protein- specific CAR (SARS2-CAR) to a subject (e.g., a human) having a disease, disorder, or condition in an amount effective to treat the disease, disorder, or condition. For example, in some embodiments, the methods treat a disease or disorder associated with elevated expression or specific expression of the SARS2 spike protein or an antigenic fragment thereof by administering to the subject an effective amount of a pharmaceutical compositionincluding cells modified to express recombinant SARS2-CAR. In some forms, the methods treat a subject having a disease, disorder, or condition by administering to the subject an effective amount of a pharmaceutical composition including a genetically modified cell, wherein the cell is modified by introducing to the cell:

[0232] (i) a nucleic acid (e.g., vector or mRNA) encoding a SARS2-CAR; and (ii) causing the SARS2-CAR to be expressed by the cell.

[0233] The cell can have been isolated from the subject having the disease, disorder, or condition, or from a healthy donor, prior to genetic modification.

[0234] In some embodiments, the methods include administering a SARS2 spike protein binder to a subject (e.g., a human) having a disease, disorder, or condition in an amount effective to treat the disease, disorder, or condition. In some embodiments, the molecule or antibody includes a drug conjugated thereto and / or has antibody-dependent cell-mediated cytotoxicity (ADCC) activity, complement-dependent cytotoxicity (CDC) activity, and / or is a bispecific engager, optionally a T cell engager.

[0235] As used herein, the term “subject” refers to any member of the subphylum Chordata, including, but not limited to, humans and other primates, including non-human primates such as chimpanzees and other apes and monkey species; farm animals such as cattle, sheep, pigs, goats and horses; domestic mammals such as dogs and cats; laboratory animals including rodents such as mice, rats and guinea pigs; birds, including domestic, wild and game birds such as chickens, turkeys and other gallinaceous birds, ducks, geese, and the like. The term does not denote a particular age. Thus, both adult and newborn individuals are intended to be covered. The system described above is intended for use in any of the above vertebrate species, since the immune systems of all of these vertebrates operate similarly.

[0236] As used herein, the terms “pharmaceutically acceptable” or “pharmacologically acceptable” refer to a material which is not biologically or otherwise undesirable, i.e., the material may be administered to an individual in a formulation or composition without causing any unacceptable biological effects or interacting in a deleterious manner with any of the components of the composition in which it is contained. For example, the phrase “pharmaceutically acceptable” includes compositions, polymers and other materials and / or dosage forms which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of human beings and animals without excessive toxicity, irritation, allergic response, or other problem or complication, commensurate with a reasonable benefit / risk ratio.

[0237] As used herein, the phrase “pharmaceutically acceptable carrier” refers to pharmaceutically acceptable materials, compositions or vehicles, such as a liquid or solid filler, diluent, solvent orencapsulating material involved in carrying or transporting any subject composition, from one organ, or portion of the body, to another organ, or portion of the body. Each carrier must be “acceptable” in the sense of being compatible with the other ingredients of a subject composition and not injurious to the patient.

[0238] As used herein, the term “treatment” refers to the medical management of a patient with the intent to cure, ameliorate, stabilize, or prevent a disease, pathological condition, or disorder. This term includes active treatment, that is, treatment directed specifically toward the improvement of a disease, pathological condition, or disorder, and also includes causal treatment, that is, treatment directed toward removal of the cause of the associated disease, pathological condition, or disorder. In addition, this term includes palliative treatment, that is, treatment designed for the relief of symptoms rather than the curing of the disease, pathological condition, or disorder; preventative treatment, that is, treatment directed to minimizing or partially or completely inhibiting the development of the associated disease, pathological condition, or disorder; and supportive treatment, that is, treatment employed to supplement another specific therapy directed toward the improvement of the associated disease, pathological condition, or disorder. For example, “treatment” includes any of (i) the prevention of infection or reinfection with a coronavirus using neutralizing antibodies, (ii) the reduction or elimination of symptoms, and (iii) the substantial or complete elimination of infection of a coronavirus or variants thereof. Treatment can be prophylactic (prior to infection) or therapeutic (following infection).

[0239] As used herein, the term “therapeutically effective amount” refers to an amount of the therapeutic agent that, when incorporated into and / or onto particles described herein, produces some desired effect at a reasonable benefit / risk ratio applicable to any medical treatment. The effective amount may vary depending on such factors as the disease or condition being treated, the particular targeted constructs being administered, the size of the subject, or the severity of the disease or condition. One of ordinary skill in the art may empirically determine the effective amount of a particular compound without necessitating undue experimentation. In some embodiments, the term “effective amount” refers to an amount of a therapeutic agent or prophylactic agent to reduce or diminish the symptoms of one or more diseases or disorders of the brain, such as reducing tumor size (e.g., tumor volume).

[0240] In certain embodiments, the coronavirus is SARS-CoV-2. In another aspect, the coronavirus is SARS, MERS, 229E (alpha), NL63 (alpha), OC43 (beta), HKU1 (beta), SARS-CoV-2, or an emerging variant thereof. SARS-CoV-2 variants include the Wuhan parental sequence with or without the D614G mutation, Alpha (B.1.1.7 and Q lineages), Beta (B.1.351 and descendent lineages), Gamma (P.l and descendent lineages), Epsilon (B. 1.427 and B.1.429), Eta (B.1.525), Iota(B.1.526), Kappa (B.1.617.1), Mu (B.1.621, B.1.621.1), Zeta (P.2), Delta (B.l.617.2 and AY lineages), and Omicron (B. l. 1.529 or its subvariants, BA.l, BA.2, and BA.3). In another aspect, the mutant coronavirus spike proteins are formed into dimers, trimers, multimers, or nanoparticles. In another aspect, the nanoparticles comprise ferritin nanoparticles, polymeric nanoparticles, or both.

[0241] As used herein, the term “effective dose” refers to that amount of the antibodies or antigenbinding portions thereof of the present invention against SARS-CoV-2 sufficient to induce immunity, to prevent and / or ameliorate an infection or to reduce at least one symptom of an infection and / or to enhance the efficacy of another dose of a SARS-CoV-2 antibody, or prophylactic or therapeutic treatment. An effective dose may refer to the amount of the antibodies or antigen-binding portions thereof of the present invention against SARS-CoV-2 sufficient to delay or minimize the onset of an infection. An effective dose may also refer to the amount of the antibodies or antigen-binding portions thereof of the present invention against SARS-CoV-2 that provides a therapeutic benefit in the treatment or management of an infection. Further, an effective dose is the amount of the antibodies or antigen-binding portions thereof of the present invention against SARS-CoV-2, or in combination with other therapies, that provides a therapeutic benefit in the treatment or management of an infection. An effective dose may also be the amount sufficient to enhance a subject's (e.g., a human's) own immune response against subsequent exposure to SARS-CoV-2. Levels of immunity can be monitored, e.g., by measuring amounts of neutralizing secretory and / or serum antibodies, e.g., by plaque neutralization, complement fixation, enzyme-linked immunosorbent, or microneutralization assay. In the case of a vaccine, an “effective dose” is one that prevents disease and / or reduces the severity of symptoms.

[0242] As used herein, the term “immune stimulator” refers to a compound that enhances an immune response via the body's own chemical messengers (cytokines). These molecules comprise various cytokines, lymphokines and chemokines with immunostimulatory, immunopotentiating, and pro- inflammatory activities, such as interferons, interleukins (e.g., IL-1, IL-2, IL-3, IL-4, IL-12, IL-13); growth factors (e.g., granulocyte-macrophage (GM)-colony stimulating factor (CSF)); and other immunostimulatory molecules, such as macrophage inflammatory factor, Flt3 ligand, B7.1; B7.2, etc. The immune stimulator molecules can be administered with the antibodies or antigen-binding portions thereof of the present invention against SARS-CoV-2, or can be administered separately. Either the protein or an expression vector encoding the protein can be administered to produce an immunostimulatory effect.

[0243] As used herein, the term “protective immune response” or “protective response” refers to an immune response mediated by the antibodies or antigen-binding portions thereof of the presentinvention against SARS-CoV-2, which is exhibited by a vertebrate (e.g., a human), which prevents or ameliorates an infection or reduces at least one symptom thereof.

[0244] The invention further provides kits, including SARS-CoV-2 antibodies, nucleic acids, agents, drugs and pharmaceutical formulations, packaged into suitable packaging material, optionally in combination with instructions for using the kit components, e.g., instructions for performing a method of detection of the invention. In one non-limiting example, the antibody can be detected by an immunoassay selected from radioimmunoassay, enzyme-linked immunosorbent assay (ELISA), sandwich assays, Western blot, immunoprecipitation, immunohistochemistry, immunofluorescence, antibody microarray, dot blotting, and fluorescence-activated cell sorting (FACS).1. Infections / Diseases to Be Treated

[0245] The SARS-CoV-2 spike protein forms part of the infectious SARS-CoV-2 virus, which is implicated in COVID-19 infection and pathogenesis, and is associated with various stages and mechanisms of disease progression. The spike protein is essential for viral entry into host cells, particularly respiratory epithelial cells. It facilitates attachment to cellular receptors, such as angiotensin-converting enzyme 2 (ACE2), leading to viral uptake and systemic dissemination. The spike protein is implicated in the development of COVID- 19-associated respiratory distress, inflammation, and other systemic symptoms. As the spike protein mediates cell entry and viral fusion, it contributes to symptoms such as fever, cough, fatigue, and, in severe cases, acute respiratory distress syndrome (ARDS). It is further associated with immune dysregulation and cytokine storm observed in critical cases. The spike protein is also involved in immune evasion mechanisms, allowing the virus to persist within the host through antigenic variation and glycan shielding. In the absence of early treatment or immune priming (e.g., vaccination), the spike protein’s role in viral propagation may lead to progressive disease and severe complications.

[0246] Any of the disclosed compositions and methods can be used to detect, diagnose, prognose, and / or treat any stage, mechanism, symptom, or complication associated with SARS-CoV-2 infection, the spike protein, or spike protein-related diseases and disorders.

[0247] In some forms, the SARS-CoV-2 polypeptides (e.g., recombinant monoclonal antibodies) and pharmaceutical compositions thereof, can be used to treat a subject with a coronavirus infection or a subject at risk of developing one or more symptoms associated with a coronavirus infection. In some forms, the SARS-CoV-2 polypeptides (e.g., recombinant monoclonal antibodies) can be used to treat a subject having an elevated risk of developing one or more symptoms associated with a severe coronavirus infection, such as systemic inflammatory response syndrome, sepsis, or septic shock. In some forms, the methods of treatment with the pharmaceutical compositions containing the SARS- CoV-2 polypeptides (e.g., recombinant monoclonal antibodies) are based on determining that thesubject has one or more viral markers that are known in art to increase risk for the subject to develop moderate to severe infection with a coronavirus.

[0248] In some forms, when the pathogenic coronavirus is SARS-CoV-2, the SARS-CoV-2 variant can be a variant of the wild-type strain of the coronavirus. “Wild-type” as used herein, refers to the original strain of coronavirus considered to be the background strain of the coronavirus containing no major mutations.

[0249] In some forms, the coronavirus disease to be treated is COVID associated with SARS-CoV - 2 betacoronavirus of the subgenus Sarbecovirus. SARS-CoV-2 viruses share approximately 79% genome sequence identity with the SARS-CoV virus identified in 2003. The genome organization of SARS-CoV-2 viruses is shared with other betacoronaviruses; six functional open reading frames (ORFs) are arranged in order from 5’ to 3’: replicase (ORFla / ORFlb), spike (S), envelope (E), membrane (M) and nucleocapsid (N). In addition, seven putative ORFs encoding accessory proteins are interspersed between the structural genes.

[0250] Coronavirus species and representative viruses thereof include [representative virus (of species)]: SARSr-CoV BtKY72 (Severe acute respiratory syndrome-related coronavirus), SARS- CoV-2 (Severe acute respiratory syndrome-related coronavirus), SARSr-CoV RaTG13 (Severe acute respiratory syndrome-related coronavirus), SARS-CoV PC4-227 (Severe acute respiratory syndrome-related coronavirus), SARS-CoV (Severe acute respiratory syndrome-relatecl coronavirus), Bat-Hp-BetaCovC (Bat Hp-betacoronavirus Zhejiang2013), Ro-BatCoV GCCDC1 (Rousettus bat coronavirus GCCDC1), Ro-BatCoV HKU9 (Rousettus bat coronavirus HKU9), Ei- BatCoV C704 (Eidolon bat coronavirus C704), Pi-BatCoV HKU5 (Pipistrellus bat coronavirus HKU5), Ty-BatCoV HKU4 (Tylonycteris bar coronavirus HKU4), MERS-CoV (Middle East respiratory' syndrome -related coronavirus), EriCoV Eledgehog coronavirus), MHV (murine coronavirus), HCoV HKU1 (Human coronavirus HKU1), ChRCoV HKU24 (China Rattus coronavirus HKU24), ChRCovC HKU24 (Betacoronavirus 1), MrufCoV 2JL14 (Myodes coronavirus 2JL14), HCoV NL63 (Human coronavirus NL63), HCoV 229E (Human coronavirus 229E), and HCoV OC43 (Human coronavirus OC43). See, e.g., Coronaviridae Study Group of the International Committee on Taxonomy of Viruses, Nat Microbiol 2020. DOI: 10.1038 / s41564-020- 0695-z), which is specifically incorporated by reference in its entirety. In some embodiments, the coronavirus is a common cold coronavirus such as 229E, NL63, OC43, and HKU1.

