Compositions and methods involving antibodies that bind to covalent peptide conjugates

Abstract

Compositions and methods are provided that include binding partners that specifically bind to peptide conjugate / MHC complexes, including peptide conjugates formed by covalent reaction of a targeted covalent inhibitor with a peptide. The binding partners are provided as antibodies and antibody derivatives that specifically bind to the peptide conjugate / MHC complex.

Description

CROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims the benefit of U.S. Provisional Application No. 63 / 366,819, filed June 22, 2022, U.S. Provisional Application No. 63 / 369,702, filed July 28, 2022, U.S. Provisional Application No. 63 / 402,606, filed August 31, 2022, U.S. Provisional Application No. 63 / 377,466, filed September 28, 2022, and U.S. Provisional Application No. 63 / 485,788, filed February 17, 2023, each of which is incorporated by reference in its entirety.

[0002] Statement Regarding Federally Sponsored Research This invention was made with government support under Grant Nos. CA016087 and CA267362 awarded by the National Institutes of Health. The government has certain rights in this invention. Background

[0003] There is an ongoing unmet need for target-binding agents, including drugs covalently attached to proteins and peptides. In particular, there is a need to improve the efficacy of targeted therapies, as well as to enhance tumor immunogenicity and improve the efficacy of immunotherapy against cancers driven by intracellular oncogene or tumor suppressor gene loss. The present disclosure addresses these needs.

[0004] The present disclosure provides compositions and methods comprising binding partners that specifically bind to target sites on proteins or peptides containing covalently attached molecules. This is believed to be the first disclosure of binding partners with such binding functionality. The present disclosure illustrates this approach using binding partners in the form of numerous antibodies and antibody derivatives that specifically bind to proteins and peptides covalently modified by the attachment of molecules, exemplified by various drugs. Furthermore, the present disclosure demonstrates that binding partners that specifically bind to peptides covalently modified by the attachment of small molecule drugs are specific for the covalently modified peptides presented in the context of human leukocyte antigens (HLA; HLA is a representative example of the major histocompatibility complex [MHC]). Thus, binding partners specific for peptide-drug conjugates in HLA complexes are demonstrated. The present disclosure includes polynucleotides encoding the described binding partners and cells engineered to express the binding partners. The present disclosure includes diagnostic, prophylactic, and therapeutic approaches using the binding partners.

[0005] In one aspect, the present disclosure provides a binding partner that specifically binds to a peptide conjugate / MHC complex, wherein the peptide conjugate / MHC complex comprises: (a) a peptide conjugate formed by the covalent reaction of a targeted covalent inhibitor, or a fragment thereof, with a peptide; and (b) an MHC.

[0006] In some embodiments, the binding partner binds to the peptide conjugate / MHC complex with higher affinity than it binds to the peptide or free target covalent inhibitor, ie, the affinity of the binding partner for the peptide conjugate / MHC complex is 100-10,000 times greater than the affinity of the binding partner for the peptide or free target covalent inhibitor.

[0007] In some embodiments, the MHC is a human leukocyte antigen (HLA), and optionally the HLA is HLA-A, HLA-B, or HLA-C. In some embodiments, the HLA molecule is an HLA-A*02:01, HLA-A*03:01, HLA-A*01:01, HLA-A*11:01, HLA-A*24:02, HLA-A*26:01, HLA-B*07:02, HLA-B*08:01, HLA-B*27:05, HLA-B*39:01, HLA-B*40:01, HLA-B*58:01, and / or HLA-B*15:01 molecule.

[0008] In some embodiments, the peptide comprises a nucleophilic or electrophilic residue, which is optionally one of cysteine, lysine, tyrosine, histidine, serine, arginine, or threonine. In some embodiments, the peptide comprises a cysteine residue. In some embodiments, the peptide conjugate is formed by the covalent reaction of a targeted covalent inhibitor with a cysteine residue in the peptide. In some embodiments, the peptide is a segment of a protein associated with cancer, optionally wherein the protein is encoded by a gene that is mutated in the cancer. In some embodiments, the peptide is a segment of an enzyme, wherein the targeted covalent inhibitor is an inhibitor of the enzyme. In some embodiments, the enzyme is a kinase or a GTPase.

[0009] In some embodiments, the peptide is or is derived from: RAS (e.g., KRAS, HRAS, or NRAS), Bruton's tyrosine kinase (BTK), any epidermal growth factor receptor (EGFR) family member selected from EGFR (ERBB1), HER2 / NEU (ERBB2), HER3 (ERBB3), and HER4 (ERBB4); MET (HGFR); any fibroblast growth factor receptor (FGFR); any cyclin-dependent kinase (CDK); acetylcholinesterase (ACHE); p90 ribosomal S6 kinase (RSK); a cathepsin selected from TP53, IDH1, GNAS, FBXW7, CTNNB1, DNMT3A, cathepsin B, C, F, H, K, L, O, S, V, W, and X; any caspase; a protein involved in obesity, optionally being pancreatic lipase or METAP2; any cancer-testis antigen (CTA); long interspersed nuclear element-1 (LINE-1) element-1); a short interspersed repeat element, which is optionally Alu; and any endogenous retroviral protein, optionally wherein said RAS is KRAS, HRAS, or NRAS.

[0010] In some embodiments, the peptide is a KRAS G12C , KRAS G12D , KRAS G12R , KRAS G12S , HRAS G12C , HRAS G12D , HRAS G12R , HRAS G12S NRAS G12C , NRAS G12D , NRAS G12R , or NRAS G12S Includes segments of. In some embodiments, the peptide comprises the amino acid sequence of VVVGACGVGK, VVGACGVGK, or KLVVVGACGV.

[0011] In some embodiments, the targeted covalent inhibitor is (i) a tri-complex KRAS G12C inhibitors or KRAS G12C degraders, (ii) tricomplex KRAS G12D inhibitors or KRAS G12D degraders, (iii) tricomplex KRAS G12R inhibitors or KRAS G12R degraders, or (iv) tricomplex KRAS G12S inhibitors or KRAS G12S It is a decomposing agent.

[0012] In some embodiments, the peptide is a KRAS G12C Targeted covalent inhibitors contain KRAS mutations G12C In some embodiments, the peptide is a KRAS inhibitor. G12D Targeted covalent inhibitors contain KRAS mutations G12D In some embodiments, the peptide is a KRAS inhibitor. G12R Targeted covalent inhibitors contain KRAS mutations G12R In some embodiments, the peptide is a KRAS inhibitor. G12S Targeted covalent inhibitors contain KRAS mutations G12S It is an inhibitor.

[0013] In some embodiments, the targeted covalent inhibitor is osimertinib, ibrutinib, neratinib, sotorasib, ARS-853, ARS-1620, ARS-3248, MRTX849, JNJ74699157, LY3499446, LY3537982, MRTX-1257, JDQ443, MRTX-1133, RMC-6291, RMC-9805, GDC-6036, D-1553, 2E07, 6H05, SML-8-73-1 or BI 182391.

[0014] In some embodiments, the peptide comprises a segment of EGFR. In some embodiments, the peptide comprises the amino acid sequence of QLMPFGCLL, LMPFGCLLDY or MPFGCLLDY. In some embodiments, the peptide comprises a segment of EGFR and the targeted covalent inhibitor is an inhibitor of an EGFR family kinase. In some embodiments, the targeted covalent inhibitor is osimertinib or neratinib. In some embodiments, the peptide comprises a segment of BTK. In some embodiments, the peptide comprises the amino acid sequence YMANGCLLNY. In some embodiments, the targeted covalent inhibitor is ibrutinib.

[0015] In some embodiments, the peptide is a full-length protein that is processed intracellularly after covalent reaction with a target covalent inhibitor to produce smaller peptide fragments.

[0016] In some embodiments, the peptide conjugate comprises a compound selected from the group consisting of compounds 1-8: [ka] [ka] These are covalently linked to cysteine residues of peptides containing the amino acid sequence VVVGACGVGK, VVGACGVGK or KLVVVGACGV.

[0017] In some embodiments, the MHC is HLA-A*02:01, HLA-A*03:01 and / or HLA-A*11:01. In some embodiments, the peptide comprises the amino acid sequence of VVVGACGVGK or VVGACGVGK, and the MHC is HLA-A*03:01 or HLA-A*11:01. In some embodiments, the peptide comprises the amino acid sequence of KLVVVGACGV, and the MHC is HLA-A*02:01.

[0018] In some embodiments, the peptide conjugate comprises compound 9: [ka] It is covalently attached to a cysteine residue of a peptide containing the amino acid sequence QLMPFGCLL, LMPFGCLLDY or MPFGCLLDY.

[0019] In some embodiments, the MHC is HLA-A*02, HLA-A*01, HLA-A*03 or HLA-A*26.

[0020] In some embodiments, the peptide conjugate comprises compound 10: [ka] It is covalently attached to a cysteine residue of a peptide containing the amino acid sequence YMANGCLLNY.

[0021] In some embodiments, the MHC is HLA-A*01:01.

[0022] In some embodiments, the binding partner specifically binds to a peptide conjugate / MHC complex, wherein the peptide conjugate is a combination of sotorasib and KRAS G12CThe binding partner is formed by a covalent bond reaction with a peptide, wherein the binding partner comprises a heavy chain variable region (VH) and a light chain variable region (VL). In some embodiments, the VH comprises: (i) a CDR-H1 comprising the amino acid sequence of DYSIH, or a variant thereof comprising one to three amino acid changes; (ii) a CDR-H2 comprising the amino acid sequence of SISSSSGSTSYADSVKG, or a variant thereof comprising one to five amino acid changes; and / or (iii) a CDR-H3 comprising the amino acid sequence of GX1WX2X3AMDY, where X1 is G, R, H, S, or K, X2 is Y or I, and X3 is P or A. In some embodiments, the VL comprises: (i) a CDR-L1 comprising the amino acid sequence of RASQSVSSAVA, or a variant thereof comprising 1 to 5 amino acid changes; (ii) a CDR-L2 comprising the amino acid sequence of SASSLYS, or a variant thereof comprising 1 to 5 amino acid changes; and / or (iii) a CDR-L3 comprising the amino acid sequence of QQX1SYVX2X3X4IT, where X1 is I, A, P, V, or S, X2 is K, R, A, or H, X3 is K or R, and X4 is L, T, K, R, V, A, or E.

[0023] In some embodiments, the binding partner specifically binds to a peptide conjugate / MHC complex, wherein the peptide conjugate is a combination of sotorasib and KRAS G12C The binding partner is formed by a covalent reaction with a peptide, wherein the binding partner comprises the CDR-H1, CDR-H2, and CDR-H3 amino acid sequences of the VH amino acid sequence and / or the CDR-L1, CDR-L2, and CDR-L3 amino acid sequences of the VL amino acid sequence of the binding partner, and the binding partner is selected from the group consisting of RA_D11, RA_D01-RA_D04, RA_D06-RA_D09, RA_D12-RA_D14, RA_D16, RA_D18-RA_D21, RA_D23, and RA_D24; or a variant thereof comprising 1 to 5 amino acid changes in one or more of the CDR amino acid sequences.

[0024] In some embodiments, the binding partner comprises the CDR-H1, CDR-H2, CDR-H3, CDR-L1, CDR-L2, and / or CDR-L3 amino acid sequence of a binding partner selected from the group consisting of RA_D11, RA_D01-RA_D04, RA_D06-RA_D09, RA_D12-RA_D14, RA_D16, RA_D18-RA_D21, RA_D23, and RA_D24 (Tables G and H), or a variant thereof comprising 1 to 5 amino acid changes in one or more of the CDR amino acid sequences.

[0025] In some embodiments, the binding partner comprises a VH and / or VL amino acid sequence that is 90%, 95%, or 100% identical to the following VH and VL sequences: TIFF2025526240000005.tif110165TIFF2025526240000006.tif243168TIFF2025526240000007.tif162164

[0026] In some embodiments, the binding partner specifically binds to a peptide conjugate / MHC complex, wherein the peptide conjugate is formed by a covalent reaction between osimertinib and an EGFR peptide, and wherein the binding partner comprises a heavy chain variable region (VH) and a light chain variable region (VL). In some embodiments, the VH comprises: (i) a CDR-H1 comprising the amino acid sequence of SSYIH, or a variant thereof comprising one to five amino acid changes; (ii) a CDR-H2 comprising the amino acid sequence of YISPSYGSTSYADSVKG, or a variant thereof comprising one to five amino acid changes; and / or (iii) a CDR-H3 comprising the amino acid sequence of EX1X2X3MX4X5DY, wherein X1 is Y, L, S, or E, X2 is V, T, or I, X3 is T or I, X4 is A, T, or S, and X5 is L, A, I, K, P, or T. In some embodiments, the VL comprises: (i) a CDR-L1 comprising the amino acid sequence of RASQSVSSAVA, or a variant thereof comprising 1 to 5 amino acid changes; (ii) a CDR-L2 comprising the amino acid sequence of SASSLYS, or a variant thereof comprising 1 to 5 amino acid changes; and / or (iii) a CDR-L3 comprising the amino acid sequence of QQYX1X2WPX3T, where X1 is S or A or S; X2 is Y, H, A, D, E, K, S or G; and X3 is I or E.

[0027] In some embodiments, the binding partner specifically binds to the peptide conjugate / MHC complex, and the peptide conjugate is formed by a covalent reaction of osimertinib with an EGFR peptide, wherein the binding partner comprises the CDR-H1, CDR-H2, and CDR-H3 amino acid sequences of the VH amino acid sequence, and / or the CDR-L1, CDR-L2, and CDR-L3 amino acid sequences of the VL amino acid sequence of the binding partner selected from the group consisting of OEA2-5, EO_Q01-EO_Q18, and EO_Q20-EO_Q24, or variants thereof comprising 1 to 5 amino acid changes in one or more of the CDR amino acid sequences.

[0028] In some embodiments, the binding partner comprises the CDR-H1, CDR-H2, CDR-H3, CDR-L1, CDR-L2 and / or CDR-L3 amino acid sequence of a binding partner selected from the group consisting of OEA2-5, EO_Q01-EO_Q18 and EO_Q20-EO_Q24 (Tables I and J), or a variant thereof comprising 1 to 5 amino acid changes in one or more of the CDR amino acid sequences.

[0029] In some embodiments, the binding partner comprises a VH and / or VL amino acid sequence that is 90%, 95%, or 100% identical to the following VH and VL sequences: TIFF2025526240000008.tif107163TIFF2025526240000009.tif242163TIFF2025526240000010.tif243163

[0030] In some embodiments, the binding partner has a higher affinity for a peptide conjugate / MHC complex comprising a first HLA than for a peptide conjugate / MHC complex comprising a second HLA.

[0031] In some embodiments, the first and second HLA molecules are each selected from the group consisting of HLA-A*02:01, HLA-A*03:01, HLA-A*01:01, HLA-A*11:01, HLA-A*24:02, HLA-A*26:01, HLA-B*07:02, HLA-B*08:01, HLA-B*27:05, HLA-B*39:01, HLA-B*40:01, HLA-B*58:01 and / or HLA-B*15:01.

[0032] In some embodiments, the binding partner is an intact antibody, a bispecific antibody, a multispecific antibody, an antigen-binding (Fab) fragment, a Fab' fragment, a (Fab')2 fragment, an Fd, an Fv, a dAb, a single domain fragment or a single monomeric variable antibody domain, a Dual-Affinity Retargeting (DART) molecule, a diabody (Db), a single-chain diabody (scDb), a single-chain variable fragment (scFv), a bispecific T-cell engager (BiTE), a bispecific killer cell engager (BiKE), a CrossMab, a camelid antibody, a trispecific binding partner, a chimeric antigen receptor (CAR), a monobody (also known as an adnectin), a DARPin, an anticalin, an affibody, or an affimer. In some embodiments, the binding partner is bispecific. In some embodiments, the binding partner specifically binds to the peptide conjugate / MHC complex and the T cell antigen. In some embodiments, the binding partner specifically binds to the peptide conjugate / MHC complex and human CD3.

[0033] In some embodiments, the binding partner comprises the following VH and / or VL amino acid sequences: TIFF2025526240000011.tif27166

[0034] In some embodiments, the binding partner comprises a sequence that is at least 90% similar to any of the following sequences (excluding the underlined sequences): TIFF2025526240000012.tif127162

[0035] In various embodiments, the binding partners described herein do not comprise the amino acid sequence LEGGGGLNDIFEAQKIEWHESRHHHHHH. In various embodiments, the binding partners described herein do not comprise an AviTag or a HisTag. In various embodiments, the binding partner administered to a subject does not comprise the amino acid sequence LEGGGGLNDIFEAQKIEWHESRHHHHHH. In various embodiments, the binding partner administered to a subject does not comprise an AviTag or a HisTag.

[0036] In some embodiments, the binding partner is a single-chain diabody (scDb) and comprises a sequence that is at least 90% identical to any of the following sequences (excluding the underlined sequences): TIFF2025526240000013.tif130164

[0037] In some embodiments, the binding partner comprises a heavy chain constant region selected from the group consisting of human IgM, IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2. In some embodiments, the heavy chain constant region comprises one or more amino acid substitutions in the Fc region. In some embodiments, the binding partner comprises a human kappa light chain constant region or a human lambda light chain constant region. In some embodiments, the binding partner is in a CrossMab format. In some embodiments, the binding partner is conjugated to a detectable label, a chemotherapeutic agent, a radioisotope, or a toxin. In some embodiments, the binding partner is comprised within a chimeric antigen receptor. In some embodiments, the binding partner is expressed by a T cell, a macrophage, a neutrophil, or a natural killer cell. In some embodiments, binding of the binding partner to the peptide conjugate / MHC complex is not inhibited by a free target covalent inhibitor.

[0038] In another aspect, the present disclosure provides a complex comprising a binding partner disclosed herein and a peptide conjugate / MHC complex. In another aspect, the present disclosure provides polynucleotides encoding the binding partners disclosed herein. In another aspect, the present disclosure provides polynucleotides encoding the heavy chain variable region and / or the light chain variable region of a binding partner disclosed herein. In another aspect, the present disclosure provides a vector comprising a polynucleotide disclosed herein. In some embodiments, the vector is a viral vector, hi some embodiments, the viral vector is an adenoviral vector, a lentiviral vector, a retroviral vector, or an adeno-associated viral vector.

[0039] In another aspect, the present disclosure provides a recombinant host cell comprising: (a) a polynucleotide disclosed herein; (b) a vector disclosed herein; (c) a first polynucleotide encoding the VH or heavy chain of a binding partner disclosed herein, and a second polynucleotide encoding the VL or light chain of a binding partner disclosed herein; or (d) a first vector comprising a first polynucleotide encoding the VH or heavy chain of a binding partner disclosed herein, and a second vector comprising a second polynucleotide encoding the VL or light chain of a binding partner disclosed herein.

[0040] In another aspect, the present disclosure provides a pharmaceutical composition comprising a binding partner disclosed herein, a polynucleotide disclosed herein, a vector disclosed herein, or a host cell disclosed herein, and a pharmaceutically acceptable carrier or excipient.

[0041] In another aspect, the disclosure provides a method of producing a binding partner, the method comprising culturing a host cell disclosed herein under appropriate conditions such that the polynucleotide is expressed and the binding partner is produced.

[0042] In another aspect, the disclosure provides a eukaryotic cell comprising a polynucleotide disclosed herein or a vector disclosed herein. In some embodiments, the cell is optionally a totipotent stem cell, a multipotent stem cell, or a pluripotent stem cell. In some embodiments, the stem cell has an induced stem cell phenotype, or the cell is optionally a leukocyte, optionally a CD4+ T cell, optionally a CD8+ T cell, optionally a γδ T cell, optionally a natural killer cell, a natural killer T cell, a mucosal-associated invariant T (MAIT) cell, a neutrophil, or a macrophage.

[0043] In another aspect, the present disclosure provides a method comprising administering a binding partner disclosed herein, a polynucleotide disclosed herein, a vector disclosed herein, a pharmaceutical composition disclosed herein, or a cell disclosed herein to an individual in need thereof.

[0044] In another aspect, the disclosure provides a method of producing a peptide conjugate / MHC complex, the method comprising contacting a cell with a targeted covalent inhibitor and isolating the peptide conjugate / MHC complex. In some embodiments, the method comprises identifying the peptide conjugate / MHC complex.

[0045] In another aspect, the present disclosure provides a cell-free peptide conjugate / MHC complex comprising: (a) an isolated peptide conjugate formed by the covalent reaction of a targeted covalent inhibitor with a peptide; and (b) MHC.

[0046] In some embodiments, the peptide comprises a segment of: RAS (e.g., KRAS G12C , KRAS G12D , KRAS G12R , KRAS G12S , HRAS G12C , HRAS G12D , HRAS G12R , HRAS G12S NRAS G12C , NRAS G12D , NRAS G12R or NRAS G12S ), EGFR, BTK, HER2 / NEU (ERBB2), HER3 (ERBB3), HER4 (ERBB4), MET (HGFR); FGFR, CDK, acetylcholinesterase (ACHE), p90 ribosomal S6 kinase (RSK), TP53, IDH1, GNAS, FBXW7, CTNNB1, DNMT3A, cathepsin B, cathepsin C, cathepsin F, cathepsin H, cathepsin K, cathepsin L, cathepsin O, cathepsin S, cathepsin V, cathepsin W, cathepsin X, caspase, pancreatic lipase, METAP2, any cancer-testis antigen (CTA), long interspersed nucleotide sequence-1 (LINE-1), short interspersed nucleotide sequence (optionally Alu), or any endogenous retroviral protein.

[0047] In some embodiments, the targeted covalent inhibitor is osimertinib, ibrutinib, neratinib, sotorasib, ARS-853, ARS-1620, ARS-3248, MRTX849, JNJ74699157, LY3499446, LY3537982, MRTX-1257, JDQ443, MRTX-1133, RMC-6291, RMC-9805, GDC-6036, D-1553, 2E07, 6H05, SML-8-73-1 or BI 182391. In another aspect, the present disclosure provides a cell-free peptide conjugate / MHC complex comprising: (a) a peptide conjugate / MHC complex; (a) a compound selected from the group consisting of compounds 1 and 4 to 8: [ka] [ka] which are covalently linked to a cysteine residue of a peptide containing the amino acid sequence VVVGACGVGK, VVGACGVGK, or KLVVVGACGV; and (b) MHC.

[0048] In another aspect, the present disclosure provides a cell-free peptide conjugate / MHC complex comprising: (a) a peptide conjugate / MHC complex; (a) Compound 2 [ka] which is covalently attached to a cysteine residue in a peptide containing the amino acid sequence QLMPFGCLL, LMPFGCLLDY, or MFPGCLLDY; and (b) MHC.

[0049] In another aspect, the present disclosure provides a cell-free peptide conjugate / MHC complex comprising: (a) a peptide conjugate / MHC complex; (a) Compound 3 [ka] which is covalently attached to a cysteine residue of a peptide containing the amino acid sequence YMANGCLLNY; and (b) MHC.

[0050] In some embodiments, the MHC is HLA, optionally wherein the HLA is an HLA-A*02:01, HLA-A*03:01, HLA-A*01:01, HLA-A*11:01, HLA-A*24:02, HLA-A*26:01, HLA-B*07:02, HLA-B*08:01, HLA-B*27:05, HLA-B*39:01, HLA-B*40:01, HLA-B*58:01, or HLA-B*15:01 molecule.

[0051] In another aspect, the present disclosure provides a recombinant cell or particle comprising on its outer surface a cell-free peptide conjugate / MHC complex disclosed herein.

[0052] In another aspect, the present disclosure provides a method for identifying one or more binding partners that specifically bind to an HLA-presented peptide conjugate, the method comprising contacting a cell-free peptide conjugate / MHC complex disclosed herein with multiple binding partners and selecting one or more binding partners that specifically bind to the cell-free peptide conjugate / MHC complex. In some embodiments, the method further comprises determining the sequence of the one or more selected binding partners. In some embodiments, the method further comprises producing the one or more selected binding partners.

[0053] In another aspect, the present disclosure provides a method for identifying binding partners that specifically bind to a peptide conjugate presented by two or more HLAs, the method comprising providing a sample of cells from a subject treated with a targeted covalent inhibitor and having different HLA types, or providing a cell-free peptide conjugate / MHC complex disclosed herein that is presented by two or more HLAs, and screening for binding partners to thereby identify binding partners that specifically bind to the peptide conjugate or antigen presented by the two or more HLAs. In some embodiments, the two or more HLA types include HLA-A*02:01 and at least one additional HLA type. In some embodiments, each of the two or more HLAs is an HLA-A*02:01, HLA-A*03:01, HLA-A*01:01, HLA-A*11:01, HLA-A*24:02, HLA-A*26:01, HLA-B*07:02, HLA-B*08:01, HLA-B*27:05, HLA-B*39:01, HLA-B*40:01, HLA-B*58:01, or HLA-B*15:01 molecule.