[0251] In particularly preferred embodiments, the virus is a Severe acute respiratory syndrome- related virus, such as, SARSr-CoV BtKY72, SARS-CoV-2, SARSr-CoV RaTG13, SARS-CoV PC4- 227, or SARS-CoV, preferably one that infects humans such as SARS-CoV or SARS-CoV-2.

[0252] The sequence WIV04 / 2019, belonging to the GISAID S clade / PANGO A lineage I Nextstrain 19B clade, is thought to most closely reflect the sequence of the original SARS-CoV-2 infecting humans. It is known as "sequence zero", and is widely used as a reference sequence. Subsequent to the initial isolate from Wuhan, China, numerous SARS-CoV-2 virus sequence variants of WIV04 / 2019 have been identified, some of which may be of particular importance due to their potential for increased transmissibility, increased virulence, and reduced effectiveness of vaccines against them. SARS-CoV-2 having a variation of the WIV04 / 2019 sequence include, but are not limited to:

[0253] B.l.1.7 lineage (a.k.a. 201 / 501 Y. VI Variant of Concern (VOC) 202012 / 01). This variant has a mutation in the receptor binding domain (RBD) of the spike protein at position 501, where the amino acid asparagine (N) has been replaced with tyrosine (Y). The shorthand for this mutation is N501Y. This variant also has several other mutations, including: 69 / 70 deletion: occurred spontaneously many times and likely leads to a conformational change in the spike protein. P681H: near the S1 / S2 furin cleavage site, a site with high variability in coronaviruses. This mutation has also emerged spontaneously multiple times.

[0254] B.1.351 lineage (a.k.a. 20H / 501Y.V2). This variant has multiple mutations in the spike protein, including K417N, E484K, N501Y. Unlike the B.l.1.7 lineage detected in the UK, this variant does not contain the deletion at 69 / 70.

[0255] P.l lineage (a.k.a. 20J / 501Y.V3). The P.l variant is a branch off the B. 1.1.28 lineage that was first reported by the National Institute of Infectious Diseases (NIID) in Japan in four travelers from Brazil, sampled during routine screening at Haneda airport outside Tokyo. The P.l lineage contains three mutations in the spike protein receptor binding domain: K417T, E484K, and N501Y.

[0256] Other lineages and mutations of interest include, but are not limited to, B.1.1.207, B.1.429, B.1.427, B.1.525, and other two-mutation (e.g., N501T-G142D), or three-mutation (e.g., N501T- G142D-F486L) variants in the Spike protein. All of these lineages and sequence alternatives relative to the WIV04 / 2019 strain are also considered SARS-CoV-2 virus. Tn some embodiments, SARS- CoV-2 is a strain or isolate with elevated or potentially elevated risk for causing human disease relative to WIV04 / 2019. See, e.g., Science Brief, Emerging SARS-CoV-2 variants (CDC website, Updated Jan. 28, 2021), Horby, et al., “NERVTAG note on B.l.1.7 severity.” SAGE meeting report. January 21, 2021 ; Wu, et al. “mRNA-1273 vaccine induces neutralizing antibodies against spike mutants from global SARS-CoV-2 variants.” bioRxiv. Posted January 25, 2021 ; Xie, et al., “Neutralization of N501Y mutant SARS-CoV-2 by BNT162b2 vaccine-elicited sera.” bioRxiv. Posted January 7, 2021; Greaney, et al. “Comprehensive mapping of mutations to the SARS-CoV-2 receptor-binding domain that affect recognition by polyclonal human serum antibodies.” bioRxiv.[Preprint posted online January 4, 2021]; Weisblum, et al., “Escape from neutralizing antibodies by SARS-CoV-2 spike protein variants.” eLife 2020;9:e61312; Resende, at al. “Spike E484K mutation in the first SARS-CoV-2 reinfection case confirmed in Brazil,” 2020. [Posted on virological.org on January 10, 2021]

[0257] Various strains and isolates of the foregoing viruses are known and include the representative genomic sequences those sequences and accession numbers provided in, e.g., Coronaviridae Study Group of the International Committee on Taxonomy of Viruses, Nat Microbiol 2020. DOI: 10.1038 / s41564-020-0695-z), as well as NCBI and GISAID which provide hundreds of SARS-CoV- 2 sequences.

[0258] In some embodiments, the SARS-CoV-2 is of the B.1.1.7, B.1.351, P.l, B.1.1.207, B.1.429, B. 1.427, or B.1.525 lineage.

[0259] In an exemplary embodiment, the SARS-CoV-2 is isolate USA / CA_CDC_5574 / 2020, or another isolate sharing one or mutations therewith relative to the original Wuhan isolate. Under the nomenclature system introduced by GISAID (Global Initiative on Sharing All Influenza Data), SARS-CoV-2, isolate USA / CA_CDC_5574 / 2020 is assigned lineage B. l.1.7 and GISAID clade GR using Phylogenetic Assignment of Named Global Outbreak LINeages (PANGOLIN) tool (GISAID website, 3. Rambaut, et al., Nat. Microbiol. 5 (2020): 1403-1407. PubMed: 32669681 ; Mercatelli, et al., Front. Microbiol. (2020): doi.org / 10.3389 / fmicb.2020.01800. PubMed: 32793182. The complete genome of SARS-CoV-2, isolate USA / CA_CDC_5574 / 2020 has been sequenced (GISAID: EPI_ISL_751801). The following mutations are present in the clinical isolate: Spike A570D, Spike D614G, Spike D1118H, Spike H69del, Spike N501Y, Spike P681H, Spike S982A, Spike T716I, Spike V70del, Spike Y145del, M (Membrane protein) V70L, N (Nucleocapsid protein) D3L, N G204R, N R203K, N S235F, NS3 T223I, NS8 (Non- structural protein 8) Q27stop, NS8 R52I, NS8 Y73C, NSP3 (Non-structural protein 3) A890D, NSP3 I1412T, NSP3 T183I, NSP6 (Non-structural protein 6) F108del, NSP6 G107del, NSP6 S 106del, NSP12 (Non-structural protein 12) P323L, NSP13 (Non-structural protein 13) A454V, NSP13 K460R. One additional SNP in ORFl ab L3826F was reported in the deposited passage two virus, in comparison to the clinical specimen. See also BEI Resources, Catalog No. NR-54011, and its description, which is specifically incorporated by reference herein in its entirety.2. Subjects to Be Treated

[0260] Patients with SARS-CoV-2 infection can experience a range of clinical manifestations, from no symptoms to critical illness. In general, adults with SARS-CoV-2 infection can be grouped into the following severity of illness categories; however, the criteria for each category may overlap or vary across clinical guidelines and clinical trials, and a patient’s clinical status may change over time.

[0261] (i) Asymptomatic or pre-symptomatic infection: individuals who test positive for S ARS-CoV- 2 using a virologic test ( / '.<?., a nucleic acid amplification test or an antigen test) but who have no symptoms that are consistent with COVID-19.

[0262] (ii) Mild illness: individuals who have any of the various signs and symptoms of COVID-19 (e.g., fever, cough, sore throat, malaise, headache, muscle pain, nausea, vomiting, diarrhea, loss of taste and smell) but who do not have shortness of breath, dyspnea, or abnormal chest imaging.

[0263] (hi) Moderate Illness: Individuals who show evidence of lower respiratory disease during clinical assessment or imaging and who have an oxygen saturation (SpO2) >94% on room air at sea level.

[0264] (iv) Severe illness: individuals who have SpO2 <94% on room air at sea level, a ratio of arterial partial pressure of oxygen to fraction of inspired oxygen (PaO2 / FiO2) <300 mm Hg, a respiratory rate >30 breaths / min, or lung infiltrates >50%. These patients may experience rapid clinical deterioration. Oxygen therapy should be administered immediately using a nasal cannula or a high-flow oxygen device. If secondary bacterial pneumonia or sepsis is suspected, administer empiric antibiotics, re-evaluate the patient daily, and de-escalate or stop antibiotics if there is no evidence of bacterial infection.

[0265] (v) Critical illness: individuals who have acute respiratory distress syndrome, septic shock that may represent virus-induced distributive shock, cardiac dysfunction, an exaggerated inflammatory response, and / or exacerbation of underlying comorbidities. In addition to pulmonary disease, patients with critical illness may also experience cardiac, hepatic, renal, central nervous system, or thrombotic disease.

[0266] Patients with certain underlying comorbidities are at a higher risk of progressing to severe COVID- 19. These comorbidities include being aged >65 years; having cardiovascular disease, chronic lung disease, sickle cell disease, diabetes, cancer, obesity, or chronic kidney disease; being pregnant; being a cigarette smoker; being a transplant recipient; and receiving immunosuppressive therapy.

[0267] In some cases, patients with COVID-19 may have additional infections that are noted when they present for care or that develop during the course of treatment. These coinfections may complicate treatment and recovery. Older patients or those with certain comorbidities or immunocompromising conditions may be at higher risk for these infections.

[0268] In some forms, the SARS-CoV-2 polypeptides (e.g., recombinant monoclonal antibodies) and pharmaceutical compositions thereof, can be used to reduce the replication of SARS-CoV-2 variants that predispose the host to developing severe CO VID. In further forms, the pharmaceutical composition containing the SARS-CoV-2 polypeptides (e.g., recombinant monoclonal antibodies)can be used to treat patients having an elevated risk of developing one or more symptoms associated with severe COVID- 19 as a result of SARS-CoV-2 infection. In these cases, the patients carrying these SARS-CoV-2 variants are likely to develop one or more symptoms associated with severe illness, critical illness, and additional complications. The disclosed the pharmaceutical composition containing the SARS-CoV-2 polypeptides (e.g., recombinant monoclonal antibodies), can be used to treat a subject as at risk of developing severe COVID in order to reduce the infection and replication of the virus at least by 50%, 60%, 70%, 80%, 90%, 95%, 97%, or 99%.3. Modes of Administration

[0269] In some embodiments the methods administer the composition in combination with a pharmaceutically acceptable carrier. The compositions described herein can be conveniently formulated into pharmaceutical compositions composed of one or more of the compounds in association with a pharmaceutically acceptable carrier. See, e.g., Remington's Pharmaceutical Sciences, latest edition, by E.W. Martin Mack Pub. Co., Easton, PA, which discloses typical carriers and conventional methods of preparing pharmaceutical compositions that can be used in conjunction with the preparation of formulations of the therapeutics described herein and which is incorporated by reference herein. These most typically would be standard carriers for administration of compositions to humans. In one aspect, for humans and non-humans, these include solutions such as sterile water, saline, and buffered solutions at physiological pH. Other therapeutics can be administered according to standard procedures used by those skilled in the art.

[0270] The pharmaceutical compositions can include, but are not limited to, carriers, thickeners, diluents, buffers, preservatives, surface active agents and the like in addition to the therapeutic(s) of choice.

[0271] Pharmaceutical compositions can be administered to the subject in a number of ways depending on whether local or systemic treatment is desired, and on the area to be treated. Thus, for example, a pharmaceutical composition including modified cells, such as therapeutic T cells, can be administered as an intravenous infusion, or directly injected into a specific site. Moreover, a pharmaceutical composition can be administered to a subject as an ophthalmic solution and / or ointment to the surface of the eye, vaginally, rectally, intranasally, orally, by inhalation, or parenterally, for example, by intradermal, subcutaneous, intramuscular, intraperitoneal, intrarectal, intraarterial, intralymphatic, intravenous, intrathecal and intratracheal routes.

[0272] Parenteral administration, if used, is generally characterized by injection. Injectables can be prepared in conventional forms, either as liquid solutions or suspensions, solid forms suitable for solution or suspension in liquid prior to injection, or as emulsions. A more recently revised approach for parenteral administration involves use of a slow release or sustained release system such that aconstant dosage is maintained. See, e.g., U.S. Patent No. 3,610,795, which is incorporated by reference herein. Suitable parenteral administration routes include intravascular administration (e.g., intravenous bolus injection, intravenous infusion, intra-arterial bolus injection, intra-arterial infusion and catheter instillation into the vasculature); peri- and intra-tissue injection (e.g., intraocular injection, intra-retinal injection, or sub-retinal injection); subcutaneous injection or deposition including subcutaneous infusion (such as by osmotic pumps); direct application by a catheter or other placement device (e.g., an implant including a porous, non-porous, or gelatinous material).

[0273] Preparations for parenteral administration include sterile aqueous or non-aqueous solutions, suspensions, and emulsions which can also contain buffers, diluents and other suitable additives. Examples of non-aqueous solvents are propylene glycol, polyethylene glycol, vegetable oils such as olive oil, and injectable organic esters such as ethyl oleate. Aqueous carriers include water, alcoholic / aqueous solutions, emulsions or suspensions, including saline and buffered media. Parenteral vehicles include sodium chloride solution, Ringer's dextrose, dextrose and sodium chloride, lactated Ringer's, or fixed oils. Intravenous vehicles include fluid and nutrient replenishers, electrolyte replenishers (such as those based on Ringer's dextrose), and the like. Preservatives and other additives can also be present such as, for example, antimicrobials, anti-oxidants, chelating agents, and inert gases and the like.

[0274] Administration of the pharmaceutical compositions can be localized (i.e., to a particular region, physiological system, tissue, organ, or cell type) or systemic.4. Dosage Units and Effective Amounts

[0275] The methods for reducing the replication of a coronavirus infection, or methods for achieving a desired alleviation of coronavirus-associated disease symptoms, include administering to an animal, such as a mammal, especially a human being, an effective amount of a combination of a pharmaceutical composition comprising one or more caspase-6 inhibitor (s) and optionally one or more therapeutic, prophylactic or diagnostic agents, such as part of the same composition, or administered separately and independently at the same time or at different times (i.e., administration of the one or more therapeutic, prophylactic or diagnostic agents, and the SARS-CoV-2 polypeptides (e.g., recombinant monoclonal antibodies) is separated by a finite period of time from each other). Therefore, the term “combination” or “combined” is used to refer to either concomitant, simultaneous, or sequential administration of the SARS-CoV-2 polypeptides (e.g., recombinant monoclonal antibodies) and one or more optional therapeutic, prophylactic or diagnostic agents. The combinations can be administered either concomitantly (e.g., as an admixture), separately but simultaneously (e.g., via separate intravenous lines into the same subject; one agent is given orallywhile the other agent is given by infusion or injection, etc.), or sequentially (e.g., one agent is given first followed by the second).

[0276] Formulations of including the SARS-CoV-2 polypeptides (e.g., recombinant monoclonal antibodies) and one or more therapeutic, prophylactic, and / or diagnostic agents typically include an effective amount of an admixture of the SARS-CoV-2 polypeptides (e.g., recombinant monoclonal antibodies) and one or more therapeutic, prophylactic, and / or diagnostic agents. Effective amounts of the SARS-CoV-2 polypeptides (e.g., recombinant monoclonal antibodies)are provided herein. It will be appreciated that in some forms the effective amount of the SARS-CoV-2 polypeptides (e.g., recombinant monoclonal antibodies) and one or more therapeutic, prophylactic, and / or diagnostic agents is different from the amount that would be effective for the one or more therapeutic, prophylactic, and / or diagnostic agents to achieve the same result when administered in the absence of the SARS-CoV-2 polypeptides (e.g., recombinant monoclonal antibodies).

[0277] When used for treating a coronavirus infection in a subject, the amount of the SARS-CoV-2 polypeptides (e.g., recombinant monoclonal antibodies) present in a pharmaceutical dosage unit, or otherwise administered to a subject, can be the amount effective to reduce coronavirus replication in small airways and alveoli of the lungs. For example, the amount of the SARS-CoV-2 polypeptides (e.g., recombinant monoclonal antibodies) present in the pharmaceutical dosage unit, is administered to the subject in an amount effective to reduce coronavirus replication by 50% or more 24 hours following administration.

[0278] In some forms, the amount of the SARS-CoV-2 polypeptides (e.g., recombinant monoclonal antibodies )present in a pharmaceutical dosage unit, or otherwise administered to a subject, can be the amount effective to reduce bronchiolitis, alveolitis, and vasculitis. In some forms, the amount of the SARS-CoV-2 polypeptides (e.g., recombinant monoclonal antibodies )present in a pharmaceutical dosage unit, or otherwise administered to a subject, can be the amount effective to reduce to reduce bronchiolar epithelial cell death and desquamation, alveolar space mononuclear cell infiltration, protein rich fluid exudation, alveolar hemorrhage, damage to alveolar structure, pulmonary blood vessel wall inflammation and endothelium infiltration, focal alveolar septal congestion and perivascular infiltration, and lower alveolar space immune cells. In some forms, the amount of the SARS-CoV-2 polypeptides (e.g., recombinant monoclonal antibodies) present in a pharmaceutical dosage unit, or otherwise administered to a subject, can be the amount effective to reduce body weight loss and improve survival.

[0279] In some forms, the amount of the SARS-CoV-2 polypeptides (e.g., recombinant monoclonal antibodies) present in a pharmaceutical dosage unit, or otherwise administered to a subject, can be the amount effective to reduce the expression of one or more pro-inflammatory cytokines and / orchemokines selected from the group comprising IL6, ILip, TNFa, IFNy, IP10, IP6, MCP1, CXCL3, CXCL5, and / or CXCL9. In some forms, the amount of caspase-6 inhibitor (s) present in a pharmaceutical dosage unit, or otherwise administered to a subject, can be the amount effective to upregulates the expression of one or more IFN-signaling genes selected from the group comprising IFIT1, IFIT2, IFIT3, IFITM3, TRIM22, and / or OAS1 following treatment.

[0280] In some forms, the pharmaceutical composition containing the SARS-CoV-2 polypeptides (e.g., recombinant monoclonal antibodies) is administered to a subject in need thereof, to deliver the caspase-6 inhibitor in an amount between about 0.1 mg and about 1,000 mg, inclusive, preferably between about 0.5 mg and about 100 mg, inclusive, more preferably between about 1 mg and about 15 mg, inclusive, for example, 0.5 mg, 1 mg, 2 mg, 3 mg, 4 mg, 5 mg, 6 mg, 7 mg, 8 mg, 9 mg, 10 mg, 11 mg, 12 mg, 13 mg, 14 mg, and / or 15 mg.

[0281] A dosage regimen of the pharmaceutical composition containing the SARS-CoV-2 polypeptides (e.g., recombinant monoclonal antibodies) and optionally one or more therapeutic, prophylactic, and / or diagnostic agents, can include one or multiple administrations of the pharmaceutical composition.

[0282] The effective amount of the pharmaceutical composition will vary based on the active agent and from subject to subject, depending on the species, age, weight and general condition of the subject, the severity of the disorder being treated, and its mode of administration. Thus, it is not possible to specify an exact amount for every pharmaceutical composition. However, an appropriate amount can be determined by one of ordinary skill in the art using only routine experimentation given the teachings herein. For example, effective dosages and schedules for administering the pharmaceutical composition can be determined empirically. In some forms, the dosage ranges for the administration of the composition are those large enough to resolve mucosal hyper-reactivity throughout the respiratory tract.

[0283] Preferably, the dosage is not so large as to cause adverse side effects, such as unwanted crossreactions, anaphylactic reactions, and the like. Generally, the dosage will vary with the age, condition, and sex of the patient, route of administration, whether other drugs are included in the regimen, and the type, stage, and location of the disease to be treated. The dosage can be adjusted by the individual physician in the event of any counter-indications. It will also be appreciated that the effective dosage of the composition can increase or decrease over the course of a particular treatment. Changes in dosage can result and become apparent from the results of diagnostic assays.

[0284] Dosage can vary, and can be administered in one or more dose administrations daily, for one or several days. Guidance can be found in the literature for appropriate dosages for given classes of pharmaceutical products. Optimal dosing schedules can be calculated from measurements of drugaccumulation in the body of the subject or patient. Persons of ordinary skill can determine optimum dosages, dosing methodologies and repetition rates. Optimum dosages can vary depending on the relative potency of individual pharmaceutical compositions, and can generally be estimated based on EC50s found to be effective in in vitro and in vivo animal models.

[0285] In some forms, administration of the composition will be given as a long-term treatment regimen whereby pharmacokinetic steady state conditions will be reached.

[0286] For example, some forms, antibodies are packaged in a hermetically sealed container, such as an ampoule or sachette, indicating the quantity of antibody. In some forms, the antibodies are supplied as a dry sterilized lyophilized powder or water free concentrate in a hermetically sealed container and can be reconstituted, e.g., with water or saline to the appropriate concentration for administration to a subject. For example, antibodies can be supplied as a dry sterile lyophilized powder in a hermetically sealed container at a unit dosage of at least 5 mg, more preferably at least 10 mg, at least 15 mg, at least 25 mg, at least 35 mg, at least 45 mg, at least 50 mg, or at least 75 mg. The lyophilized antibodies can be stored at between 2 and 8°C in their original container and the antibodies can be administered within 12 hours, preferably within 6 hours, within 5 hours, within 3 hours, or within 1 hour after being reconstituted. In an alternative form, antibodies can be supplied in liquid form in a hermetically sealed container indicating the quantity and concentration of the antibody, fusion protein, or conjugated molecule. Preferably, the liquid form of the antibodies are supplied in a hermetically sealed container at least 1 mg / ml, more preferably at least 2.5 mg / ml, at least 5 mg / ml, at least 8 mg / ml, at least 10 mg / ml, at least 15 mg / ml, at least 25 mg / ml, at least 50 mg / ml, at least 100 mg / ml, at least 150 mg / ml, at least 200 mg / ml of the antibodies.

[0287] For antibodies, the dosage administered to a patient is typically 0.01 mg / kg to 100 mg / kg of the patient’s body weight. Preferably, the dosage administered to a patient is between 0.01 mg / kg and 20 mg / kg, 0.01 mg / kg and 10 mg / kg, 0.01 mg / kg and 5 mg / kg, 0.01 and 2 mg / kg, 0.01 and 1 mg / kg, 0.01 mg / kg and 0.75 mg / kg, 0.01 mg / kg and 0.5 mg / kg, 0.01 mg / kg to 0.25 mg / kg, 0.01 to 0.15 mg / kg, 0.01 to 0.10 mg / kg, 0.01 to 0.05 mg / kg, or 0.01 to 0.025 mg / kg of the patient’s body weight. In particular, the invention contemplates that the dosage administered to a patient is 0.2 mg / kg, 0.3 mg / kg, 1 mg / kg, 3 mg / kg, 6 mg / kg or 10 mg / kg. A dose as low as 0.01 mg / kg may show appreciable pharmacodynamic effects. Dose levels of 0.10 - 1 mg / kg are predicted to be most appropriate. Higher doses (e.g., 1-30 mg / kg) are also contemplated. Generally, human antibodies have a longer half-life within the human body than antibodies from other species due to the immune response to the foreign polypeptides. Thus, lower dosages of human antibodies and less frequent administration is often possible. Further, the dosage and frequency of administration of antibodiesmay be reduced by enhancing uptake and tissue penetration of the antibodies by modifications such as, for example, lipidation.

[0288] Injections and infusion of the disclosed compositions can be repeated as often and as many times as the patient can tolerate until the desired response is achieved. Thus, antibodies can also be administered once or multiple times at these dosages. The optimal dosage and treatment regime for a particular patient can be determined by one skilled in the art of medicine by monitoring the patient for signs of disease and adjusting the treatment accordingly. In some forms, the unit dosage is in a unit dosage form for intravenous injection. In some forms, the unit dosage is in a unit dosage form for oral administration. In some forms, the unit dosage is in a unit dosage form for inhalation. In some forms, the unit dosage is in a unit dosage form for subcutaneous injection.

[0289] In some forms, the pharmaceutical compositions are administered at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24 or 30 hours, or more than 30 hours, up to 36 or 48 hours prior to or after the detection of the coronavirus in the patient. In other forms, the pharmaceutical compositions are administered at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24 or 30 hours, or more than 30 hours, up to 36 or 48 hours prior to or after administering a separate therapeutic, prophylactic, or diagnostic agent. In certain forms, additive or more than additive effects of the administration of the pharmaceutical composition containing SARS-CoV-2 polypeptides (e.g., recombinant monoclonal antibodies) in combination with one or more therapeutic and / or prophylactic agent (s) is evident after one day, two days, three days, four days, five days, six days, one week, two weeks, three weeks, or more than three weeks following administration.

[0290] An effective amount of the pharmaceutical compositions and optionally one or more therapeutic and / or prophylactic agents can be administered as a single unit dosage (e.g., as dosage unit), or sub-therapeutic doses that are administered over a finite time interval. Such unit doses may be administered on a daily basis for a finite time period, such as up to 3 days, or up to 5 days, or up to 7 days, or up to 10 days, or up to 15 days or up to 20 days or up to 25 days, are all specifically contemplated.5. Combination Therapy

[0291] In some embodiments, the compositions are administered in combination with other therapeutic agents or treatment modalities. Any of the disclosed pharmaceutical compositions, such as anti-SARS2 spike protein binders or CAR-modified cells targeting SARS2, can be used alone or in combination with other therapeutic agents or treatment modalities, for example, antiviral therapies, immunomodulatory agents, or vaccination strategies. As used herein, “combination” or “combined” refer to either concomitant, simultaneous, or sequential administration of the therapeutics.

[0292] In some forms, the pharmaceutical compositions and other therapeutic agents are administered separately through the same route of administration. In other forms, the pharmaceutical compositions and other therapeutic agents are administered separately through different routes of administration. The combinations can be administered either concomitantly (e.g., as an admixture), separately but simultaneously (e.g., via separate intravenous lines into the same subject; one agent is given orally while the other agent is given by infusion or injection), or sequentially (e.g., one agent is given first followed by the second).