[0054] In another aspect, the present disclosure provides a method of killing cancer cells in a subject, the method comprising administering to the subject: (a) a targeted covalent inhibitor, and (b) a binding partner disclosed herein, a polynucleotide disclosed herein, a vector disclosed herein, a pharmaceutical composition disclosed herein, or a cell disclosed herein.

[0055] In some embodiments, the targeted covalent inhibitor targets: RAS (e.g., KRAS, HRAS, NRAS), Bruton's tyrosine kinase (BTK), EGFR (ERBB1), HER2 / NEU (ERBB2), HER3 (ERBB3), HER4 (ERBB4), fibroblast growth factor receptor (FGFR), MET, BRAF, cyclin-dependent kinase (CDK), acetylcholinesterase (ACHE), TP53, IDH1, GNAS, FBXW7, CTNNB1, DNMT3A, cathepsin, caspase, pancreatic lipase, METAP2, any cancer-testis antigen, endogenous retroviral protein, long interspersed nucleotide sequence-1 (LINE-1), or short interspersed nucleotide sequence (SINE).

[0056] In another embodiment, the present disclosure provides a method for treating EGFR, BTK, or RAS (e.g., KRAS G12C , KRAS G12D , KRAS G12R , KRAS G12S , HRAS G12C , HRAS G12D , HRAS G12R , HRAS G12S NRAS G12C , NRAS G12D , NRAS G12R or NRAS G12S In subjects treated with a covalent inhibitor targeting EGFR, BTK, or RAS (e.g., KRAS G12C , KRAS G12D , KRAS G12R , KRAS G12S , HRAS G12C , HRAS G12D , HRAS G12R , HRAS G12S NRAS G12C , NRAS G12D , NRAS G12R or NRAS G12S), the method comprising administering to a subject a binding partner disclosed herein, a polynucleotide disclosed herein, a vector disclosed herein, a pharmaceutical composition disclosed herein, or a cell disclosed herein.

[0057] In some embodiments, the subject has cancer. In another aspect, the present disclosure provides a method of treating cancer in a subject treated with a targeted covalent inhibitor, the method comprising administering to the subject a binding partner disclosed herein, a polynucleotide disclosed herein, a vector disclosed herein, a pharmaceutical composition disclosed herein, or a cell disclosed herein.

[0058] In another aspect, the present disclosure provides a method of treating a disease or disorder in a subject treated with a targeted covalent inhibitor, the method comprising administering to the subject a binding partner disclosed herein, a polynucleotide disclosed herein, a vector disclosed herein, a pharmaceutical composition disclosed herein, or a cell disclosed herein. In some embodiments, the disease or disorder is an autoimmune disease or a fibrotic disease.

[0059] In another aspect, the present disclosure provides a method for detecting RAS (e.g., KRAS) mutations in a subject with a cancer that exhibits a RAS or EGFR mutation. G12C , KRAS G12D , KRAS G12R , KRAS G12S , HRAS G12C , HRAS G12D , HRAS G12R , HRAS G12S , NRAS G12C , NRAS G12D , NRAS G12R or NRAS G12S ) or EGFR mutations, the method comprising administering to the subject: (a) a RAS (e.g., KRAS G12C, KRAS G12D , KRAS G12R , KRAS G12S , HRAS G12C , HRAS G12D , HRAS G12R , HRAS G12S , NRAS G12C , NRAS G12D , NRAS G12R or NRAS G12S ) or an EGFR inhibitor, and a binding partner disclosed herein, a polynucleotide disclosed herein, a vector disclosed herein, a pharmaceutical composition disclosed herein, or a cell disclosed herein. In some embodiments, the subject has previously been diagnosed with a RAS (e.g., KRAS G12C , KRAS G12D , KRAS G12R , KRAS G12S , HRAS G12C , HRAS G12D , HRAS G12R , HRAS G12S , NRAS G12C , NRAS G12D , NRAS G12R or NRAS G12S ) or being treated with an EGFR inhibitor.

[0060] In some embodiments, the inhibitor is osimertinib, ibrutinib, neratinib, sotorasib, ARS-853, ARS-1620, ARS-3248, MRTX849, JNJ74699157, LY3499446, LY3537982, MRTX-1257, JDQ443, MRTX-1133, RMC-6291, RMC-9805, GDC-6036, D-1553, 2E07, 6H05, SML-8-73-1 or BI 182391. In some embodiments, the targeted covalent inhibitor targets RAS (e.g., KRAS, HRAS, or NRAS), Bruton's tyrosine kinase (BTK), EGFR (ERBB1), HER2 / NEU (ERBB2), HER3 (ERBB3), HER4 (ERBB4), fibroblast growth factor receptor (FGFR), MET, BRAF, cyclin-dependent kinase (CDK), acetylcholinesterase (ACHE), TP53, IDH1, GNAS, FBXW7, CTNNB1, DNMT3A, cathepsin, caspase, pancreatic lipase, METAP2, any cancer-testis antigen, endogenous retroviral protein, long interspersed nucleotide sequence-1 (LINE-1), or short interspersed nucleotide sequence (SINE).

[0061] In some embodiments, the cancer is renal cell carcinoma, breast cancer, prostate cancer, pancreatic cancer, lung cancer, liver cancer, ovarian cancer, cervical cancer, colon or colorectal cancer, esophageal cancer, glioma, glioblastoma, brain cancer (brain tumor), gastric cancer, bladder cancer, testicular cancer (testicular cancer), head and neck cancer, melanoma, skin cancer, sarcoma, fibrosarcoma, angiosarcoma, osteosarcoma, rhabdomyosarcoma, leukemia, lymphoma, or myeloma.

[0062] In some embodiments, the targeted covalent inhibitor is osimertinib, ibrutinib, neratinib, sotorasib, ARS-853, ARS-1620, ARS-3248, MRTX849, JNJ74699157, LY3499446, LY3537982, MRTX-1257, JDQ443, MRTX-1133, RMC-6291, RMC-9805, GDC-6036, D-1553, 2E07, 6H05, SML-8-73-1 or BI 182391.

[0063] In some embodiments, the methods disclosed herein further comprise administering an additional therapeutic agent, hi some embodiments, the additional therapeutic agent is a chemotherapy, an immunomodulatory agent, or another targeted covalent inhibitor. In some embodiments, the immunomodulatory agent is a checkpoint targeting agent, and optionally the checkpoint targeting agent is selected from the group consisting of: an antagonistic anti-PD-1 antibody, an antagonistic anti-PD-L1 antibody, an antagonistic anti-PD-L2 antibody, an antagonistic anti-CTLA-4 antibody, an antagonistic anti-BTLA antibody, an antagonistic anti-TREMR antibody, an antagonistic anti-TIGIT antibody, an antagonistic anti-VISTA antibody, an antagonistic anti-TIM-3 antibody, an antagonistic anti-LAG-3 antibody, an antagonistic anti-CEACAM1 antibody, an agonist anti-GITR antibody, an agonist anti-OX40 antibody and an agonist anti-CD137 antibody, an agonist anti-DR3 antibody, an agonist anti-TNFSF14 antibody, an agonist anti-CD27 antibody, an agonist anti-ICOS antibody, and an agonist anti-CD28 antibody; a cytokine (optionally, the cytokine is anchored IL2 or engineered IL2); or an extracellular adenosine (eADO) signaling inhibitor.

[0064] In another aspect, the present disclosure provides a method for detecting a peptide conjugate / MHC complex in a biological sample, the method comprising contacting the sample with a binding partner disclosed herein, wherein the peptide conjugate / MHC complex comprises a peptide conjugate formed by covalent reaction of a targeted covalent inhibitor with the peptide. In some embodiments, the peptide binds to EGFR, BTK, or RAS (e.g., KRAS G12C , KRAS G12D , KRAS G12R , KRAS G12S , HRAS G12C , HRAS G12D , HRAS G12R , HRAS G12S NRAS G12C , NRAS G12D , NRAS G12R or NRAS G12S In some embodiments, the targeted covalent inhibitor is an EGFR, BTK, or RAS (e.g., KRAS) peptide. G12C , KRAS G12D , KRAS G12R , KRAS G12S , HRAS G12C , HRAS G12D , HRAS G12R , HRAS G12S NRAS G12C , NRAS G12D , NRAS G12R or NRAS G12S ) inhibitor. In some embodiments, the targeted covalent inhibitor is osimertinib, ibrutinib, neratinib, sotorasib, ARS-853, ARS-1620, ARS-3248, MRTX849, JNJ74699157, LY3499446, LY3537982, MRTX-1257, JDQ443, MRTX-1133, RMC-6291, RMC-9805, GDC-6036, D-1553, 2E07, 6H05, SML-8-73-1 or BI 182391. In some embodiments, the biological sample is blood or serum.

[0065] In another aspect, the present disclosure provides a method of identifying a cell that contains a peptide conjugate / MHC complex disclosed herein, the method comprising contacting the cell with a binding partner disclosed herein.

[0066] In another aspect, the present disclosure provides a method of generating one or more binding partners that specifically bind to a peptide conjugate, wherein the peptide conjugate comprises a peptide covalently attached to a non-peptide molecule, the method comprising: (a) exposing the peptide conjugate to a plurality of binding partners; and (b) selecting binding partners that specifically bind to the peptide conjugate to provide one or more selected binding partners. In some embodiments, the methods disclosed herein further comprise selecting binding partners that specifically bind to the peptide conjugate but do not detectably bind, or bind with lower affinity, to the peptide or the non-peptide molecule when they are not covalently attached to each other.

[0067] In another aspect, the present disclosure provides a method for generating one or more binding partners that specifically bind to a peptide conjugate presented in the context of an MHC molecule, or a fragment or derivative thereof, wherein the peptide conjugate comprises a peptide covalently attached to a non-peptide molecule, the method comprising: (a) providing a complex of the peptide conjugate and the MHC molecule, or a fragment or derivative thereof; (b) exposing the complex to multiple binding partners; and (c) selecting binding partners that specifically bind to the complex, thereby providing one or more selected binding partners. In some embodiments, the methods disclosed herein further comprise selecting a binding partner that specifically binds to the complex but does not detectably bind, or binds with low affinity, to a complex of the peptide and the MHC molecule, or a fragment thereof, wherein the peptide is not covalently attached to the non-peptide molecule. In some embodiments, the methods disclosed herein further comprise selecting a binding partner that specifically binds to the peptide conjugate presented in the context of two or more different MHC molecules.

[0068] In some embodiments, the MHC molecule is an MHC class I molecule and the peptide is 7-15 amino acids in length, or the MHC molecule is a non-classical MHC class I molecule and the peptide is 7-15 amino acids in length, hi some embodiments, the MHC molecule is an MHC class II molecule and the peptide is 9-30 amino acids in length.

[0069] In some embodiments, the complex is immobilized on a solid support. In some embodiments, the complex is present on the surface of a cell.

[0070] In some embodiments, the method further comprises determining whether the one or more binding partners selected in step (c) can mediate immune cell-mediated killing, or antibody-dependent cellular cytotoxicity (ADCC), antibody-dependent cellular phagocytosis (ADCP), or complement-dependent cytotoxicity (CDC)-mediated killing of the cells, or determining whether the binding partner can be used as an immunotoxin, as an antibody-drug conjugate (ADC), or as a radioconjugate. In some embodiments, the plurality of binding partners is generated by a phage display library or a yeast display library.

[0071] In another aspect, the present disclosure provides a method of killing cancer cells, the method comprising administering to the cells a binding partner that specifically binds to a peptide conjugate, wherein the peptide conjugate (i) comprises a peptide derived from a target protein in the cells, the peptide being covalently linked to (ii) a non-peptide molecule, and wherein the binding partner mediates immune cell-mediated killing, or antibody-drug conjugate (ADC)-mediated, antibody-dependent cellular phagocytosis (ADCP), or complement-dependent cytotoxicity (CDC) immunotoxin killing of the cells, or kills the cancer cells by using the binding partner as an immunotoxin, or by the action of a radioconjugate.

[0072] In some embodiments, the peptide conjugate is presented on the cell in the context of an MHC molecule. In some embodiments, a binding partner specifically binds to a complex comprising the peptide conjugate and the MHC molecule. In some embodiments, a binding partner does not detectably bind to a complex of the peptide and the MHC molecule (wherein the peptide is not covalently bound to the non-peptide molecule).

[0073] In some embodiments, the methods disclosed herein further comprise administering the non-peptide molecule to the cells, wherein the non-peptide molecule forms a covalent bond with the target protein in the cells. In some embodiments, the non-peptide molecule is administered to the cells before administering the binding partner. In some embodiments, the cells are isolated from a subject. In some embodiments, the cells are in a subject and the binding partner is administered to the subject. In some embodiments, the cells are in a subject and the non-peptide molecule and the binding partner are administered to the subject. In some embodiments, the target protein is alternatively spliced or overexpressed in cancer cells but not alternatively spliced or overexpressed by cancer cells. In some embodiments, the target protein is encoded by a gene that is mutated in cancer cells but not mutated in non-cancer cells. In some embodiments, the non-peptide molecule is a covalent inhibitor of the target protein.

[0074] In some embodiments, the binding partner is an intact antibody, a bispecific antibody, a multispecific antibody, an antigen-binding (Fab) fragment, a Fab' fragment, a (Fab')2 fragment, an Fd, an Fv, a dAb, a single domain fragment or a single monomeric variable antibody domain, a single-chain diabody (scDb), a diabody (Db), a Dual-Affinity Retargeting (DART) molecule, a single-chain variable fragment (scFv), a bispecific T cell engager (BiTE), a bispecific killer cell engager (BiKE), a CrossMab, a camelid antibody, a trispecific binding partner, a chimeric antigen receptor (CAR), a monobody (also known as an Adnectin), a DARPin, an anticalin, an affibody, or an affimer. In some embodiments, the CAR is present on a T cell, a natural killer (NK) cell, a neutrophil, or a macrophage. In some embodiments, the binding partner is an antibody-drug conjugate (ADC), a radioconjugate, or a toxin conjugate.

[0075] In some embodiments, the target protein is selected from the following: RAS (e.g., KRAS, HRAS, or NRAS), Bruton's tyrosine kinase (BTK), a member of the epidermal growth factor receptor (EGFR) family, fibroblast growth factor receptor (FGFR), MET, BRAF, a cyclin-dependent kinase (CDK), acetylcholinesterase (ACHE), TP53, IDH1, GNAS, FBXW7, CTNNB1, DNMT3A, a cathepsin, a caspase, pancreatic lipase, METAP2, a cancer-testis antigen, a viral polymerase, a protein required for viral cell entry, a protein encoded by a transposable element (e.g., an endogenous retrovirus), or a mutant thereof. In some embodiments, the target protein is a KRAS protein containing a G12C mutation, and the non-peptide molecule is selected from sotorasib, ARS-853, ARS-1620, ARS-3248, MRTX849, JNJ74699157, LY3499446, LY3537982, MRTX-1257, JDQ443, RMC-6291, DC-6036, D-1553, 2E07, 6H05, SML-8-73-1 or BI 182391, and derivatives thereof. In some embodiments, the target protein is EGFR and the non-peptide molecule is selected from PD168393, PF00299804 (dacomitinib), EKB569 (pelitinib), afatinib, WZ4002, osimertinib (AZD9291), PF-06459988, nazartinib, nacotinib, olmutinib, avitinib, rociletinib, neratinib, pyrotinib, poziotinib, and derivatives thereof. In some embodiments, the target protein is Bruton's tyrosine kinase (BTK) and the non-peptide molecule is selected from ibrutinib, acalabrutinib, zanubrutinib, CHMFL-BTK-11, ONO / GS-405, PRN1008, CC-292, and derivatives thereof.In some embodiments, the target protein is p90 ribosomal S6 kinase (RSK), and the non-peptide molecule is fluoromethyl ketone (FMK), dimethyl fumarate, or a derivative thereof. In some embodiments, the target protein is FGFR, and the non-peptide molecule is selected from FIIN-1, FIIN-2, FIIN-3, BGJ398, AZD4547, PRN1371, FGF401, and a derivative thereof.

[0076] In another aspect, the present disclosure provides a method of treating cancer in a subject in need thereof, the method comprising administering to the subject an effective amount of a binding partner that specifically binds to a peptide conjugate, wherein the peptide conjugate comprises: (i) a peptide derived from a target protein present in cancer cells of the subject, the peptide being covalently linked to (ii) a non-peptide molecule, wherein the binding partner mediates immune cell-mediated killing or antibody-drug conjugate (ADC)-mediated killing of cancer cells, antibody-dependent cellular phagocytosis (ADCP)-mediated killing, or complement-dependent cytotoxicity (CDC)-mediated killing of cancer cells, or by using the binding partner as an immunotoxin or radioconjugate in the subject, thereby killing cancer cells. In some embodiments, the peptide conjugate is presented on the subject's cancer cells in association with an MHC molecule. In some embodiments, the binding partner specifically binds to a complex comprising the peptide conjugate and the MHC molecule. In some embodiments, the binding partner does not detectably bind to a complex of the peptide and the MHC molecule (wherein the peptide is not covalently bound to the non-peptide molecule). In some embodiments, the subject has previously been administered the non-peptide molecule. In some embodiments, the methods disclosed herein further comprise administering to the subject an effective amount of the non-peptide molecule, wherein the non-peptide molecule forms a covalent bond with the target protein in cancer cells of the subject. In some embodiments, the non-peptide molecule is administered to the subject prior to administering the binding partner.

[0077] In another aspect, the present disclosure provides a method for improving the effectiveness of anti-cancer therapy in a subject in need thereof, wherein the anti-cancer therapy comprises administering to the subject a non-peptide molecule that forms a covalent bond with a target protein in cancer cells of the subject; and the method further comprises administering to the subject an effective amount of a binding partner that specifically binds to a peptide conjugate, wherein the peptide conjugate comprises a peptide derived from the target protein covalently bound to the non-peptide molecule, and wherein the binding partner mediates immune cell-mediated or antibody-drug conjugate (ADC)-mediated killing of cancer cells, or antibody-dependent cellular phagocytosis (ADCP)-mediated or complement-dependent cytotoxicity (CDC)-mediated killing of cancer cells, or kills cancer cells by using the binding partner as an immunotoxin or radioconjugate in the subject. In some embodiments, the peptide conjugate is presented on the subject's cancer cells in the context of an MHC molecule. In some embodiments, the binding partner specifically binds to a complex comprising the peptide conjugate and the MHC molecule. In some embodiments, the binding partner does not detectably bind to a complex of the peptide and the MHC molecule (wherein the peptide is not covalently bound to the non-peptide molecule), hi some embodiments, the subject has been administered the non-peptide molecule prior to administering the binding partner.

[0078] In another aspect, the present disclosure provides a kit, said kit comprising: (i) a non-peptide molecule, wherein the non-peptide molecule forms a covalent bond with a target protein in a cell; (ii) a binding partner that specifically binds to a peptide conjugate, wherein the peptide conjugate comprises a peptide derived from the target protein covalently attached to the non-peptide molecule; and and (iii) instructions for use. Includes:

[0079] In another aspect, the present disclosure provides an isolated peptide conjugate comprising a peptide of 7 to 30 amino acids in length comprising an amino acid sequence at least 80% identical to the amino acid sequence VVGACGVGK, wherein the peptide is conjugated to sotorasib or a derivative thereof.

[0080] In another aspect, the present disclosure provides an isolated peptide conjugate comprising a peptide of 7 to 30 amino acids in length comprising an amino acid sequence that is at least 80% identical to the amino acid sequence QLMPFGCLL, wherein the peptide is conjugated to osimertinib or a derivative thereof.

[0081] In another aspect, the present disclosure provides an isolated peptide conjugate comprising a peptide of 7 to 30 amino acids in length comprising an amino acid sequence that is at least 80% identical to the amino acid sequence YMANGCLLNY, wherein the peptide is conjugated to ibrutinib or a derivative thereof.

[0082] In another aspect, the present disclosure provides an isolated molecular complex comprising a peptide conjugate disclosed herein and an MHC molecule, or a fragment or derivative thereof. In another aspect, the present disclosure provides a host cell comprising the molecular complex disclosed herein. In another aspect, the present disclosure provides a solid surface support comprising a molecular complex disclosed herein.

[0083] In another aspect, the present disclosure provides a kit comprising (i) a peptide conjugate disclosed herein, a molecular complex disclosed herein, a cell disclosed herein, a solid surface support disclosed herein, or any combination thereof, and, optionally, (ii) instructions for use.

[0084] In another aspect, the present disclosure provides an isolated binding partner that specifically binds to the peptide conjugate or molecular complex disclosed herein. In some embodiments, the binding partner is an antibody or an antigen-binding fragment thereof. In some embodiments, the binding partner is a component of a chimeric antigen receptor (CAR).

[0085] In another aspect, the present disclosure provides a fusion protein comprising a binding partner disclosed herein, wherein the fusion protein comprises: (i) at least one cytokine, and / or (ii) an additional antibody or fragment thereof that is an immune checkpoint inhibitor. In some embodiments, the cytokine is selected from the group consisting of interleukin (IL)-2, IL-7, IL-8, IL-15, IL-17, IL-18, and combinations thereof, and / or (ii) Additional antibodies or fragments thereof that are immune checkpoint inhibitors are anti-PD-1 antibodies, antagonistic anti-PD-L1 antibodies, antagonistic anti-PD-L2 antibodies, antagonistic anti-CTLA-4 antibodies, antagonistic anti-BTLA antibodies, antagonistic anti-TREMR antibodies, antagonistic anti-TIGIT antibodies, antagonistic anti-VISTA antibodies, antagonistic anti-TIM-3 antibodies, antagonistic anti-LAG-3 antibodies, antagonistic anti-CEACAM1 antibodies, agonistic anti-GITR antibodies, agonistic anti-OX40 antibodies, and agonistic anti-CD137 antibodies, agonistic anti-DR3 antibodies, agonistic anti-TNFSF14 antibodies, agonistic anti-CD27 antibodies, agonistic anti-ICOS antibodies, or agonistic anti-CD28 antibodies. In some embodiments, the binding partner comprises a moiety that binds to a T cell or natural killer (NK) cell protein, wherein the T cell or NK cell protein is selected from the group consisting of a T cell receptor protein, CD4, CD8, CD28, CD16A, NKG2D, NKp30, NKp46, and combinations thereof.

[0086] In another aspect, the disclosure provides a polypeptide comprising an antigen-binding domain comprising a heavy chain variable region (VH) and / or a light chain variable region (VL). In some embodiments, the VH comprises a heavy chain complementarity-determining region 3 (CDR-H3) comprising the amino acid sequence of GX1WX2X3AMDY, where X1 is G, R, H, S, or K, X2 is Y or I, and X3 is P or A. In some embodiments, the VL comprises a light chain complementarity-determining region 3 (CDR-L3) comprising the amino acid sequence of QQX1SYVX2X3X4IT, where X1 is I, A, P, V, or S, X2 is K, R, A, or H, X3 is K or R, and X4 is L, T, K, R, V, A, or E.

[0087] In another aspect, the present disclosure provides a polypeptide comprising an antigen-binding domain, wherein the antigen-binding domain binds to a peptide conjugate / MHC complex, wherein the peptide conjugate of the peptide conjugate / MHC complex is a peptide covalently linked to a targeted covalent inhibitor or fragment thereof; and wherein the polypeptide binds to an epitope of the MHC.

[0088] In another aspect, the present disclosure provides a polypeptide comprising an antigen-binding domain, wherein the antigen-binding domain binds to a peptide conjugate / MHC complex, wherein the peptide conjugate of the peptide conjugate / MHC complex is a peptide covalently attached to a targeted covalent inhibitor or fragment thereof; and wherein the antigen-binding domain has a dissociation constant (K D ) binds to the peptide conjugate / MHC complex.

[0089] In another aspect, the present disclosure provides a polypeptide comprising an antigen-binding domain, wherein the antigen-binding domain binds to a peptide conjugate / MHC complex, wherein the peptide conjugate of the peptide conjugate / MHC complex is a peptide covalently attached to a targeted covalent inhibitor or fragment thereof; and wherein the antigen-binding domain has a dissociation constant (KD ) and binds to the free target covalent inhibitor.

[0090] In another aspect, the present disclosure provides a polypeptide comprising an antigen-binding domain, wherein the antigen-binding domain binds to a peptide conjugate / MHC complex, wherein the peptide conjugate of the peptide conjugate / MHC complex is a peptide covalently bound to a targeted covalent inhibitor or fragment thereof; and wherein: (a) the polypeptide binds to the peptide conjugate / MHC complex at an angle between the axis of the MHC and the axis of the polypeptide of about 10° to 60°; (b) when the polypeptide is bound to the peptide conjugate / MHC complex, the angle between the axis of the polypeptide and the axis of the peptide conjugate / MHC complex is about 10° to 60°; (c) when the polypeptide is bound to the peptide conjugate / MHC complex, the angle between the axis of the polypeptide and the axis of the peptide conjugate / MHC complex is different from the angle between the axis of the TCR and the axis of the peptide conjugate / MHC complex when the TCR is bound to the peptide conjugate / MHC complex; (d) When the polypeptide is bound to the peptide conjugate / MHC complex, the angle between the axis of the polypeptide and the axis of the peptide conjugate / MHC complex is about 10° to 60° less than, or about 10° to 60° more than, the angle between the axis of the TCR and the axis of the peptide conjugate / MHC complex when the TCR is bound to the peptide conjugate / MHC complex; (e) the polypeptide binds to the peptide conjugate / MHC complex at an angle that is different from the angle at which the T cell receptor binds to the peptide conjugate / MHC complex; and / or (f) the polypeptide binds to the peptide conjugate / MHC complex at an angle that is about 10° to 60° less than, or about 10° to 60° more than, the angle at which the T cell receptor binds to the peptide conjugate / MHC complex.