[0293] Examples of preferred additional therapeutic agents include other conventional or experimental therapies known in the art for treating SARS-CoV-2 infection or associated complications. The compositions and methods described herein may be used as a first-line therapy, second-line therapy, third-line therapy, or in combination with other types of therapies known in the art.

[0294] The disclosed pharmaceutical compositions and / or other therapeutic agents, procedures, or modalities can be administered during periods of active infection, or during a period of convalescence, remission, or post-infection complication management. The pharmaceutical compositions can be administered before, concurrently with, or after the additional treatment, or during ongoing monitoring or management of SARS-CoV-2-related disease or disorder. When administered in combination, the disclosed pharmaceutical compositions and the additional therapeutic agents (e.g., a second or third agent), or all, can be administered in an amount or dose that is higher, lower, or the same as the amount or dosage of each agent used individually, e.g., as a monotherapy. In certain forms, the administered amount or dosage of the disclosed pharmaceutical composition, the additional therapeutic agent (e.g., second or third agent), or all, is lower (e.g., at least 20%, at least 30%, at least 40%, or at least 50%) than the amount or dosage of each agent used individually (e.g., required to achieve the same therapeutic effect).IV. Kits

[0295] In one embodiment, a kit includes a SARS-CoV-2 antibody, subsequence or fragment and instructions for detecting the S2 domain of the SARS-CoV-2 Spike protein. In another embodiment, a kit includes a SARS-CoV-2 antibody, subsequence or fragment and instructions for treating a subject in need of treatment (e.g., a subject having a disease, disorder, pathology, or condition amendable or that may respond to treatment or therapy) with the SARS-CoV-2 antibody, subsequence or fragment.

[0296] The term "packaging material" refers to a physical structure housing the components of the kit. The packaging material can maintain the components sterilely, and can be made of material commonly used for such purposes (e.g., paper, corrugated fiber, glass, plastic, foil, ampules, etc.).The label or packaging insert can include appropriate written instructions, for example, a diagnostic or treatment method of the invention. Instructions can therefore include instructions for practicing any of the methods of the invention described herein. Thus, in various embodiments, a kit includes a label or packaging insert including instructions for practicing a method of the invention in solution, in vitro, in vivo, or ex vivo. In one aspect, the instructions include administering or delivering the anti-S2 domain of the Spike protein of SARS-CoV-2 antibody, subsequence or binding fragment thereof into a subject locally, regionally or systemically in a treatment or therapeutic method of the invention.

[0297] Instructions may additionally include indications of a satisfactory clinical endpoint or any adverse symptoms or complications that may occur. Instructions may further include storage information, expiration date, or any information required by regulatory agencies such as the Food and Drug Administration for use in a human subject.

[0298] The instructions may be on “printed matter”, e.g., on paper or cardboard within the kit, on a label affixed to the kit or packaging material or attached to a vial or tube containing a component of the kit. Instructions may comprise audio or video medium and additionally be included on a computer readable medium, such as a disk (floppy diskette or hard disk), optical CD such as CD- or DVD- ROM / RAM, magnetic tape, electrical storage media such as RAM and ROM and hybrids of these such as magnetic / optical storage media.

[0299] Invention kits can additionally include a buffering agent, a preservative, or a stabilizing agent. The kit can also include control components for assaying for activity, e.g., a control sample or a standard. Each component of the kit can be enclosed within an individual container or in a mixture and all of the various containers can be within single or multiple packages.

[0300] In accordance with the invention further provided are cell-free (e.g., in solution, in solid phase) and cell-based (e.g., in vitro or in vivo) methods of screening, detecting and identifying SARS- CoV-2. The methods can be performed in solution, in vitro using a biological material or sample, and in vivo, for example, a sample of cells from an animal. In one embodiment, a method includes contacting a biological material or sample with an antibody that binds to SARS-CoV-2 under conditions allowing binding of the antibody to SARS-CoV-2; and assaying for binding of the antibody to SARS-CoV-2. The binding of the antibody to SARS-CoV-2 detects the presence of SARS-CoV-2. In one aspect, SARS-CoV-2 is present in a cell or tissue. In another aspect, the biological material or sample is obtained from a mammalian subject.

[0301] As used herein, the term “contacting” when used in reference to a composition such as a protein (e.g., SARS-CoV-2 antibody), material, sample, or treatment, means a direct or indirect interaction between the composition (e.g., SARS-CoV-2 antibody) and the other referenced entity.A particular example of direct interaction is binding. A particular example of an indirect interaction is where the composition acts upon an intermediary molecule, which in turn acts upon the referenced entity. Thus, for example, contacting a cell (e.g., a lymphocyte) with the SARS-CoV-2 antibody includes allowing the antibody to bind to the cell (e.g., through binding to SARS-CoV-2), or allowing the antibody to act upon an intermediary that in turn acts upon the cell.

[0302] As used herein, the terms “assaying” and “measuring” and grammatical variations thereof are used interchangeably herein and refer to either qualitative or quantitative determinations, or both qualitative and quantitative determinations. When the terms are used in reference to binding, any means of assessing the relative amount, affinity or specificity of binding is contemplated, including the various methods set forth herein and known in the art. For example, SARS-CoV-2 antibody binding to SARS-CoV-2 can be assayed or measured by an ELISA assay.

[0303] The practice of the present invention employs, unless otherwise indicated, conventional methods of chemistry, biochemistry, molecular biology, immunology and pharmacology, within the skill of the art to make the antibodies or antigen-binding portions thereof of the present invention against SARS-CoV-2. Such techniques are explained fully in the literature. See, e.g., Remington’s Pharmaceutical Sciences, 18th Edition (Easton, Pa.: Mack Publishing Company, 1990); Methods In Enzymology (S. Colowick and N. Kaplan, eds., Academic Press, Inc.); and Handbook of Experimental Immunology, Vols. LIV (D. M. Weir and C. C. Blackwell, eds., 1986, Blackwell Scientific Publications); Sambrook, et al., Molecular Cloning: A Laboratory Manual (2nd Edition, 1989); Short Protocols in Molecular Biology, 4th ed. (Ausubel et al. eds., 1999, John Wiley & Sons); Molecular Biology Techniques: An Intensive Laboratory Course, (Ream et al., eds., 1998, Academic Press); PCR (Introduction to Biotechniques Series), 2nd ed. (Newton & Graham eds., 1997, Springer Verlag); Fundamental Virology, Second Edition (Fields & Knipe eds., 1991, Raven Press, New York), relevant portion incorporated herein by reference.

[0304] As embodied and broadly described herein, an aspect of the present disclosure relates to a pharmaceutical composition antibody comprising, consisting essentially of, or consisting of: a pharmaceutically acceptable excipient and a therapeutically effective amount of a monoclonal antibody or antigen-binding fragment thereof comprising a heavy chain variable region comprising an amino acid sequence at least about 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, or 100% identical to an amino acid sequence of SEQ ID NOS : 1 -29. In another aspect, the antibody light chain variable region comprises an amino acid sequence at least about 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, or 100% identical to an amino acid sequence of SEQ ID NOS:30-58. In alternative aspects, the antibody or antigen-binding fragment thereof comprises those heavy chain complementarity determining regions (CDR)l, CDR2, and CDR3 as set forth in any one of SEQ ID NOS 1-29 and / or those light chainCDR1, CDR2, and CDR3 as set forth in any one of SEQ ID NOS 30-58. In one aspect, the antibody or antigen-binding fragment that binds to a domain of SARS CoV-2 Spike protein. In another aspect, the antigen-binding fragment is a recombinant single-chain fragment variable (scFV) antibody, Fab fragment, F(ab’)2 fragment, or Fv fragment. In another aspect, the antibody or antigen-binding fragment is chimeric, humanized, fully human, bispecific, or multimeric. In another aspect, the antibody or antigen-binding fragment comprises an Fc portion mutated to at least one of: eliminate or enhance Fc Receptor (FcR) interactions to change a half-life, increase or decrease antibodydependent cellular cytotoxicity, or increase or decrease complement activation. In another aspect, the antibody or binding fragment thereof of further comprising a peptide linker in a hinge region between the variable and constant domains of the heavy chain. In another aspect, the antibody or antigenbinding fragment is adapted for administration or genetic delivery with an RNA or DNA sequence or vector encoding the antibody or antigen-binding fragment. In another aspect, the antibody or antigenbinding fragment is formulated for nasal, pulmonary, alveolar, intravenous, administration.

[0305] As embodied and broadly described herein, an aspect of the present disclosure relates to a method of treating SARS-CoV-2 in a subject in need thereof, the method antibody comprising, consisting essentially of, or consisting of: administering to the subject a monoclonal antibody or antigen-binding fragment thereof comprising those heavy chain complementarity determining regions (CDR)l, CDR2, and CDR3 as set forth in any one of SEQ ID NOS: 1-29, respectively; and light chain CDR1, CDR2, and CDR3 as set forth in any one of SEQ ID NOS:30-58, respectively. In one aspect, the antibody or antigen-binding fragment binds to a domain of SARS CoV-2 Spike protein. In another aspect, the antibody heavy chain variable region comprises an amino acid sequence at least about 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, or 100% identical to an amino acid sequence of SEQ ID NOS: 1-29. In another aspect, the antibody light chain variable region comprises an amino acid sequence at least about 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, or 100% identical to an amino acid sequence of SEQ ID NOS:30-58. In another aspect, the antigen-binding fragment is a recombinant single-chain fragment variable (scFV) antibody, Fab fragment, F(ab’)2 fragment, or Fv fragment. In another aspect, the antibody or antigen-binding fragment is chimeric, humanized, fully human, bispecific, or multimeric. In another aspect, the antibody or antigen-binding fragment comprises an Fc portion mutated to at least one of: eliminate or enhance Fc Receptor (FcR) interactions to change a half-life, increase or decrease antibody-dependent cellular cytotoxicity, or increase or decrease complement activation. In another aspect, the antibody or antigen-binding fragment is adapted for administration or genetic delivery with an RNA or DNA sequence or vector encoding the antibody or antigen-binding fragment.

[0306] As embodied and broadly described herein, an aspect of the present disclosure relates to a pharmaceutical device suitable for nasal or pulmonary delivery of monoclonal antibody or antigenbinding fragment thereof antibody comprising, consisting essentially of, or consisting of: heavy chain complementarity determining regions (CDR) 1 , CDR2, and CDR3 as set forth in any one of SEQ ID NOS:l-29, respectively; and light chain CDR1, CDR2, and CDR3 as set forth in any one of SEQ ID NOS:30-58, respectively, selected from an inhaler for liquids, a nebulizer, metered-dose inhaler, aerosol, and a dry powder inhaler. In one aspect, the antibody or antigen-binding fragment that binds to a domain of SARS CoV-2 Spike protein.

[0307] As embodied and broadly described herein, an aspect of the present disclosure relates to a method for detecting a Spike protein of SARS-CoV-2 (SARS2-S) antibody comprising, consisting essentially of, or consisting of: obtaining or having obtained a biological sample suspected of comprising a SARS-CoV-2 virus; contacting the biological sample with a monoclonal antibody or antigen-binding fragment thereof comprising heavy chain complementarity determining regions (CDR)l , CDR2, and CDR3 as set forth in any one of SEQ ID NOS: 1-29, respectively; and light chain CDR1, CDR2, and CDR3 as set forth in any one of SEQ ID NOS:30-58, respectively, wherein the antibody or antigen-binding fragment that binds to a domain of SARS CoV-2 Spike protein with a light chain of SEQ ID NO:9; and detecting binding of the antibody or antigen-binding fragment to the SARS-CoV-2 virus. In one aspect, the method further comprises the step of detecting the antibody or antigen-binding fragment is diagnostic for the detection of SARS-CoV-2 in the biological sample. In one aspect, the method further comprises detecting SARS-CoV-2 by performing an immunoassay on the biological sample from a subject; wherein the immunoassay uses an antibody or antigenbinding fragment comprising the monoclonal antibody or antigen-binding fragment thereof comprising heavy chain complementarity determining regions (CDR)l, CDR2, and CDR3 as set forth in any one of SEQ ID NOS: 1-29, respectively; and light chain CDR1, CDR2, and CDR3 as set forth in any one of SEQ ID NOS:30-58, respectively. In another aspect, the immunoassay is selected from radioimmunoassay, enzyme-linked immunosorbent assay (ELISA), sandwich assays, Western blot, immunoprecipitation, immunohistochemistry, immunofluorescence, antibody microarray, dot blotting, and fluorescence- activated cell sorting (FACS).

[0308] As embodied and broadly described herein, an aspect of the present disclosure relates to a kit for detection of a Spike protein of SARS-CoV-2 (SARS2-S) comprising a monoclonal antibody or antigen-binding fragment thereof antibody comprising, consisting essentially of, or consisting of: heavy chain complementarity determining regions (CDR) 1 , CDR2, and CDR3 as set forth in any one of SEQ ID NOS: 1-29, respectively; and light chain CDR1, CDR2, and CDR3 as set forth in any one of SEQ ID NOS:30-58, respectively, for isolation and / or detection of SARS-CoV-2 in one or moresample types of human or nonhuman origin, including: blood, plasma, serum, saliva, tears, cerebrospinal fluid, lymph, urine, feces, exhaled breath condensate, perspiration, amniotic fluid, exosomes, cell and tissue lysates.

[0309] As embodied and broadly described herein, an aspect of the present disclosure relates to a recombinant nucleic acid molecule encoding an antibody or antigen-binding fragment thereof encoding a monoclonal antibody or antigen-binding fragment thereof antibody comprising, consisting essentially of, or consisting of: heavy chain complementarity determining regions (CDR)l, CDR2, and CDR3 as set forth in any one of SEQ ID NOS: 1-29, respectively; and light chain CDR1, CDR2, and CDR3 as set forth in any one of SEQ ID NOS:30-58, respectively.

[0310] As embodied and broadly described herein, an aspect of the present disclosure relates to a recombinant expression vector antibody comprising, consisting essentially of, or consisting of: an expression control sequence operatively linked to the recombinant nucleic acid molecule encoding a monoclonal antibody or antigen-binding fragment thereof comprising heavy chain complementarity determining regions (CDR)l, CDR2, and CDR3 as set forth in any one of SEQ ID NOS:l-29; and light chain CDR1, CDR2, and CDR3 as set forth in any one of SEQ ID NOS: 30-58, respectively.

[0311] As embodied and broadly described herein, an aspect of the present disclosure relates to a host cell antibody comprising, consisting essentially of, or consisting of: the recombinant nucleic acid molecule encoding a monoclonal antibody or antigen-binding fragment thereof comprising heavy chain complementarity determining regions (CDR)l, CDR2, and CDR3 as set forth in any one of SEQ ID NOS:l-29, respectively; and light chain CDR1, CDR2, and CDR3 as set forth in any one of SEQ ID NQS:30-58, respectively.

[0312] As embodied and broadly described herein, an aspect of the present disclosure relates to a method of making an antibody or antigen-binding fragment of an antibody comprising, consisting essentially of, or consisting of: culturing a recombinant host cell comprising a recombinant expression construct comprising an expression control sequence operatively linked to a recombinant nucleic acid molecule encoding a monoclonal antibody or binding fragment thereof comprising heavy chain complementarity determining regions (CDR) 1 , CDR2, and CDR3 as set forth in any one of SEQ ID NOS:l-29; and light chain CDR1, CDR2, and CDR3 as set forth in any one of SEQ ID NOS:30-58, respectively, wherein the host cell produces the antibody or antigen-binding fragment of an antibody, and isolating the antibody or antigen-binding fragment of an antibody from the recombinant host cell.

[0313] It is contemplated that any embodiment discussed in this specification can be implemented with respect to any method, kit, reagent, or composition of the invention, and vice versa. Furthermore, compositions of the invention can be used to achieve methods of the invention.

[0314] It will be understood that particular embodiments described herein are shown by way of illustration and not as limitations of the invention. The principal features of this invention can be employed in various embodiments without departing from the scope of the invention. Those skilled in the art will recognize, or be able to ascertain using no more than routine experimentation, numerous equivalents to the specific procedures described herein. Such equivalents are considered to be within the scope of this invention and are covered by the claims.

[0315] All publications and patent applications mentioned in the specification are indicative of the level of skill of those skilled in the art to which this invention pertains. All publications and patent applications are herein incorporated by reference to the same extent as if each individual publication or patent application was specifically and individually indicated to be incorporated by reference.

[0316] The use of the word “a” or “an” when used in conjunction with the term “comprising” in the claims and / or the specification may mean “one,” but it is also consistent with the meaning of “one or more,” “at least one,” and “one or more than one.” The use of the term “or” in the claims is used to mean “and / or” unless explicitly indicated to refer to alternatives only or the alternatives are mutually exclusive, although the disclosure supports a definition that refers to only alternatives and “and / or.” Throughout this application, the term “about” is used to indicate that a value includes the inherent variation of error for the device, the method being employed to determine the value, or the variation that exists among the study subjects.

[0317] As used in this specification and claim(s), the words “comprising” (and any form of comprising, such as “comprise” and “comprises”), “having” (and any form of having, such as “have” and “has”), “including” (and any form of including, such as “includes” and “include”) or “containing” (and any form of containing, such as “contains” and “contain”) are inclusive or open-ended and do not exclude additional, unrecited elements or method steps. In embodiments of any of the compositions and methods provided herein, “comprising” may be replaced with “consisting essentially of’ or “consisting of”. As used herein, the phrase “consisting essentially of’ requires the specified integer(s) or steps as well as those that do not materially affect the character or function of the claimed invention. As used herein, the term “consisting” is used to indicate the presence of the recited integer (e.g., a feature, an element, a characteristic, a property, a method / process step or a limitation) or group of integers (e.g., feature(s), element(s), characteristic(s), propertie(s), method / process steps or limitation(s)) only.

[0318] The term “or combinations thereof’ as used herein refers to all permutations and combinations of the listed items preceding the term. For example, “A, B, C, or combinations thereof’ is intended to include at least one of: A, B, C, AB, AC, BC, or ABC, and if order is important in a particular context, also BA, CA, CB, CBA, BCA, ACB, BAC, or CAB. Continuing with this example, expresslyincluded are combinations that contain repeats of one or more item or term, such as BB, AAA, AB, BBC, AAABCCCC, CBBAAA, CAB ABB, and so forth. The skilled artisan will understand that typically there is no limit on the number of items or terms in any combination, unless otherwise apparent from the context.

[0319] As used herein, words of approximation such as, without limitation, “about”, "substantial" or "substantially" refers to a condition that when so modified is understood to not necessarily be absolute or perfect but would be considered close enough to those of ordinary skill in the art to warrant designating the condition as being present. The extent to which the description may vary will depend on how great a change can be instituted and still have one of ordinary skilled in the art recognize the modified feature as still having the required characteristics and capabilities of the unmodified feature. In general, but subject to the preceding discussion, a numerical value herein that is modified by a word of approximation such as “about” may vary from the stated value by at least ±1, 2, 3, 4, 5, 6, 7, 10, 12 or 15%.

[0320] Additionally, the section headings herein are provided for consistency with the suggestions under 37 CFR 1.77 or otherwise to provide organizational cues. These headings shall not limit or characterize the invention(s) set out in any claims that may issue from this disclosure. Specifically, and by way of example, although the headings refer to a “Field of Invention,” such claims should not be limited by the language under this heading to describe the so-called technical field. Further, a description of technology in the “Background of the Invention” section is not to be construed as an admission that technology is prior art to any invention(s) in this disclosure. Neither is the “Summary” to be considered a characterization of the invention(s) set forth in issued claims. Furthermore, any reference in this disclosure to “invention” in the singular should not be used to argue that there is only a single point of novelty in this disclosure. Multiple inventions may be set forth according to the limitations of the multiple claims issuing from this disclosure, and such claims accordingly define the invention(s), and their equivalents, that are protected thereby. In all instances, the scope of such claims shall be considered on their own merits in light of this disclosure, but should not be constrained by the headings set forth herein.

[0321] All of the compositions and / or methods disclosed and claimed herein can be made and executed without undue experimentation in light of the present disclosure. While the compositions and methods of this invention have been described in terms of preferred embodiments, it will be apparent to those of skill in the art that variations may be applied to the compositions and / or methods and in the steps or in the sequence of steps of the method described herein without departing from the concept, spirit and scope of the invention. All such similar substitutes and modifications apparentto those skilled in the art are deemed to be within the spirit, scope and concept of the invention as defined by the appended claims.

[0322] To aid the Patent Office, and any readers of any patent issued on this application in interpreting the claims appended hereto, applicants wish to note that they do not intend any of the appended claims to invoke paragraph 6 of 35 U.S.C. § 112, U.S.C. § 112 paragraph (f), or equivalent, as it exists on the date of filing hereof unless the words “means for” or “step for” are explicitly used in the particular claim.

[0323] For each of the claims, each dependent claim can depend both from the independent claim and from each of the prior dependent claims for each and every claim so long as the prior claim provides a proper antecedent basis for a claim term or element.