[0091] In another aspect, the present disclosure provides a polypeptide comprising an antigen-binding domain, wherein the antigen-binding domain binds to a peptide conjugate / MHC complex, wherein the peptide conjugate of the peptide conjugate / MHC complex is a peptide covalently attached to a targeted covalent inhibitor or fragment thereof; and wherein said polypeptide contacts one or more residues of the α1 domain or region and one or more residues of the α2 domain or region of the heavy chain of the MHC of the peptide conjugate / MHC complex.

[0092] In another aspect, the present disclosure provides a polypeptide comprising an antigen-binding domain, wherein the antigen-binding domain binds to a peptide conjugate / MHC complex, wherein the peptide conjugate of the peptide conjugate / MHC complex is a peptide covalently linked to a targeted covalent inhibitor or fragment thereof; and wherein said polypeptide binds to residues 62-66, 106-109, and / or 150-170 of MHC, or one or more residues thereof.

[0093] In another aspect, the present disclosure provides a polypeptide comprising an antigen-binding domain, wherein the antigen-binding domain binds to a peptide conjugate / MHC complex, wherein the peptide conjugate of the peptide conjugate / MHC complex is a peptide covalently attached to a targeted covalent inhibitor, or a fragment thereof; and wherein the interface area between the peptide conjugate / MHC complex and the polypeptide is at least about 500 Å. 2In some embodiments, the polypeptides disclosed herein comprise a heavy chain variable region (VH) and / or a light chain variable region (VL). In some embodiments, the VH comprises a CDR-H3 comprising the amino acid sequence of GX1WX2X3AMDY, where X1 is G, R, H, S, or K, X2 is Y or I, and X3 is P or A. In some embodiments, the VL comprises a CDR-L3 comprising the amino acid sequence of QQX1SYVX2X3X4IT, where X1 is I, A, P, V, or S, X2 is K, R, A, or H, X3 is K or R, and X4 is L, T, K, R, V, A, or E. In some embodiments, the VH comprises: (i) a CDR-H1 comprising the amino acid sequence DYSIH, or a variant thereof comprising 1 to 3 amino acid changes; (ii) a CDR-H2 comprising the amino acid sequence SISSSSGSTSYADSVKG, or a variant thereof comprising 1 to 5 amino acid changes; and / or (iii) a CDR-H3 comprising the amino acid sequence GX1WX2X3AMDY, where X1 is G, R, H, S, or K, X2 is Y or I, and X3 is P or A. In some embodiments, the VL comprises: (i) a CDR-L1 comprising the amino acid sequence RASQSVSSAVA, or a variant thereof comprising 1 to 5 amino acid changes; (ii) a CDR-L2 comprising the amino acid sequence SASSLYS, or a variant thereof comprising 1 to 5 amino acid changes; and / or (iii) a CDR-L3 comprising the amino acid sequence QQX1SYVX2X3X4IT, where X1 is I, A, P, V, or S, X2 is K, R, A, or H, X3 is K or R, and X4 is L, T, K, R, V, A, or E.

[0094] In some embodiments, the VH comprises a CDR-H1 having the sequence of DYSIH, a CDR-H2 having the sequence of SISSSSGSTSYADSVKG, and a CDR-H3 having the sequence of GGWIAAMDY. In some embodiments, the VL comprises a CDR-L1 having the sequence of RASQSVSSAVA, a CDR-L2 having the sequence of SASSLYS, and a CDR-L3 having the sequence of QQASYVRKTIT. In some embodiments, the VH comprises an amino acid sequence having at least about 90%, 95%, or 100% sequence identity to the amino acid sequence EVQLVESGGGLVQPGGSLRLSCAASGFTFSDYSIHWVRQAPGKGLEWVASISSSSGSTSYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARGGWIAAMDYWGQGTLVTVSS. In some embodiments, the VL comprises an amino acid sequence having at least about 90%, 95%, or 100% sequence identity to the amino acid sequence DIQMTQSPSSLSASVGDRVTITCRASQSVSSAVAWYQQKPGKAPKLLIYSASSLYSGVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQASYVRKTITFGQGTKVEIKRTV.

[0095] In some embodiments, the antigen-binding domain comprises a heavy chain variable region (VH), wherein the VH comprises a heavy chain complementarity-determining region 3 (CDR-H3) comprising the amino acid sequence GSWIHAMDY. In some embodiments, the VH comprises a CDR-H2 comprising the amino acid sequence SISSSWGVTSYADSVKG. In some embodiments, the VH comprises a CDR-H1 comprising the amino acid sequence FHWYSIH. In some embodiments, the antigen-binding domain further comprises a light chain variable region (VL), wherein the VL comprises a CDR-L3 comprising the amino acid sequence QQASYVRKTIT. In some embodiments, the VL comprises a CDR-L2 comprising the amino acid sequence SASSLYS. In some embodiments, the VL comprises a CDR-L1 comprising the amino acid sequence RASQSVSSAVA. In some embodiments, the antigen-binding domain comprises the following: a CDR-H3 sequence of GSWIHAMDY, a CDR-H2 sequence of SISSSWGVTSYADSVKG, a CDR-H1 sequence of FHWYSIH, a CDR-L3 sequence of QQASYVRKTIT, a CDR-L2 sequence of SASSLYS, and a CDR-L1 sequence of RASQSVSSAVA. In some embodiments, the VH comprises a sequence having at least 80% sequence identity to the sequence EVQLVESGGGLVQPGGSLRLSCAASGFTFHWYSIHWVRQAPGKGLEWVASISSSWGVTSYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARGSWIHAMDYWGQGTLVTVSS. In some embodiments, the VL comprises a sequence having at least 80% sequence identity to the sequence DIQMTQSPSSLSASVGDRVTITCRASQSVSSAVAWYQQKPGKAPKLLIYSASSLYSGVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQASYVRKTITFGQGTKVEIKRTV. In some embodiments, the antigen-binding domain comprises a heavy chain variable region (VH), wherein the VH comprises a heavy chain complementarity-determining region 3 (CDR-H3) comprising the amino acid sequence GHWIAAMDY. In some embodiments, the VH comprises a CDR-H2 comprising the amino acid sequence SIASSSGSTGYADSVKG.In some embodiments, the VH comprises a CDR-H1 comprising the amino acid sequence FSWYSIH. In some embodiments, the antigen-binding domain further comprises a light chain variable region (VL), wherein the VL comprises a light chain complementarity-determining region 3 (CDR-L3) comprising the amino acid sequence QQASYVRKTIT. In some embodiments, the VL comprises a CDR-L2 comprising the amino acid sequence SASSLYS. In some embodiments, the VL comprises a CDR-L1 comprising the amino acid sequence RASQSVSSAVA. In some embodiments, the antigen-binding domain comprises: a CDR-H3 sequence of GHWIAAMDY, a CDR-H2 sequence of SIASSSGSTGYADSVKG, a CDR-H1 sequence of FSWYSIH, a CDR-L3 sequence of QQASYVRKTIT, a CDR-L2 sequence of SASSLYS, and a CDR-L1 sequence of RASQSVSSAVA. In some embodiments, the VH comprises a sequence having at least 80% sequence identity to the sequence EVQLVESGGGLVQPGGSLRLSCAASGFTFSWYSIHWVRQAPGKGLEWVASIASSSGSTGYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARGHWIAAMDYWGQGTLVTVSS. In some embodiments, the VL comprises a sequence having at least 80% sequence identity to the sequence DIQMTQSPSSLSASVGDRVTITCRASQSVSSAVAWYQQKPGKAPKLLIYSASSLYSGVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQASYVRKTITFGQGTKVEIKRTV.

[0096] In some embodiments, the antigen-binding domain comprises a heavy chain variable region (VH), wherein the VH comprises a heavy chain complementarity-determining region 3 (CDR-H3) comprising the amino acid sequence GGVIHAMDY. In some embodiments, the VH comprises a CDR-H2 comprising the amino acid sequence SILSRWGVTSYADSVKG. In some embodiments, the VH comprises a CDR-H1 comprising the amino acid sequence FSPYSIH. In some embodiments, the antigen-binding domain further comprises a light chain variable region (VL), wherein the VL comprises a CDR-L3 comprising the amino acid sequence QQASYVRKTIT. In some embodiments, the VL comprises a CDR-L2 comprising the amino acid sequence SASSLYS. In some embodiments, the VL comprises a CDR-L1 comprising the amino acid sequence RASQSVSSAVA. In some embodiments, the antigen-binding domain comprises the following: a CDR-H3 sequence of GGVIHAMDY, a CDR-H2 sequence of SILSRWGVTSYADSVKG, a CDR-H1 sequence of FSPYSIH, a CDR-L3 sequence of QQASYVRKTIT, a CDR-L2 sequence of SASSLYS, and a CDR-L1 sequence of RASQSVSSAVA. In some embodiments, the VH comprises a sequence having at least 80% sequence identity to the sequence EVQLVESGGGLVQPGGSLRLSCAASGFTFSPYSIHWVRQAPGKGLEWVASILSRWGVTSYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARGGVIHAMDYWGQGTLVTVSS. In some embodiments, the VL comprises a sequence having at least 80% sequence identity to the sequence DIQMTQSPSSLSASVGDRVTITCRASQSVSSAVAWYQQKPGKAPKLLIYSASSLYSGVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQASYVRKTITFGQGTKVEIKRTV.

[0097] In some embodiments, the antigen-binding domain comprises a heavy chain variable region (VH), wherein the VH comprises a heavy chain complementarity-determining region 3 (CDR-H3) comprising the amino acid sequence GSWIAAMDY. In some embodiments, the VH comprises a CDR-H2 comprising the amino acid sequence SISSWHGETGYADSVKG. In some embodiments, the VH comprises a CDR-H1 comprising the amino acid sequence FSPYSIH. In some embodiments, the antigen-binding domain further comprises a light chain variable region (VL), wherein the VL comprises a CDR-L3 comprising the amino acid sequence QQASYVRKTIT. In some embodiments, the VL comprises a CDR-L2 comprising the amino acid sequence SASSLYS. In some embodiments, the VL comprises a CDR-L1 comprising the amino acid sequence RASQSVSSAVA. In some embodiments, the antigen-binding domain comprises the following: a CDR-H3 sequence of GSWIAAMDY, a CDR-H2 sequence of SISSWHGETGYADSVKG, a CDR-H1 sequence of FSPYSIH, a CDR-L3 sequence of QQASYVRKTIT, a CDR-L2 sequence of SASSLYS, and a CDR-L1 sequence of RASQSVSSAVA. In some embodiments, the VH comprises a sequence having at least 80% sequence identity to the sequence EVQLVESGGGLVQPGGSLRLSCAASGFTFSPYSIHWVRQAPGKGLEWVASISSWHGETGYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARGSWIAAMDYWGQGTLVTVSS. In some embodiments, the VL comprises a sequence having at least 80% sequence identity to the sequence DIQMTQSPSSLSASVGDRVTITCRASQSVSSAVAWYQQKPGKAPKLLIYSASSLYSGVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQASYVRKTITFGQGTKVEIKRTV.