[0324] The disclosed invention can be further understood by the following numbered paragraphs: Paragraph 1 : A monoclonal antibody or antigen-binding fragment thereof comprising: a heavy chain complementarity determining regions (CDR) 1 , CDR2, and CDR3 as set forth in any one of SEQ ID NOS: 1-29, respectively; and light chain CDR1, CDR2, and CDR3 as set forth in any one of SEQ ID NOS:30-58, respectively.Paragraph 2: The monoclonal antibody or antigen-binding fragment thereof of paragraph 1 , wherein the antibody heavy chain variable region comprises an amino acid sequence at least about 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, or 100% identical to an amino acid sequence of SEQ ID NOS: 1-29.Paragraph 3 : The monoclonal antibody or antigen-binding fragment thereof of paragraph 1 , wherein the antibody light chain variable region comprises an amino acid sequence at least about 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, or 100% identical to an amino acid sequence of SEQ ID NOS:30-58.Paragraph 4: The monoclonal antibody or antigen-binding fragment thereof of any one of paragraphs 1 to 3, wherein the antigen-binding fragment is a recombinant single-chain fragment variable (scFV) antibody, Fab fragment, F(ab’)2 fragment, or Fv fragment.Paragraph 5 : The monoclonal antibody or antigen-binding fragment thereof of any one of paragraphs 1 to 4, wherein the antibody or antigen-binding fragment is chimeric, humanized, fully human, bispecific, or multimeric.Paragraph 6: The monoclonal antibody or antigen-binding fragment thereof of any one of paragraphs 1 to 5, wherein the antibody or antigen-binding fragment comprises an Fc portion mutated to at least one of: eliminate or enhance Fc Receptor (FcR) interactions to change a half-life, increase or decrease antibody-dependent cellular cytotoxicity, or increase or decrease complement activation. Paragraph 7 : The monoclonal antibody or antigen-binding fragment thereof of any one of paragraphs 1 to Error! Reference source not found., wherein the antibody or antigen-bindingfragment is adapted for administration or genetic delivery with an RNA or DNA sequence or vector encoding the antibody or antigen-binding fragment.Paragraph 8: The monoclonal antibody or antigen- binding fragment thereof of any one of paragraphs 1 to 7, further comprising a peptide linker in a hinge region between the variable and constant domains of the heavy chain.Paragraph 9: A pharmaceutical composition comprising a pharmaceutically acceptable excipient and a therapeutically effective amount of a monoclonal antibody or antigen-binding fragment thereof comprising: heavy chain complementarity determining regions (CDR)l, CDR2, and CDR3 as set forth in any one of SEQ ID NOS:l-29, respectively; and a light chain CDR1, CDR2, and CDR3 as set forth in any one of SEQ ID NOS:30-58, respectively.Paragraph 10: The pharmaceutical composition of paragraph 9, wherein the antibody heavy chain comprises an amino acid sequence at least about 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, or 100% identical to an amino acid sequence of SEQ ID NOS: 1-29.Paragraph 11 : The pharmaceutical composition of paragraph 9, wherein the antibody light chain comprises an amino acid sequence at least about 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, or 100% identical to an amino acid sequence of SEQ ID NOS:30-58.Paragraph 12: The pharmaceutical composition of any one of paragraphs 9 to 11, wherein the antigen-binding fragment is a recombinant single-chain fragment variable (scFV) antibody, Fab fragment, F(ab’)2 fragment, or Fv fragment.Paragraph 13: The pharmaceutical composition of any one of paragraphs 9 to 12, wherein the antibody or antigen-binding fragment is chimeric, humanized, fully human, bispecific, or multimeric. Paragraph 14: The pharmaceutical composition of any one of paragraphs 9 to 13, wherein the antibody or antigen-binding fragment comprises an Fc portion mutated to at least one of: eliminate or enhance Fc Receptor (FcR) interactions to change a half-life, increase or decrease antibodydependent cellular cytotoxicity, or increase or decrease complement activation.Paragraph 15: The pharmaceutical composition of any one of paragraphs 9 to 14, further comprising a peptide linker in a hinge region between the variable and constant domains of the heavy chain.Paragraph 16: The pharmaceutical composition of any one of paragraphs 9 to 14, wherein the antibody or antigen-binding fragment is adapted for administration or genetic delivery with an RNA or DNA sequence or vector encoding the antibody or antigen-binding fragment.Paragraph 17: The pharmaceutical composition of any one of paragraphs 9 to 14, wherein the antibody or antigen-binding fragment is formulated for nasal, pulmonary, alveolar, intravenous, administration.Paragraph 18: A method of treating SARS-CoV-2 in a subject in need thereof, the method comprising administering to the subject a monoclonal antibody or antigen-binding fragment thereof comprising: a heavy chain complementarity determining regions (CDR)l, CDR2, and CDR3 as set forth in any one of SEQ ID NOS: 1-29, respectively; and a light chain CDR1, CDR2, and CDR3 as set forth in any one of SEQ ID NOS:3Q-58, respectively.Paragraph 19: The method of paragraph 18, wherein the antibody heavy chain variable region comprises an amino acid sequence at least about 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, or 100% identical to an amino acid sequence of SEQ ID NOS: 1-29.Paragraph 20: The method of paragraph 19, wherein the antibody light chain variable region comprises an amino acid sequence at least about 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, or 100% identical to an amino acid sequence of SEQ ID NOS:30-58Paragraph 21: The method of any one of paragraphs 19 to 20, wherein the antigen-binding fragment is a recombinant single-chain fragment variable (scFV) antibody, Fab fragment, F(ab’)2 fragment, or Fv fragment.Paragraph 22: The method of any one of paragraphs 19 to 21, wherein the antibody or antigenbinding fragment is chimeric, humanized, fully human, bispecific, or multimeric.Paragraph 23: The method of any one of paragraphs 19 to 22, wherein the antibody or antigenbinding fragment comprises an Fc portion mutated to at least one of: eliminate or enhance Fc Receptor (FcR) interactions to change a half-life, increase or decrease antibody -dependent cellular cytotoxicity, or increase or decrease complement activation.Paragraph 24: The method of any one of paragraphs 19 to 23, wherein the antibody or antigenbinding fragment is adapted for administration or genetic delivery with an RNA or DNA sequence or vector encoding the antibody or antigen-binding fragment.Paragraph 25: A pharmaceutical device suitable for nasal or pulmonary delivery of monoclonal antibody or antigen-binding fragment thereof comprising: a heavy chain complementarity determining regions (CDR) 1 , CDR2, and CDR3 as set forth in any one of SEQ ID NOS: 1-29, respectively; and a light chain CDR1, CDR2, and CDR3 as set forth in any one of SEQ ID NOS:30-58, respectively, selected from an inhaler for liquids, a nebulizer, metered-dose inhaler, aerosol, and a dry powder inhaler.Paragraph 26: A method for detecting a Spike protein of SARS-CoV-2 (SARS2-S) comprising: obtaining or having obtained a biological sample suspected of comprising a SARS-CoV-2 virus; contacting the biological sample with a monoclonal antibody or antigen-binding fragment thereof comprising: a heavy chain complementarity determining regions (CDR) 1 , CDR2, and CDR3 as set forth in any one of SEQ ID NOS: 1-29, respectively; and a light chain CDR1, CDR2, and CDR3 as set forth in any one of SEQ ID NOS:30-58, respectively; and detecting binding of the antibody or antigen-binding fragment to the SARS-CoV-2 virus. Paragraph 27: The method of paragraph 26, further comprising the step of detecting the antibody or antigen-binding fragment is diagnostic for the detection of SARS-CoV-2 in the biological sample. Paragraph 28: The method of paragraph 26, further comprising detecting SARS-CoV-2 by performing an immunoassay on the biological sample from a subject; wherein the immunoassay uses an antibody or antigen-binding fragment of any of paragraphs 1 to 17.Paragraph 29: The method of paragraph 28, wherein the immunoassay is selected from radioimmunoassay, enzyme-linked immunosorbent assay (ELISA), sandwich assays, Western blot, immunoprecipitation, immunohistochemistry, immunofluorescence, antibody microarray, dot blotting, and fluorescence- activated cell sorting (FACS).Paragraph 30: A kit for detection of a Spike protein of SARS-CoV-2 (SARS2-S) comprising a monoclonal antibody or antigen-binding fragment thereof comprising: a heavy chain complementarity determining regions (CDR)l, CDR2, and CDR3 as set forth in any one of SEQ ID NOS: 1-29, respectively; and a light chain CDR1, CDR2, and CDR3 as set forth in any one of SEQ ID NOS:30-58, respectively, for isolation and / or detection of SARS-CoV-2 in one or more sample types of human or nonhuman origin, including: blood, plasma, serum, saliva, tears, cerebrospinal fluid, lymph, urine, feces, exhaled breath condensate, perspiration, amniotic fluid, exosomes, cell and tissue lysates.Paragraph 31 : A recombinant nucleic acid molecule encoding an antibody or antigen-binding fragment thereof of any of paragraphs 1 to 17.Paragraph 32: A recombinant expression vector comprising an expression control sequence operatively linked to the recombinant nucleic acid molecule of paragraph 31.Paragraph 33: A host cell comprising the recombinant nucleic acid molecule of paragraph 31.Paragraph 34: A method of making an antibody or antigen-binding fragment of an antibody comprising culturing a host cell comprising a recombinant expression construct comprising an expression control sequence operatively linked to a recombinant nucleic acid molecule encoding a monoclonal antibody or binding fragment thereof comprising heavy chain complementarity determining regions (CDR)l, CDR2, and CDR3 as set forth in any one of SEQ ID NOS: 1-29, respectively; and light chain CDR1, CDR2, and CDR3 as set forth in any one of SEQ ID NOS:30- 58, respectively, wherein the host cell produces the antibody or antigen-binding fragment of an antibody, and isolating the antibody or antigen-binding fragment of an antibody from the recombinant host cell.Paragraph 35: A molecule or antibody comprising an antigen binding region of an antibody comprising six complementarity determining regions (CDRs), that immunospecifically binds to a surface unit or a transmembrane unit of a SARS-CoV-2 receptor-binding protein (RBD), wherein one, two, three, four, five, or six of the CDRs is / are a CDR(s) of any one of SEQ ID NOS: 1-58, optionally the three CDRs of any one of SEQ ID NOs:l-29 in combination with the three CDRs of any one of SEQ ID NOS:30-58.Paragraph 36: A molecule or antibody comprising an antigen binding region of an antibody that immunospecifically binds SARS-CoV-2 receptor-binding protein (RBD), the antigen binding region comprising six complementarity determining regions (CDRs), wherein the CDRs include at least one CDR of the CDRs of anti-SARS-CoV-2 RBD antibodies: B_01, B_02, B_03, B_04, B_05, B_06, B_07, B_08, B_09, B_10, O_01, O_02, O_03, O_04, O_05, O_06, O_07, O_08, O_09, Q_10, O_l l, GL2_V01, GL2_V02, GL2_V03, GL2_V04, GL3_V01, GL3_V02, GL3_VO3, and GL3_V04, or a variant thereof with at least 60, 70, 75, 80, 85, 90, or 95% sequence identity thereto; with all remaining CDRs independently selected from anti- SARS-CoV-2 RBD antibodies: B_01, B_02, B_03, B_04, B_05, B_06, B_07, B_08, B_09, B_10, O_01, O_02, O_03, O_04, O_05, O_06, O_07, O_08, O_09, O_10, O_l l, GL2_V01, GL2_V02, GL2_V03, GL2_V04, GL3_V01, GL3_V02, GL3_V03, and GL3_V04, and variants thereof with at least 60, 70, 75, 80, 85, 90, or 95% sequence identity thereto. Paragraph 37: The molecule or antibody of paragraph 35 or 36, wherein the CDRs include three heavy chain variable region CDRs independently selected from the heavy chain variable region CDRs of anti-SARS-CoV-2 RBD antibodies B_01, B_02, B_03, B_04, B_05, B_06, B_07, B_08, B_09, B_10, Q_01, Q_02, O_03, Q_04, Q_05, Q_06, Q_07, Q_08, Q_09, O_10, O_l l, GL2_V01, GL2_V02, GL2_V03, GL2_V04, GL3_V01, GL3_V02, GL3_VO3, and GL3_V04, and variants thereof with at least 60, 70, 75, 80, 85, 90, or 95% sequence identity thereto in combination with three light chain variable region CDRs independently selected from the light chain variable region CDRs of anti-SARS-CoV-2 RBD antibodies B_01, B_02, B_03, B_04, B_05, B_06, B_07, B_08, B_09,B_10, O_01, O_02, O_03, O_04, O_05, O_06, O_07, O_08, O_09, O_10, O_l l, GL2_V01, GL2_V02, GL2_V03, GL2_V04, GL3_V01, GL3_V02, GL3_VO3, and GL3_V04, and variants thereof with at least 60, 70, 75, 80, 85, 90, or 95% sequence identity thereto.Paragraph 38: The molecule or antibody of any one of paragraphs 35-37, wherein the CDRs include the three heavy chain variable region CDRs of anti-SARS-CoV-2 RBD antibody B_01, B_02, B_03, B_04, B_05, B_06, B_07, B_08, B_09, B_10, O_01, O_02, O_03, O_04, O_05, O_06, O_07, O_08, O_09, O_10, O_l l, GL2_V01, GL2_V02, GL2_V03, GL2_V04, GL3_V01, GL3_V02, GL3_V03, and GL3_V04,or variants thereof with at least 60, 70, 75, 80, 85, 90, or 95% sequence identity thereto in combination with the three light chain variable region CDRs for anti-SARS-CoV-2 RBD antibody B_01, B_02, B_03, B_04, B_05, B_06, B_07, B_08, B_09, B_10, O_01, O_02, O_03, O_04, O_05, O_06, O_07, O_08, O_09, O_10, O_l l, GL2_V01, GL2_V02, GL2_V03, GL2_V04, GL3_V01, GL3_V02, GL3_V03, and GL3_V04 or variants thereof with at least 60, 70, 75, 80, 85, 90, or 95% sequence identity thereto.Paragraph 39: The molecule or antibody of any one of paragraphs 35-38, wherein the six CDRs are:(i) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody B_01 ;(ii) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody B_02;(iii) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody B_03;(iv) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody B_04;(v) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody B_05;(vi) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody B_06;(vii) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody B_07;(viii) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody B_08;(ix) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody B_09;(x) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody B_10;(xi) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody O_01;(xii) the three light chain and the three heavy chain CDRs of anti-S ARS2 RBD antibody O_02;(xiii) the three light chain and the three heavy chain CDRs of anti-S ARS2 RBD antibody O_03;(xiv) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody O_04;(xv) the three light chain and the three heavy chain CDRs of anti-S ARS2 RBD antibody O_05;(xvi) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody O_06;(xvii) the three light chain and the three heavy chain CDRs of anti-S ARS2 RBD antibody O_07; (xviii) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody O_08;(xix) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody O_09;(xx) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody O_10;(xxi) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody O_11 ;(xxii) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody GL2_V01;(xxiii) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody GL2_V02;(xxiv) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody GL2_V03;(xxv) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody GL2_V04;(xxvi) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody GL3_V01;(xxvii) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody GL3_V02;(xxviii) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody GL3_V03; or(xxix) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody GL3_V04.Paragraph 40: The molecule or antibody of any one of paragraphs 35-39, wherein the six CDRs are oriented and in the same orientation as in the anti-SARS-CoV-2 RBD antibody from which they were selected.Paragraph 41: The molecule or antibody of any one of paragraphs 35-40, wherein the heavy chain variable region and light chain variable regions of antibody B_01, B_02, B_03, B_04, B_05, B_06, B_07, B_08, B_09, B_10, O_01, O_02, O_03, O_04, O_05, O_06, O_07, O_08, O_09, O_10, O_l l, GL2_V01, GL2_V02, GL2_V03, GL2_V04, GL3_V01, GL3_V02, GL3_V03, and GL3_V04, are the heavy chain variable regions of B_01 , B_02, B_03, B_04, B_05, B_06, B_07, B_08, B_09, B_10, O_01, O_02, O_03, O_04, O_05, O_06, O_07, O_08, O_09, O_10, O_l l, GL2_V01, GL2_V02, GL2_V03, GL2_V04, GL3_V01, GL3_V02, GL3_V03, and GL3_V04 according to Tables 1 or variants thereof with at least 60, 70, 75, 80, 85, 90, or 95% sequence identity thereto, and the light chain variable regions of B_01, B_02, B_03, B_04, B_05, B_06, B_07, B_08, B_09, B_10, O_01, O_02, O_03, O_04, O_05, O_06, O_07, 0_08, O_09, 0_10, O_11, GL2_V01, GL2_V02, GL2_V03, GL2_V04, GL3_V01, GL3_V02, GL3_VO3, and GL3_V04 according to Table 2 or variants thereof with at least 60, 70, 75, 80, 85, 90, or 95% sequence identity thereto.Paragraph 42: The molecule or antibody of any one of paragraphs 35-41, wherein the molecule or antibody is an antibody.Paragraph 43: The molecule or antibody of paragraph 42, wherein the antibody is an intact antibody and functional antibody fragment or fusion protein.Paragraph 44: The molecule or antibody of paragraph 43, wherein the functional fragment or fusion protein is selected from Fab fragments, F(ab')2 fragments, Fab' fragments, Fv fragments, recombinant IgG (rlgG) fragments, single chain antibody fragments optionally single chain variable fragments (scFv), and single region antibodies optionally selected from sdAb, sdFv, and nanobody fragments. Paragraph 45: The molecule or antibody of any one of paragraphs 42-44, wherein the antibody is selected from intrabodies, peptibodies, chimeric antibodies, fully human antibodies, humanized antibodies, and heteroconjugate antibodies, and multispecific antibodies optionally selected bispecific antibodies, diabodies, triabodies, tetrabodies, tandem di-scFv, and tandem tri-scFv.Paragraph 46: The molecule or antibody of any one of paragraphs 42-45, wherein the antibody is an IgM, IgE, IgA, IgD, or IgG optionally an IgGl, IgG2, IgG3, or IgG4.Paragraph 47: The molecule or antibody of any one of paragraphs 35-46, wherein the molecule or antibody is detectably labeled or comprises a conjugated toxin, drug, receptor, enzyme, receptor ligand.Paragraph 48: A fusion protein comprising the molecule or antibody of any one of paragraphs 35-47 and heterologous amino acid sequence.Paragraph 49: A chimeric antigen receptor (CAR) polypeptide comprising the molecule or antibody of any one of paragraphs 35-48.Paragraph 50: The chimeric antigen receptor of paragraph 49 comprising an extracellular antigen binding region, a spacer region, a transmembrane region, and intracellular signaling region.Paragraph 51 : The chimeric antigen receptor of paragraph 50 comprising a co-stimulatory domain. Paragraph 52: A nucleic acid or acids encoding the molecule or antibody of any one of paragraphs 35-47, fusion protein of paragraph 48, or CAR of any one of paragraphs 49-51 .Paragraph 53: The nucleic acid or acids of paragraph 52 comprising an expression control sequence operably linked thereto, optionally wherein the expression control sequence comprises a promoter. Paragraph 54: A vector comprising the nucleic acid or acids of paragraph 52 or paragraph 53.Paragraph 55: A host cell comprising the molecule or antibody of any one of paragraphs 35-47, fusion protein of paragraphs 48, CAR of any one of paragraphs 49-51, nucleic acid or acids(s) of paragraphs 52-53, or vector of paragraph 54.Paragraph 56: A CAR immune cell comprising the CAR of any one of paragraphs 49-51, optionally wherein the immune cell is a T cell.Paragraph 57: A pharmaceutical composition comprising the CAR immune cells of paragraph 56, or the molecule or antibody of any one of paragraphs 35-47 optionally wherein the molecule or antibody comprises a drug conjugated thereto and / or has antibody-dependent cell-mediated cytotoxicity (ADCC) activity, or complement-dependent cytotoxicity activity (CDC), and / or is a bispecific immune cell engager.Paragraph 58: A method of treating a subject in need thereof comprising administering the subject an effective amount of pharmaceutical composition of paragraph 57.Paragraph 59: The method of paragraph 58, wherein the subject has a disease or disorder caused by or characterized by increased presence of SARS-CoV-2 spike protein or a fragment thereof.Paragraph 60: The method of paragraph 58 or 59, wherein the subject has a SARS-CoV-2 infection, is at risk for developing symptoms of SARS-CoV-2 infection, or complications caused by SARS- CoV-2 infection.Paragraph 61: The method of paragraph 60, wherein the subject has SARS-CoV-2 infection and the SARS-CoV-2 spike protein or fragment thereof is an antigen of the SARS-CoV-2 infection.Paragraph 62: A method of detecting SARS-CoV-2 spike protein or fragment thereof comprising contacting a biological sample with one or more molecules or antibodies of any one of paragraphs 35-47, and detecting binding between the molecule(s) or antibody(ies) and the SARS-CoV-2 spike protein or fragment thereof.Paragraph 63: The method of paragraph 62 further comprising determining that the sample includes increased SARS-CoV-2 spike protein or fragment thereof if the level of detected binding is higher in the biological sample than in a control.Paragraph 64: The method of paragraphs 62 or 63, wherein binding is detected by an immunoassay, immunohistochemistry, Western blotting, surface plasmon resonance, flow cytometry (FACS) analysis, or a biochip.Paragraph 65: The method of paragraph 64, wherein the immunoassay is selected from an enzyme immunoassay (EIA), radioimmunoassay (RIA), fluoroimmunoassay (FIA), chemiluminescent immunoassay (CLIA) and counting immunoassay (CIA), homogeneous enzyme-multiplied immunoassays (“EMIT”), apoenzyme reactivation immunoassay (“ARIS”), dipstick immunoassays, or immuno-chromatography assays.Paragraph 66: The method of any one of paragraphs 62-65, wherein the biological sample is cells, respiratory tract cells, a cell lysate or a fraction thereof, or a fluid, optionally blood, saliva, or urine. Paragraph 67: The method of paragraph 66, wherein the cells or cell lysate or fraction thereof are derived from a biopsy from a subject.Paragraph 68: The method of paragraph 67, wherein the biopsy sample contains, or is suspected to contain, SARS-CoV-2 infection.Paragraph 69: A method of diagnosing a subject with a SARS-CoV-2-related disease or disorder comprising detecting SARS-CoV-2 spike protein or fragment thereof according to the method of any one of paragraphs 62-68.Paragraph 70: The method of paragraph 69, wherein the SARS-CoV-2-related disease or disorder is a SARS-CoV-2 infection, is a risk for developing symptoms of SARS-CoV-2 infection, or complications caused by SARS-CoV-2 infection.Paragraph 71: The method of any one of paragraphs 62-70 further comprising treating the subject.Paragraph 72: The method of paragraph 71, wherein the treatment comprises a therapy effective for treating a SARS-CoV-2-related disease or disorder.Paragraph 73: The method of paragraphs 71 or 72, wherein the treatment comprises administering the subject an effective amount of the pharmaceutical composition of paragraph 56 and / or an antiviral.

Claims

WHAT IS CLAIMED IS:

1. A monoclonal antibody or antigen-binding fragment thereof comprising: a heavy chain complementarity determining regions (CDR) 1 , CDR2, and CDR3 as set forth in any one of SEQ ID NOS: 1-29, respectively; and light chain CDR1, CDR2, and CDR3 as set forth in any one of SEQ ID NOS:30-58, respectively.

2. The monoclonal antibody or antigen-binding fragment thereof of claim 1, wherein the antibody heavy chain variable region comprises an amino acid sequence at least about 90, 91 , 92, 93, 94, 95, 96, 97, 98, 99, or 100% identical to an amino acid sequence of SEQ ID NOS: 1-29.

3. The monoclonal antibody or antigen-binding fragment thereof of claim 1, wherein the antibody light chain variable region comprises an amino acid sequence at least about 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, or 100% identical to an amino acid sequence of SEQ ID NOS:30-58.

4. The monoclonal antibody or antigen-binding fragment thereof of claim 1, wherein the antigen-binding fragment is a recombinant single-chain fragment variable (scFV) antibody, Fab fragment, F(ab’)2 fragment, or Fv fragment.

5. The monoclonal antibody or antigen-binding fragment thereof of claim 1, wherein the antibody or antigen-binding fragment is chimeric, humanized, fully human, bispecific, or multimeric.

6. The monoclonal antibody or antigen-binding fragment thereof of claim 1, wherein the antibody or antigen-binding fragment comprises an Fc portion mutated to at least one of: eliminate or enhance Fc Receptor (FcR) interactions to change a half-life, increase or decrease antibodydependent cellular cytotoxicity, or increase or decrease complement activation.

7. The monoclonal antibody or antigen-binding fragment thereof of claim 1, wherein the antibody or antigen-binding fragment is adapted for administration or genetic delivery with an RNA or DNA sequence or vector encoding the antibody or antigen-binding fragment.

8. The monoclonal antibody or antigen-binding fragment thereof of claim 1, further comprising a peptide linker in a hinge region between the variable and constant domains of the heavy chain.

9. A pharmaceutical composition comprising a pharmaceutically acceptable excipient and a therapeutically effective amount of a monoclonal antibody or antigen-binding fragment thereof comprising: heavy chain complementarity determining regions (CDR)l, CDR2, and CDR3 as set forth in any one of SEQ ID NOS: 1-29, respectively; anda light chain CDR1, CDR2, and CDR3 as set forth in any one of SEQ ID NOS:30-58, respectively.

10. The pharmaceutical composition of claim 9, wherein the antibody heavy chain comprises an amino acid sequence at least about 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, or 100% identical to an amino acid sequence of SEQ ID NOS: 1-29.

11. The pharmaceutical composition of claim 9, wherein the antibody light chain comprises an amino acid sequence at least about 90, 91 , 92, 93, 94, 95, 96, 97, 98, 99, or 100% identical to an amino acid sequence of SEQ ID NOS:30-58.

12. The pharmaceutical composition of claim 9, wherein the antigen-binding fragment is a recombinant single-chain fragment variable (scFV) antibody, Fab fragment, F(ab’)z fragment, or Fv fragment.

13. The pharmaceutical composition of claim 9, wherein the antibody or antigen-binding fragment is chimeric, humanized, fully human, bispecific, or multimeric.

14. The pharmaceutical composition of claim 9, wherein the antibody or antigen-binding fragment comprises an Fc portion mutated to at least one of: eliminate or enhance Fc Receptor (FcR) interactions to change a half-life, increase or decrease antibody-dependent cellular cytotoxicity, or increase or decrease complement activation.

15. The pharmaceutical composition of claim 9, further comprising a peptide linker in a hinge region between the variable and constant domains of the heavy chain.

16. The pharmaceutical composition of claim 9, wherein the antibody or antigen-binding fragment is adapted for administration or genetic delivery with an RNA or DNA sequence or vector encoding the antibody or antigen-binding fragment.

17. The pharmaceutical composition of claim 9, wherein the antibody or antigen-binding fragment is formulated for nasal, pulmonary, alveolar, intravenous, administration.

18. A method of treating SARS-CoV-2 in a subject in need thereof, the method comprising administering to the subject a monoclonal antibody or antigen-binding fragment thereof comprising: a heavy chain complementarity determining regions (CDR)l, CDR2, and CDR3 as set forth in any one of SEQ ID NOS: 1-29, respectively; and a light chain CDR1, CDR2, and CDR3 as set forth in any one of SEQ ID NOS:30-58, respectively.

19. The method of claim 18, wherein the antibody heavy chain variable region comprises an amino acid sequence at least about 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, or 100% identical to an amino acid sequence of SEQ ID NOS: 1-29.

20. The method of claim 18, wherein the antibody light chain variable region comprises an amino acid sequence at least about 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, or 100% identical to an amino acid sequence of SEQ ID NOS:30-5821 . The method of claim 19, wherein the antigen-binding fragment is a recombinant single-chain fragment variable (scFV) antibody, Fab fragment, F(ab’)2 fragment, or Fv fragment.

22. The method of claim 19, wherein the antibody or antigen-binding fragment is chimeric, humanized, fully human, bispecific, or multimeric.

23. The method of claim 19, wherein the antibody or antigen-binding fragment comprises an Fc portion mutated to at least one of: eliminate or enhance Fc Receptor (FcR) interactions to change a half-life, increase or decrease antibody-dependent cellular cytotoxicity, or increase or decrease complement activation.

24. The method of claim 19, wherein the antibody or antigen-binding fragment is adapted for administration or genetic delivery with an RNA or DNA sequence or vector encoding the antibody or antigen-binding fragment.

25. A pharmaceutical device suitable for nasal or pulmonary delivery of monoclonal antibody or antigen-binding fragment thereof comprising: a heavy chain complementarity determining regions (CDR) 1 , CDR2, and CDR3 as set forth in any one of SEQ ID NOS: 1-29, respectively; and a light chain CDR1, CDR2, and CDR3 as set forth in any one of SEQ ID NQS:30-58, respectively, selected from an inhaler for liquids, a nebulizer, metered-dose inhaler, aerosol, and a dry powder inhaler.

26. A method for detecting a Spike protein of SARS-CoV-2 (SARS2-S) comprising: obtaining or having obtained a biological sample suspected of comprising a SARS-CoV-2 virus; contacting the biological sample with a monoclonal antibody or antigen-binding fragment thereof comprising: a heavy chain complementarity determining regions (CDR) 1 , CDR2, and CDR3 as set forth in any one of SEQ ID NOS: 1-29, respectively; anda light chain CDR1, CDR2, and CDR3 as set forth in any one of SEQ ID NOS:30-58, respectively; and detecting binding of the antibody or antigen-binding fragment to the SARS-CoV-2 virus.

27. The method of claim 26, further comprising the step of detecting the antibody or antigenbinding fragment is diagnostic for the detection of SARS-CoV-2 in the biological sample.

28. The method of claim 26, further comprising detecting SARS-CoV-2 by performing an immunoassay on the biological sample from a subject; wherein the immunoassay uses the antibody or antigen-binding fragment.

29. The method of claim 28, wherein the immunoassay is selected from radioimmunoassay, enzyme-linked immunosorbent assay (ELISA), sandwich assays, Western blot, immunoprecipitation, immunohistochemistry, immunofluorescence, antibody microarray, dot blotting, and fluorescence-activated cell sorting (FACS).

30. A kit for detection of a Spike protein of SARS-CoV-2 (SARS2-S) comprising a monoclonal antibody or antigen-binding fragment thereof comprising: a heavy chain complementarity determining regions (CDR) 1 , CDR2, and CDR3 as set forth in any one of SEQ ID NOS: 1-29, respectively; and a light chain CDR1, CDR2, and CDR3 as set forth in any one of SEQ ID NQS:30-58, respectively, for isolation and / or detection of SARS-CoV-2 in one or more sample types of human or nonhuman origin, including: blood, plasma, serum, saliva, tears, cerebrospinal fluid, lymph, urine, feces, exhaled breath condensate, perspiration, amniotic fluid, exosomes, cell and tissue lysates.

31. A recombinant nucleic acid molecule encoding an antibody or antigen-binding fragment thereof of claim 1.

32. A recombinant expression vector comprising an expression control sequence operatively linked to the recombinant nucleic acid molecule of claim 31.