[0098] In some embodiments, the antigen-binding domain comprises a heavy chain variable region (VH), wherein the VH comprises a heavy chain complementarity-determining region 3 (CDR-H3) comprising the amino acid sequence GGWIAAMDY. In some embodiments, the VH comprises a CDR-H2 comprising the amino acid sequence SISSLQGDTGYADSVKG. In some embodiments, the VH comprises a CDR-H1 comprising the amino acid sequence FSWYSIH. In some embodiments, the antigen-binding domain further comprises a light chain variable region (VL), wherein the VL comprises a CDR-L3 comprising the amino acid sequence QQASYVRKTIT. In some embodiments, the VL comprises a CDR-L2 comprising the amino acid sequence SASSLYS. In some embodiments, the VL comprises a CDR-L1 comprising the amino acid sequence RASQSVSSAVA. In some embodiments, the antigen-binding domain comprises the following: a CDR-H3 sequence of GGWIAAMDY, a CDR-H2 sequence of SISSLQGDTGYADSVKG, a CDR-H1 sequence of FSWYSIH, a CDR-L3 sequence of QQASYVRKTIT, a CDR-L2 sequence of SASSLYS, and a CDR-L1 sequence of RASQSVSSAVA. In some embodiments, the VH comprises a sequence having at least 80% sequence identity to the sequence EVQLVESGGGLVQPGGSLRLSCAASGFTFSWYSIHWVRQAPGKGLEWVASISSLQGDTGYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARGGWIAAMDYWGQGTLVTVSS. In some embodiments, the VL comprises a sequence having at least 80% sequence identity to the sequence DIQMTQSPSSLSASVGDRVTITCRASQSVSSAVAWYQQKPGKAPKLLIYSASSLYSGVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQASYVRKTITFGQGTKVEIKRTV.

[0099] In some embodiments, the antigen-binding domain comprises a heavy chain variable region (VH), wherein the VH comprises a heavy chain complementarity-determining region 3 (CDR-H3) comprising the amino acid sequence GSWIAAMDY. In some embodiments, the VH comprises a CDR-H2 comprising the amino acid sequence SIASWYGDTGYADSVKG. In some embodiments, the VH comprises a CDR-H1 comprising the amino acid sequence FHYYSIH. In some embodiments, the antigen-binding domain further comprises a light chain variable region (VL), wherein the VL comprises a CDR-L3 comprising the amino acid sequence QQASYVRKTIT. In some embodiments, the VL comprises a CDR-L2 comprising the amino acid sequence SASSLYS. In some embodiments, the VL comprises a CDR-L1 comprising the amino acid sequence RASQSVSSAVA. In some embodiments, the antigen-binding domain comprises the following: a CDR-H3 sequence of GSWIAAMDY, a CDR-H2 sequence of SIASWYGDTGYADSVKG, a CDR-H1 sequence of FHYYSIH, a CDR-L3 sequence of QQASYVRKTIT, a CDR-L2 sequence of SASSLYS, and a CDR-L1 sequence of RASQSVSSAVA. In some embodiments, the VH comprises a sequence having at least 80% sequence identity to the sequence EVQLVESGGGLVQPGGSLRLSCAASGFTFHYYSIHWVRQAPGKGLEWVASIASWYGDTGYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARGSWIAAMDYWGQGTLVTVSS. In some embodiments, the VL comprises a sequence having at least 80% sequence identity to the sequence DIQMTQSPSSLSASVGDRVTITCRASQSVSSAVAWYQQKPGKAPKLLIYSASSLYSGVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQASYVRKTITFGQGTKVEIKRTV.

[0100] In some embodiments, the antigen-binding domain comprises a heavy chain variable region (VH), wherein the VH comprises a heavy chain complementarity-determining region 3 (CDR-H3) comprising the amino acid sequence GGRIEAMDY. In some embodiments, the VH comprises a CDR-H2 comprising the amino acid sequence SISSWYGKTGYADSVKG. In some embodiments, the VH comprises a CDR-H1 comprising the amino acid sequence FGYYSIH. In some embodiments, the antigen-binding domain further comprises a light chain variable region (VL), wherein the VL comprises a CDR-L3 comprising the amino acid sequence QQASYVRKTIT. In some embodiments, the VL comprises a CDR-L2 comprising the amino acid sequence SASSLYS. In some embodiments, the VL comprises a CDR-L1 comprising the amino acid sequence RASQSVSSAVA. In some embodiments, the antigen-binding domain comprises the following: a CDR-H3 sequence of GGRIEAMDY, a CDR-H2 sequence of SISSWYGKTGYADSVKG, a CDR-H1 sequence of FGYYSIH, a CDR-L3 sequence of QQASYVRKTIT, a CDR-L2 sequence of SASSLYS, and a CDR-L1 sequence of RASQSVSSAVA. In some embodiments, the VH comprises a sequence having at least 80% sequence identity to the sequence EVQLVESGGGLVQPGGSLRLSCAASGFTFGYYSIHWVRQAPGKGLEWVASISSWYGKTGYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARGGRIEAMDYWGQGTLVTVSS. In some embodiments, the VL comprises a sequence having at least 80% sequence identity to the sequence DIQMTQSPSSLSASVGDRVTITCRASQSVSSAVAWYQQKPGKAPKLLIYSASSLYSGVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQASYVRKTITFGQGTKVEIKRTV.

[0101] In some embodiments, the antigen-binding domain comprises a heavy chain variable region (VH), wherein the VH comprises a heavy chain complementarity-determining region 3 (CDR-H3) comprising the amino acid sequence GYWIEAMDY. In some embodiments, the VH comprises a CDR-H2 comprising the amino acid sequence SIASSYGSTGYADSVKG. In some embodiments, the VH comprises a CDR-H1 comprising the amino acid sequence FSKYSIH. In some embodiments, the antigen-binding domain further comprises a light chain variable region (VL), wherein the VL comprises a CDR-L3 comprising the amino acid sequence QQASYVRKTIT. In some embodiments, the VL comprises a CDR-L2 comprising the amino acid sequence SASSLYS. In some embodiments, the VL comprises a CDR-L1 comprising the amino acid sequence RASQSVSSAVA. In some embodiments, the antigen-binding domain comprises the following: a CDR-H3 sequence of GYWIEAMDY, a CDR-H2 sequence of SIASSYGSTGYADSVKG, a CDR-H1 sequence of FSKYSIH, a CDR-L3 sequence of QQASYVRKTIT, a CDR-L2 sequence of SASSLYS, and a CDR-L1 sequence of RASQSVSSAVA. In some embodiments, the VH comprises a sequence having at least 80% sequence identity to the sequence EVQLVESGGGLVQPGGSLRLSCAASGFTFSKYSIHWVRQAPGKGLEWVASIASSYGSTGYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARGYWIEAMDYWGQGTLVTVSS. In some embodiments, the VL comprises a sequence having at least 80% sequence identity to the sequence DIQMTQSPSSLSASVGDRVTITCRASQSVSSAVAWYQQKPGKAPKLLIYSASSLYSGVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQASYVRKTITFGQGTKVEIKRTV.

[0102] In some embodiments, the antigen-binding domain comprises a heavy chain variable region (VH), wherein the VH comprises a heavy chain complementarity-determining region 3 (CDR-H3) comprising the amino acid sequence GSWIAAMDY. In some embodiments, the VH comprises a CDR-H2 comprising the amino acid sequence SIHSSIGTTGYADSVKG. In some embodiments, the VH comprises a CDR-H1 comprising the amino acid sequence FGLYSIH. In some embodiments, the antigen-binding domain further comprises a light chain variable region (VL), wherein the VL comprises a CDR-L3 comprising the amino acid sequence QQASYVRKTIT. In some embodiments, the VL comprises a CDR-L2 comprising the amino acid sequence SASSLYS. In some embodiments, the VL comprises a CDR-L1 comprising the amino acid sequence RASQSVSSAVA. In some embodiments, the antigen-binding domain comprises the following: a CDR-H3 sequence of GSWIAAMDY, a CDR-H2 sequence of SIHSSIGTTGYADSVKG, a CDR-H1 sequence of FGLYSIH, a CDR-L3 sequence of QQASYVRKTIT, a CDR-L2 sequence of SASSLYS, and a CDR-L1 sequence of RASQSVSSAVA. In some embodiments, the VH comprises a sequence having at least 80% sequence identity to the sequence EVQLVESGGGLVQPGGSLRLSCAASGFTFGLYSIHWVRQAPGKGLEWVASIHSSIGTTGYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARGSWIAAMDYWGQGTLVTVSS. In some embodiments, the VL comprises a sequence having at least 80% sequence identity to the sequence DIQMTQSPSSLSASVGDRVTITCRASQSVSSAVAWYQQKPGKAPKLLIYSASSLYSGVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQASYVRKTITFGQGTKVEIKRTV.

[0103] In some embodiments, the antigen-binding domain comprises a heavy chain variable region (VH), wherein the VH comprises a heavy chain complementarity-determining region 3 (CDR-H3) comprising the amino acid sequence GSVIHAMDY. In some embodiments, the VH comprises a CDR-H2 comprising the amino acid sequence SILSWIGKTSYADSVKG. In some embodiments, the VH comprises a CDR-H1 comprising the amino acid sequence FSPYSIH. In some embodiments, the antigen-binding domain further comprises a light chain variable region (VL), wherein the VL comprises a CDR-L3 comprising the amino acid sequence QQASYVRKTIT. In some embodiments, the VL comprises a CDR-L2 comprising the amino acid sequence SASSLYS. In some embodiments, the VL comprises a CDR-L1 comprising the amino acid sequence RASQSVSSAVA. In some embodiments, the antigen-binding domain comprises the following: a CDR-H3 sequence of GSVIHAMDY, a CDR-H2 sequence of SILSWIGKTSYADSVKG, a CDR-H1 sequence of FSPYSIH, a CDR-L3 sequence of QQASYVRKTIT, a CDR-L2 sequence of SASSLYS, and a CDR-L1 sequence of RASQSVSSAVA. In some embodiments, the VH comprises a sequence having at least 80% sequence identity to the sequence EVQLVESGGGLVQPGGSLRLSCAASGFTFSPYSIHWVRQAPGKGLEWVASILSWIGKTSYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARGSVIHAMDYWGQGTLVTVSS. In some embodiments, the VL comprises a sequence having at least 80% sequence identity to the sequence DIQMTQSPSSLSASVGDRVTITCRASQSVSSAVAWYQQKPGKAPKLLIYSASSLYSGVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQASYVRKTITFGQGTKVEIKRTV.

[0104] In some embodiments, the antigen-binding domain comprises a heavy chain variable region (VH), wherein the VH comprises a heavy chain complementarity-determining region 3 (CDR-H3) comprising the amino acid sequence GGWIAAMDY. In some embodiments, the VH comprises a CDR-H2 comprising the amino acid sequence SIASRWGHTGYADSVKG. In some embodiments, the VH comprises a CDR-H1 comprising the amino acid sequence FSPYHIH. In some embodiments, the antigen-binding domain further comprises a light chain variable region (VL), wherein the VL comprises a CDR-L3 comprising the amino acid sequence QQASYVRKTIT. In some embodiments, the VL comprises a CDR-L2 comprising the amino acid sequence SASSLYS. In some embodiments, the VL comprises a CDR-L1 comprising the amino acid sequence RASQSVSSAVA. In some embodiments, the antigen-binding domain comprises the CDR-H3 sequence of GGWIAAMDY, the CDR-H2 sequence of SIASRWGHTGYADSVKG, the CDR-H1 sequence of FSPYHIH, the CDR-L3 sequence of QQASYVRKTIT, the CDR-L2 sequence of SASSLYS, and the CDR-L1 sequence of RASQSVSSAVA. In some embodiments, the VH comprises a sequence having at least 80% sequence identity to the sequence EVQLVESGGGLVQPGGSLRLSCAASGFTFSPYHIHWVRQAPGKGLEWVASIASRWGHTGYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARGGWIAAMDYWGQGTLVTVSS. In some embodiments, the VL comprises a sequence having at least 80% sequence identity to the sequence DIQMTQSPSSLSASVGDRVTITCRASQSVSSAVAWYQQKPGKAPKLLIYSASSLYSGVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQASYVRKTITFGQGTKVEIKRTV.

[0105] In some embodiments, the antigen-binding domain comprises a heavy chain variable region (VH), wherein the VH comprises a heavy chain complementarity-determining region 3 (CDR-H3) comprising the amino acid sequence GSWIAAMDY. In some embodiments, the VH comprises a CDR-H2 comprising the amino acid sequence SIASLQGITGYADSVKG. In some embodiments, the VH comprises a CDR-H1 comprising the amino acid sequence FHEYSIH. In some embodiments, the antigen-binding domain further comprises a light chain variable region (VL), wherein the VL comprises a CDR-L3 comprising the amino acid sequence QQASYVRKTIT. In some embodiments, the VL comprises a CDR-L2 comprising the amino acid sequence SASSLYS. In some embodiments, the VL comprises a CDR-L1 comprising the amino acid sequence RASQSVSSAVA. In some embodiments, the antigen-binding domain comprises the CDR-H3 sequence of GSWIAAMDY, the CDR-H2 sequence of SIASLQGITGYADSVKG, the CDR-H1 sequence of FHEYSIH, the CDR-L3 sequence of QQASYVRKTIT, the CDR-L2 sequence of SASSLYS, and the CDR-L1 sequence of RASQSVSSAVA. In some embodiments, the VH comprises a sequence having at least 80% sequence identity to the sequence EVQLVESGGGLVQPGGSLRLSCAASGFTFHEYSIHWVRQAPGKGLEWVASIASLQGITGYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARGSWIAAMDYWGQGTLVTVSS. In some embodiments, the VL comprises a sequence having at least 80% sequence identity to the sequence DIQMTQSPSSLSASVGDRVTITCRASQSVSSAVAWYQQKPGKAPKLLIYSASSLYSGVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQASYVRKTITFGQGTKVEIKRTV.

[0106] In some embodiments, the antigen-binding domain comprises a heavy chain variable region (VH), wherein the VH comprises a heavy chain complementarity-determining region 3 (CDR-H3) comprising the amino acid sequence GGVIHAMDY. In some embodiments, the VH comprises a CDR-H2 comprising the amino acid sequence SILSRWGVTSYADSVKG. In some embodiments, the VH comprises a CDR-H1 comprising the amino acid sequence FSDYSIH. In some embodiments, the antigen-binding domain further comprises a light chain variable region (VL), wherein the VL comprises a CDR-L3 comprising the amino acid sequence QQASYVRKTIT. In some embodiments, the VL comprises a CDR-L2 comprising the amino acid sequence SASSLYS. In some embodiments, the VL comprises a CDR-L1 comprising the amino acid sequence RASQSVSSAVA. In some embodiments, the antigen-binding domain comprises the CDR-H3 sequence of GGVIHAMDY, the CDR-H2 sequence of SILSRWGVTSYADSVKG, the CDR-H1 sequence of FSDYSIH, the CDR-L3 sequence of QQASYVRKTIT, the CDR-L2 sequence of SASSLYS, and the CDR-L1 sequence of RASQSVSSAVA. In some embodiments, the VH comprises a sequence having at least 80% sequence identity to the sequence EVQLVESGGGLVQPGGSLRLSCAASGFTFSDYSIHWVRQAPGKGLEWVASILSRWGVTSYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARGGVIHAMDYWGQGTLVTVSS. In some embodiments, the VL comprises a sequence having at least 80% sequence identity to the sequence DIQMTQSPSSLSASVGDRVTITCRASQSVSSAVAWYQQKPGKAPKLLIYSASSLYSGVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQASYVRKTITFGQGTKVEIKRTV.

[0107] In some embodiments, the antigen-binding domain comprises a heavy chain variable region (VH), wherein the VH comprises a heavy chain complementarity-determining region 3 (CDR-H3) comprising the amino acid sequence GGVIHAMDY. In some embodiments, the VH comprises a CDR-H2 comprising the amino acid sequence SISSRWGVTSYADSVKG. In some embodiments, the VH comprises a CDR-H1 comprising the amino acid sequence FSDYSIH. In some embodiments, the antigen-binding domain further comprises a light chain variable region (VL), wherein the VL comprises a CDR-L3 comprising the amino acid sequence QQASYVRKTIT. In some embodiments, the VL comprises a CDR-L2 comprising the amino acid sequence SASSLYS. In some embodiments, the VL comprises a CDR-L1 comprising the amino acid sequence RASQSVSSAVA. In some embodiments, the antigen-binding domain comprises the CDR-H3 sequence of GGVIHAMDY, the CDR-H2 sequence of SISSRWGVTSYADSVKG, the CDR-H1 sequence of FSDYSIH, the CDR-L3 sequence of QQASYVRKTIT, the CDR-L2 sequence of SASSLYS, and the CDR-L1 sequence of RASQSVSSAVA. In some embodiments, the VH comprises a sequence having at least 80% sequence identity to the sequence EVQLVESGGGLVQPGGSLRLSCAASGFTFSDYSIHWVRQAPGKGLEWVASISSRWGVTSYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARGGVIHAMDYWGQGTLVTVSS. In some embodiments, the VH comprises a sequence having at least 80% sequence identity to the sequence EVQLVESGGGLVQPGGSLRLSCAASGFTFSDYSIHWVRQAPGKGLEWVASISSRWGVTSYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARGGVIHAMDYWGQGTLVTVSS.

[0108] In some embodiments, the antibody or antigen-binding fragment specifically binds to the peptide conjugate / MHC complex, wherein the antibody or antigen-binding fragment interacts with the MHC of the peptide conjugate / MHC complex. In some embodiments, the peptide conjugate / MHC complex comprises: (a) a peptide conjugate comprising a peptide covalently bound to a targeted covalent inhibitor or fragment thereof; and (b) an MHC. In some embodiments, the MHC is a human leukocyte antigen (HLA). In some embodiments, the HLA is an HLA-A*03:01, HLA-A*11:01, and / or HLA-A*02:01 molecule.

[0109] In some embodiments, the antigen-binding domain binds to the peptide conjugate / MHC complex with higher affinity than the peptide or free target covalent inhibitor. In some embodiments, the antigen-binding domain binds to the peptide conjugate / MHC complex with a dissociation constant (K) of at most about 50 nM, at most about 40 nM, at most about 30 nM, at most about 20 nM, at most about 10 nM, at most about 1 nM, at most about 0.1 nM, at most about 10 pM, at most about 1 pM, or at most about 0.1 pM. D ) that bind to a peptide conjugate / MHC complex disclosed herein. In some embodiments, the antigen-binding domain binds to a peptide conjugate / MHC complex disclosed herein with a dissociation constant (K) of about 0.01 nM to about 20 nM. D In some embodiments, the antigen-binding domain binds to the peptide conjugate / MHC complex with a dissociation constant (K) of at least about 100 nM, at least about 200 nM, at least about 300 nM, at least about 400 nM, at least about 500 nM, at least about 1 μM, at least about 10 μM, at least about 20 μM, at least about 30 μM, at least about 40 μM, at least about 50 μM, or at least 100 μM. D ) binds to a free target covalent inhibitor. In some embodiments, the antigen-binding domain is a K of an antibody or antigen-binding fragment that binds to a peptide conjugate / MHC complex. Da dissociation constant (K) that is at least about 10-fold, 100-fold, 10,000-fold, or 100,000-fold higher than D ) binds to the free target covalent inhibitor. In some embodiments, the antigen-binding domain does not detectably bind to the free target covalent inhibitor. In some embodiments, the antigen-binding domain binds to the free target covalent inhibitor with an IC of at least about 50 nM. 50 In some embodiments, the antigen-binding domain binds to a free target covalent inhibitor at a dissociation constant (K) of greater than about 100 nM, greater than about 200 nM, greater than about 300 nM, greater than about 400 nM, greater than about 500 nM, greater than 1 μM, greater than 10 μM, greater than 20 μM, greater than 30 μM, greater than 40 μM, greater than 50 μM, or greater than 100 μM. D In some embodiments, the antigen-binding domain binds to the free peptide conjugate at a K D at least about 10-fold, 100-fold, 10,000-fold, or 100,000-fold higher dissociation constants (K D In some embodiments, the antigen-binding domain binds to the free peptide conjugate at a K D At least 2.5 times higher dissociation constant (K D In some embodiments, the antigen-binding domain binds to the free peptide conjugate at a K D higher dissociation constant (K D ) and binds to the free peptide conjugate, where the MHC is HLA-A02:01. In some embodiments, the antigen-binding domain comprises a K D at least 2.5-fold higher dissociation constant (K D), where the MHC is HLA-A03:01. In some embodiments, the antigen-binding domain binds to the peptide conjugate / MHC complex with an affinity that is at least 100-fold, at least 200-fold, at least 300-fold, at least 400-fold, at least 500-fold, at least 600-fold, at least 700-fold, at least 800-fold, at least 900-fold, at least 1,000-fold, at least 2,500-fold, at least 5,000-fold, or at least 10,000-fold greater than the affinity of the antibody or antigen-binding fragment for the free target covalent inhibitor or the free peptide conjugate.

[0110] In some embodiments, the peptide conjugate comprises a free target covalent inhibitor and a KRAS G12C Peptides, KRAS G12D Peptides, KRAS G12R , or KRAS G12S The peptide conjugate is formed by a covalent bond reaction with a KRAS peptide. In some embodiments, the free target covalent inhibitor is osimertinib, ibrutinib, neratinib, sotorasib, ARS-853, ARS-1620, ARS-3248, MRTX849, JNJ74699157, LY3499446, LY3537982, MRTX-1257, JDQ443, MRTX-1133, RMC-6291, RMC-9805, GDC-6036, D-1553, 2E07, 6H05, SML-8-73-1, or BI 182391. In some embodiments, the peptide conjugate is formed by a covalent bond reaction between sotorasib and a KRAS peptide. G12C In some embodiments, the peptide comprises or consists of the amino acid sequence VVVGACGVGK, VVGACGVGK, or KLVVVGACGV.

[0111] In some embodiments, the antigen binding domain comprises: (i) having specificity for a peptide conjugate / MHC complex comprising VVGACGVGK conjugated to a targeted covalent inhibitor or fragment thereof presented by HLA-A*11:01, and / or a peptide conjugate / MHC complex comprising VVVGACGVGK conjugated to a targeted covalent inhibitor or fragment thereof presented by HLA-A*11:01; or (ii) has specificity for a peptide conjugate / MHC complex comprising VVGACGVGK conjugated to a targeted covalent inhibitor or fragment thereof presented by HLA-A*03:01, and / or a peptide conjugate / MHC complex comprising VVVGACGVGK conjugated to a targeted covalent inhibitor or fragment thereof presented by HLA-A*03:01. In some embodiments, the antigen binding domain has specificity for a peptide conjugate / MHC complex comprising VVVGACGVGK conjugated to a targeted covalent inhibitor or fragment thereof presented by HLA-A*03:01, and / or a peptide conjugate / MHC complex comprising VVVGACGVGK conjugated to a targeted covalent inhibitor or fragment thereof presented by HLA-A*11:01. In some embodiments, the antigen binding domain has specificity for the following peptide conjugate / MHC complexes: a peptide conjugate / MHC complex comprising VVGACGVGK conjugated to a targeted covalent inhibitor or fragment thereof presented by HLA-A*11:01; a peptide conjugate / MHC complex comprising VVGACGVGK conjugated to a targeted covalent inhibitor or fragment thereof presented by HLA-A*03:01; a peptide conjugate / MHC complex comprising VVVGACGVGK conjugated to a targeted covalent inhibitor or fragment thereof presented by HLA-A*03:01; a peptide conjugate / MHC complex comprising VVVGACGVGK conjugated to a targeted covalent inhibitor presented by HLA-A*11:01 or a fragment thereof, and / or A peptide conjugate / MHC complex comprising KLVVVGACGV conjugated to a targeted covalent inhibitor or fragment thereof presented by HLA-A*02:01.

[0112] In some embodiments, the polypeptide binds to: (i) a peptide conjugate / HLA-A*11:01 MHC complex comprising a peptide consisting of the amino acid sequence VVGACGVGK, and a peptide conjugate / HLA-A*11:01 MHC complex comprising a peptide consisting of the amino acid sequence VVVGACGVGK; (ii) a peptide conjugate / HLA-A*03:01 MHC complex comprising a peptide consisting of the amino acid sequence VVGACGVGK, and a peptide conjugate / HLA-A*03:01 MHC complex comprising a peptide consisting of the amino acid sequence VVVGACGVGK; (iii) a peptide conjugate / HLA-A*03:01 MHC complex comprising a peptide consisting of the amino acid sequence VVGACGVGK, and a peptide conjugate / HLA-A*11:01 MHC complex comprising a peptide consisting of the amino acid sequence VVGACGVGK; (iv) a peptide conjugate / HLA-A*03:01 MHC complex comprising a peptide consisting of the amino acid sequence VVVGACGVGK, and a peptide conjugate / HLA-A*11:01 MHC complex comprising a peptide consisting of the amino acid sequence VVVGACGVGK; and / or (v) A peptide conjugate / HLA-A*02:01 MHC complex comprising a peptide consisting of the amino acid sequence KLVVVGACGV.

[0113] In some embodiments, the polypeptide binds to: a peptide conjugate / HLA-A*03:01 MHC complex comprising a peptide consisting of the amino acid sequence VVGACGVGK; a peptide conjugate / HLA-A*03:01 MHC complex comprising a peptide consisting of the amino acid sequence VVVGACGVGK; a peptide conjugate / HLA-A*11:01 MHC complex comprising a peptide consisting of the amino acid sequence VVVGACGVGK; a peptide conjugate / HLA-A*11:01 MHC complex comprising a peptide consisting of the amino acid sequence VVVGACGVGK; and a peptide conjugate / HLA-A*02:01 MHC complex comprising a peptide consisting of the amino acid sequence KLVVVGACGV.