33. A host cell comprising the recombinant nucleic acid molecule of claim 31.

34. A method of making an antibody or antigen-binding fragment of an antibody comprising culturing a host cell comprising a recombinant expression construct comprising an expression control sequence operatively linked to a recombinant nucleic acid molecule encoding a monoclonal antibody or binding fragment thereof comprising heavy chain complementarity determining regions (CDR)l, CDR2, and CDR3 as set forth in any one of SEQ ID NOS: 1-29, respectively; and light chain CDR1, CDR2, and CDR3 as set forth in any one of SEQ ID NOS:30-58, respectively, wherein the host cell produces the antibody or antigen-binding fragment of an antibody, and isolating the antibody or antigen-binding fragment of an antibody from the recombinant host cell.

35. A molecule or antibody comprising an antigen binding region of an antibody comprising six complementarity determining regions (CDRs), that immunospecifically binds to a surface unit or a transmembrane unit of a SARS-CoV-2 receptor-binding protein (RBD), wherein one, two, three, four, five, or six of the CDRs is / are a CDR(s) of any one of SEQ ID NOS: 1-58, optionally the three CDRs of any one of SEQ ID NOs:l-29 in combination with the three CDRs of any one of SEQ ID NOS:30-58.

36. A molecule or antibody comprising an antigen binding region of an antibody that immunospecifically binds to SARS-CoV-2 receptor-binding protein (RBD), the antigen binding region comprising six complementarity determining regions (CDRs), wherein the CDRs include at least one CDR of the CDRs of anti-S ARS-CoV-2 RBD antibodies: B_01 , B_02, B_03, B_04, B_05, B_06, B_07, B_08, B_09, B_10, Q_01, O_02, O_03, O_04, O_05, O_06, O_07, O_08, O_09, O_10, O_ll, GL2_V01, GL2 V02, GL2_V03, GL2_V04, GL3_V01, GL3_V02, GL3_V03, and GL3_V04, or a variant thereof with at least 60, 70, 75, 80, 85, 90, or 95% sequence identity thereto; with all remaining CDRs independently selected from anti- SARS-CoV-2 RBD antibodies: B_01, B_02, B_03, B_04, B_05, B_06, B_07, B_08, B_09, B_10, O_01, O_02, O_03, O_04, O_05, O_06, O_07, O_08, O_09, O_10, O_H, GL2_V01, GL2_V02, GL2_V03, GL2_V04, GL3_V01, GL3_V02, GL3_V03, and GL3_V04, and variants thereof with at least 60, 70, 75, 80, 85, 90, or 95% sequence identity thereto.

37. The molecule or antibody of claim 35, wherein the CDRs include three heavy chain variable region CDRs independently selected from the heavy chain variable region CDRs of anti-SARS- CoV-2 RBD antibodies B_01, B_02, B_03, B_04, B_05, B_06, B_07, B_08, B_09, B_10, Q_01, O_02, O_03, O_04, O_05, O_06, O_07, O_08, O_09, O_10, O_l l, GL2_V01, GL2_V02, GL2_V03, GL2_V04, GL3_V01, GL3_V02, GL3_VO3, and GL3_V04, and variants thereof with at least 60, 70, 75, 80, 85, 90, or 95% sequence identity thereto in combination with three light chain variable region CDRs independently selected from the light chain variable region CDRs of anti-S ARS-CoV-2 RBD antibodies B_01 , B_02, B_03, B_04, B_05, B_06, B_07, B_08, B_09, B_10, O_01, O_02, O_03, O_04, O_05, O_06, O_07, O_08, O_09, O_10, O_l l, GL2_V01, GL2_V02, GL2_V03, GL2_V04, GL3_V01, GL3_V02, GL3_VO3, and GL3_V04, and variants thereof with at least 60, 70, 75, 80, 85, 90, or 95% sequence identity thereto.

38. The molecule or antibody of claim 35, wherein the CDRs include the three heavy chain variable region CDRs of anti-SARS-CoV-2 RBD antibody B_01, B_02, B_03, B_04, B_05, B_06, B_07, B_08, B_09, B_10, O_01, O_02, O_03, O_04, O_05, O_06, O_07, O_08, O_09, O_10, O_l l, GL2_V01, GL2_V02, GL2_V03, GL2_V04, GL3_V01, GL3_V02, GL3_V03, and GL3_V04,or variants thereof with at least 60, 70, 75, 80, 85, 90, or 95% sequence identity theretoin combination with the three light chain variable region CDRs for anti-SARS-CoV-2 RBD antibody B_01, B_02, B_03, B_04, B_05, B_06, B_07, B_08, B_09, B_10, O_01, O_02, O_03, O_04, O_05, O_06, O_07, O_08, O_09, O_10, O_l l, GL2_V01, GL2_V02, GL2_V03, GL2_V04, GL3_V01, GL3_V02, GL3_V03, and GL3_V04 or variants thereof with at least 60, 70, 75, 80, 85, 90, or 95% sequence identity thereto.

39. The molecule or antibody of claim 35, wherein the six CDRs are:(i) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody B_01 ;(ii) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody B_02;(iii) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody B_03;(iv) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody B_04;(v) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody B_05;(vi) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody B_06;(vii) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody B_07;(viii) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody B_08;(ix) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody B_09;(x) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody B_10;(xi) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody O_01 ;(xii) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody O_02;(xiii) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody O_03;(xiv) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody O_04;(xv) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody O_05;(xvi) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody O_06;(xvii) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody O_07; (xviii) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody O_08;(xix) the three light chain and the three heavy chain CDRs of anti-S ARS2 RBD antibody O_09;(xx) the three light chain and the three heavy chain CDRs of anti-S ARS2 RBD antibody O_10;(xxi) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody O_11 ;(xxii) the three light chain and the three heavy chain CDRs of anti-S ARS2 RBD antibody GL2_V01;(xxiii) the three light chain and the three heavy chain CDRs of anti-S ARS2 RBD antibody GL2_V02;(xxiv) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody GL2_V03;(xxv) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody GL2_V04;(xxvi) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody GL3_V01;(xxvii) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody GL3_V02;(xxviii) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody GL3_V03; or(xxix) the three light chain and the three heavy chain CDRs of anti-SARS2 RBD antibody GL3_V04.

40. The molecule or antibody of claim 39, wherein the six CDRs are oriented and in the same orientation as in the anti-SARS-CoV-2 RBD antibody from which they were selected.41 . The molecule or antibody of claim 40, wherein the heavy chain variable region and light chain variable regions of antibody B_01, B_02, B_03, B_04, B_05, B_06, B_07, B_08, B_09, B_10, O_01, O_02, O_03, O_04, O_05, O_06, O_07, O_08, O_09, O_10, O_l l, GL2_V01, GL2_V02, GL2_V03, GL2_V04, GL3_V01, GL3_V02, GL3_V03, and GL3_V04, are the heavy chain variable regions of B_01, B_02, B_03, B_04, B_05, B_06, B_07, B_08, B_09, B_10, O_01, O_02, O_03, O_04, O_05, O_06, O_07, O_08, O_09, O_10, O_l l, GL2_V01, GL2_V02, GL2_V03, GL2_V04, GL3_V01, GL3_V02, GL3_VO3, and GL3_V04 according to Tables 1 or variants thereof with at least 60, 70, 75, 80, 85, 90, or 95% sequence identity thereto, and the light chain variable regions of B_01, B_02, B_03, B_04, B_05, B_06, B_07, B_08, B_09, B_10, O_01, O_02, O_03, O_04, O_05, O_06, O_07, O_08, O_09, O_10, O_l l, GL2_V01, GL2_V02, GL2_V03, GL2_V04, GL3_V01, GL3_V02, GL3_VO3, and GL3_V04 according to Table 2 or variants thereof with at least 60, 70, 75, 80, 85, 90, or 95% sequence identity thereto.

42. The molecule or antibody of claim 35, wherein the molecule or antibody is an antibody.

43. The molecule or antibody of claim 42, wherein the antibody is an intact antibody and functional antibody fragment or fusion protein.

44. The molecule or antibody of claim 43, wherein the functional fragment or fusion protein is selected from Fab fragments, F(ab')2 fragments, Fab’ fragments, Fv fragments, recombinant IgG (rlgG) fragments, single chain antibody fragments optionally single chain variable fragments (scFv), and single region antibodies optionally selected from sdAb, sdFv, and nanobody fragments.

45. The molecule or antibody of claim 42, wherein the antibody is selected from intrabodies, peptibodies, chimeric antibodies, fully human antibodies, humanized antibodies, andI l lheteroconjugate antibodies, and multispecific antibodies optionally selected bispecific antibodies, diabodies, triabodies, tetrabodies, tandem di-scFv, and tandem tri-scFv.

46. The molecule or antibody of claim 42, wherein the antibody is an IgM, IgE, IgA, IgD, or IgG optionally an IgGl, IgG2, IgG3, or IgG4.

47. The molecule or antibody of claim 42, wherein the molecule or antibody is detectably labeled or comprises a conjugated toxin, drug, receptor, enzyme, receptor ligand.

48. A fusion protein comprising the molecule or antibody of any one of claims 35-47 and heterologous amino acid sequence.

49. A chimeric antigen receptor (CAR) polypeptide comprising the molecule or antibody of any one of claims 35-47.

50. The chimeric antigen receptor of claim 49 comprising an extracellular antigen binding region, a spacer region, a transmembrane region, and intracellular signaling region.51 . The chimeric antigen receptor of claim 50 comprising a co-stimulatory domain.

52. A nucleic acid or acids encoding the molecule or antibody of any one of claims 35-47.

53. The nucleic acid or acids of claim 52 comprising an expression control sequence operably linked thereto, optionally wherein the expression control sequence comprises a promoter.

54. A vector comprising the nucleic acid or acids of claim 53.

55. A host cell comprising the molecule or antibody of claim 53.

56. A CAR immune cell comprising the CAR of 49, optionally wherein the immune cell is a T cell.

57. A pharmaceutical composition comprising the CAR immune cells of any one of claims 35- 47, or the molecule or antibody of any one of claims 35-47 optionally wherein the molecule or antibody comprises a drug conjugated thereto and / or has antibody-dependent cell-mediated cytotoxicity (ADCC) activity, or complement-dependent cytotoxicity activity (CDC), and / or is a bispecific immune cell engager.

58. A method of treating a subject in need thereof comprising administering the subject an effective amount of pharmaceutical composition of claim 57.

59. The method of claim 58, wherein the subject has a disease or disorder caused by or characterized by increased presence of SARS-CoV-2 spike protein or a fragment thereof.

60. The method of claims 58, wherein the subject has a SARS-CoV-2 infection, is at risk for developing symptoms of SARS-CoV-2 infection, or complications caused by SARS-CoV-2 infection.

61. The method of claim 60, wherein the subject has SARS-CoV-2 infection and the SARS- CoV-2 spike protein or fragment thereof is an antigen of the SARS-CoV-2 infection.

62. A method of detecting SARS-CoV-2 spike protein or fragment thereof comprising contacting a biological sample with one or more molecules or antibodies of any one of claims 35- 47, and detecting binding between the molecule(s) or antibody(ies) and the SARS-CoV-2 spike protein or fragment thereof.

63. The method of claim 62 further comprising determining that the sample includes increased SARS-CoV-2 spike protein or fragment thereof if the level of detected binding is higher in the biological sample than in a control.

64. The method of claims 62, wherein binding is detected by an immunoassay, immunohistochemistry, Western blotting, surface plasmon resonance, flow cytometry (FACS) analysis, or a biochip.

65. The method of claim 64, wherein the immunoassay is selected from an enzyme immunoassay (EIA), radioimmunoassay (RIA), fluoroimmunoassay (FIA), chemiluminescent immunoassay (CLIA) and counting immunoassay (CIA), homogeneous enzyme-multiplied immunoassays (“EMIT”), apoenzyme reactivation immunoassay (“ARIS”), dipstick immunoassays, or immuno-chromatography assays.

66. The method of claim 62, wherein the biological sample is cells, respiratory tract cells, a cell lysate or a fraction thereof, or a fluid, optionally blood, saliva, or urine.

67. The method of claim 66, wherein the cells or cell lysate or fraction thereof are derived from a biopsy from a subject.

68. The method of claim 67, wherein the biopsy sample contains, or is suspected to contain, SARS-CoV-2 infection.

69. A method of diagnosing a subject with a SARS-CoV-2-related disease or disorder comprising detecting SARS-CoV-2 spike protein or fragment thereof according to the method of claim 62.

70. The method of claim 69, wherein the SARS-CoV-2-related disease or disorder is a SARS- CoV-2 infection, is a risk for developing symptoms of SARS-CoV-2 infection, or complications caused by SARS-CoV-2 infection.

71. The method of claim 62, further comprising treating the subject.

72. The method of claim 71 , wherein the treatment comprises a therapy effective for treating a SARS-CoV-2-related disease or disorder.

73. The method of claim 71, wherein the treatment comprises administering the subject an effective amount of the pharmaceutical composition of claim 56 and / or an antiviral.

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