[0114] In some embodiments, the targeted covalent inhibitor has a chemical structure comprising C-R1, where C is a chemical fragment linked to R1 of any compound selected from the group consisting of: [ka] where R1 is selected from: [ka]

[0115] In some embodiments, the targeted covalent inhibitor forms a covalent bond with (i) a cysteine residue in a peptide comprising the amino acid sequence of VVVGACGVGK, VVGACGVGK, or KLVVVGACGV, (ii) an aspartic acid residue in a peptide comprising the amino acid sequence of VVVGADGVGK, VVGADGVGK, or KLVVVGADGV, (iii) a serine acid residue in a peptide comprising the amino acid sequence of VVVGASGVGK, VVGASGVGK, or KLVVVGASGV, or (iv) an arginine residue in a peptide comprising the amino acid sequence of VVVGARGVGK, VVGARGVGK, or KLVVVGARGV.

[0116] In some embodiments, the antigen binding domain of the polypeptide recognizes the C portion of a targeted covalent inhibitor of a peptide conjugate / MHC complex. In some embodiments, the antigen binding domain of the polypeptide recognizes the C portion of the targeted covalent inhibitor of the peptide conjugate / MHC complex, but does not recognize the R1 portion of the targeted covalent inhibitor.

[0117] In some embodiments, the interface area between the polypeptide and the peptide conjugate / MHC complex is at least about 500 Å 2 , 600Å 2 , 700Å 2 , 800Å 2 , 900Å 2 , 1,000Å 2 , 1,200Å 2 , 1,500Å 2 , or 2,000 Å 2 In some embodiments, the interfacial area between the polypeptide and the MHC of the peptide conjugate / MHC complex is greater than the interfacial area between the polypeptide and the peptide or targeted covalent inhibitor.

[0118] In some embodiments, the polypeptide forms a binding pocket at the interface between the VH and VL domains that accommodates a targeted covalent inhibitor of the peptide conjugate / MHC complex.

[0119] In some embodiments, the polypeptide does not bind to the peptide conjugate / MHC complex in a "head-to-head" coaxial interaction.

[0120] In some embodiments, the polypeptide binds to the peptide conjugate / MHC complex at an angle between the axis of the MHC and the axis of the polypeptide of about 10° to 60°. In some embodiments, when the polypeptide is bound to the peptide conjugate / MHC complex, the angle between the axis of the polypeptide and the axis of the peptide conjugate / MHC complex is about 10° to 60°; in some embodiments, when the polypeptide is bound to the peptide conjugate / MHC complex, the angle between the axis of the polypeptide and the axis of the peptide conjugate / MHC complex is different from the angle between the axis of the TCR and the axis of the peptide conjugate / MHC complex when the TCR is bound to the peptide conjugate / MHC complex. In some embodiments, when the polypeptide is bound to the peptide conjugate / MHC complex, the angle between the axis of the polypeptide and the axis of the peptide conjugate / MHC complex is about 10° to 60° less or about 10° to 60° greater than the angle between the axis of the TCR and the axis of the peptide conjugate / MHC complex when the TCR is bound to the peptide conjugate / MHC complex. In some embodiments, the polypeptide binds to the peptide conjugate / MHC complex at an angle that differs from the angle at which the T cell receptor binds to the peptide conjugate / MHC complex, ie, at an angle that is about 10° to 60° less than or about 10° to 60° greater than the angle at which the T cell receptor binds to the peptide conjugate / MHC complex.

[0121] In some embodiments, the polypeptide binds to the peptide conjugate / MHC complex at an angle between the axis of the MHC and the axis of the polypeptide of about 40°. In some embodiments, when the polypeptide is bound to the peptide conjugate / MHC complex, the angle between the axis of the polypeptide and the axis of the peptide conjugate / MHC complex is about 40° less than or about 40° more than the angle between the axis of the polypeptide and the axis of the peptide conjugate / MHC complex when the TCR is bound to the peptide conjugate / MHC complex. In some embodiments, the polypeptide binds to the peptide conjugate / MHC complex at an angle about 40° less than or about 40° more than the angle at which the T cell receptor binds to the peptide conjugate / MHC complex.

[0122] In some embodiments, the polypeptide contacts the α1 and α2 domains of the heavy chain of the MHC of the peptide conjugate / MHC complex. In some embodiments, the polypeptide binds to an epitope of the MHC, wherein the epitope comprises one or more residues from a region including residues 62-66, 106-109, and / or 150-170 of the MHC.

[0123] In some embodiments, the polypeptide binds to an epitope of MHC, wherein the epitope comprises one or more residues from a region including residues 62-66, 106-109, and / or 150-170 of an HLA-A*03:01 molecule or an HLA-A*11:01 molecule.

[0124] In some embodiments, the polypeptide binds to an epitope of an MHC, wherein the epitope comprises one or more residues selected from the group consisting of residues 62, 106, 108, 109, 158, 161, 162, 162, 165, 166, 167, 169 and 170 of an HLA-A*03:01 molecule.

[0125] In some embodiments, the VL domain of the antigen-binding domain binds to an epitope of MHC, wherein the epitope comprises one or more residues selected from the group consisting of residues 62, 106, 108, 109, 158, 161, 162, 162, 165, 166, 167, 169, and 170 of HLA-A*03:01.

[0126] In some embodiments, the polypeptide binds to an epitope of MHC, wherein the epitope comprises one or more residues selected from the group consisting of residues 62, 65, 66, 150, 151, 154, 155, 157 and 158 of HLA-A*03:01.

[0127] In some embodiments, the VH domain of the antigen-binding domain binds to an epitope of MHC, wherein the epitope comprises one or more residues selected from the group consisting of residues 62, 65, 66, 150, 151, 154, 155, 157 and 158 of HLA-A*03:01.

[0128] In some embodiments, the polypeptide binds to an epitope of MHC, wherein the epitope comprises one or more residues selected from the group consisting of residues 62, 106, 108, 109, 154, 157, 158, 161, 162, 163, 165, 166, 167, 169 and 170 of HLA-A*11:01.

[0129] In some embodiments, the VL domain of the antigen-binding domain binds to an epitope of MHC, wherein the epitope comprises one or more residues selected from the group consisting of residues 62, 106, 108, 109, 154, 157, 158, 161, 162, 163, 165, 166, 167, 169 and 170 of HLA-A*11:01.

[0130] In some embodiments, the polypeptide binds to an epitope of MHC, wherein the epitope comprises one or more residues selected from the group consisting of residues 62, 65, 66, 151, 154, 155 and 158 of HLA-A*11:01.

[0131] In some embodiments, the VH domain of the antigen-binding domain binds to an epitope of MHC, wherein the epitope includes one or more residues selected from the group consisting of residues 62, 65, 66, 151, 154, 155, and 158 of HLA-A*11:01. In some embodiments, the VH is linked to the VL via a linker. In some embodiments, the linker is (G4S) n or (S4G) n where n is any integer from 1 to 10. In some embodiments, the linker comprises the glycine-serine-alanine linker G4SA 3、 or a glycine-serine linker (G4S)4.

[0132] In some embodiments, the polypeptide is an intact antibody, a bispecific antibody, a multispecific antibody, an antigen-binding (Fab) fragment, a Fab' fragment, a (Fab')2 fragment, an Fd, an Fv, a dAb, a single domain fragment or a single monomeric variable antibody domain, a dual affinity retargeting (DART) molecule, a diabody (Db), a single-chain diabody (scDb), a single-chain variable fragment (scFv), a bispecific T-cell engager (BiTE), a bispecific killer cell engager (BiKE), a CrossMab, a camelid antibody, a trispecific binding partner, a chimeric antigen receptor (CAR), a monobody (also known as an adnectin), a DARPin, an anticalin, an affibody, or an affimer. In some embodiments, the polypeptide is a bispecific antibody. In some embodiments, the bispecific antibody is a bispecific T-cell engager (BiTE).

[0133] In some embodiments, the polypeptide further comprises a second antigen-binding domain that binds to a T cell surface marker. In some embodiments, the T cell surface marker is CD3 epsilon, CD3 gamma, CD3 delta, CD3 eta, TCR alpha, or TCR beta (of the TCR). In some embodiments, the antigen-binding domain and the second antigen-binding domain are linked by a linker. In some embodiments, the linker is (G4S) n or (S4G) n where n is any integer from 1 to 10.

[0134] In some embodiments, the linker is configured as follows: (a) The polypeptide binds to the peptide conjugate / MHC complex at an angle between the axis of the MHC and the axis of the polypeptide of approximately 10° to 60°; (b) when the polypeptide is bound to the peptide conjugate / MHC complex, the angle between the axis of the polypeptide and the axis of the peptide conjugate / MHC complex is between about 10° and 60°; (c) when the polypeptide is bound to the peptide conjugate / MHC complex, the angle between the axis of the polypeptide and the axis of the peptide conjugate / MHC complex is different from the angle between the axis of the TCR and the axis of the peptide conjugate / MHC complex when the TCR is bound to the peptide conjugate / MHC complex; (d) when the polypeptide is bound to the peptide conjugate / MHC complex, the angle between the axis of the polypeptide and the axis of the peptide conjugate / MHC complex is about 10° to 60° less than, or about 10° to 60° greater than, the angle between the axis of the TCR and the axis of the peptide conjugate / MHC complex when the TCR is bound to the peptide conjugate / MHC complex; (e) the polypeptide binds to the peptide conjugate / MHC complex at an angle that is different from the angle at which the T cell receptor binds to the peptide conjugate / MHC complex; and / or (f) The polypeptide binds to the peptide conjugate / MHC complex at an angle that is about 10° to 60° less than or about 10° to 60° greater than the angle at which the T cell receptor binds to the peptide conjugate / MHC complex.

[0135] In some embodiments, the polypeptide comprises a first polypeptide chain comprising an antigen-binding domain and a second polypeptide chain comprising a second antigen-binding domain. In some embodiments, the first polypeptide chain or the second polypeptide chain is further fused to a cytokine or a fragment thereof. In some embodiments, the cytokine comprises IL-2, IL-7, IL-15, IL-12, IL-18, or IL-21, or an interferon (IFN). In some embodiments, the peptide conjugate / MHC complex is displayed on the surface of the cell. In some embodiments, the cell expresses a low copy number of the peptide conjugate / MHC complex, wherein the low copy number is at most about 1000, 900, 800, 700, 600, 500, 400, 300, 200, 100, 90, 80, 70, 60, 50, 40, 30, 20, 10, 5, 3, or 1 copy per single cell.

[0136] In some embodiments, the peptide of the peptide conjugate / MHC complex is derived from an intracellular protein. In some embodiments, the polypeptide is a binding partner disclosed herein.

[0137] In another aspect, the present disclosure provides a polypeptide comprising an antigen-binding domain that specifically binds to a peptide conjugate / MHC complex, wherein the peptide conjugate / MHC complex comprises: (a) a peptide conjugate comprising a peptide covalently linked to a targeted covalent inhibitor, or fragment thereof; and (b) an MHC.

[0138] In another aspect, the present disclosure provides a pharmaceutical composition comprising a polypeptide disclosed herein and a pharmaceutically acceptable carrier. In another aspect, the present disclosure provides a method of treating cancer in a subject treated with a free covalent target inhibitor, the method comprising administering to the subject a polypeptide disclosed herein or a pharmaceutical composition disclosed herein. In some embodiments, the subject is refractory to treatment with the free covalent target inhibitor.

[0139] In another aspect, the present disclosure provides a method of treating cancer in a subject, the method comprising administering to the subject a polypeptide disclosed herein or a pharmaceutical composition disclosed herein after or simultaneously with administration of a small molecule drug. In another aspect, the present disclosure provides a method of treating cancer in a subject, the method comprising administering a targeted covalent inhibitor to the subject and administering a polypeptide disclosed herein or a pharmaceutical composition disclosed herein to the subject. In some embodiments, the polypeptide disclosed herein or the pharmaceutical composition disclosed herein is administered after administration of the targeted covalent inhibitor or simultaneously with the targeted covalent inhibitor.

[0140] In another aspect, the present disclosure provides a computer-assisted method for identifying or designing potential polypeptides comprising an antigen-binding domain that binds to a peptide conjugate / MHC complex, wherein the peptide conjugate of the peptide conjugate / MHC complex is a peptide covalently linked to a target covalent inhibitor or fragment thereof, the method comprising: (a) providing coordinates of at least two atoms of the peptide conjugate / MHC complex (i) of Figures 57A-C, 58A-B, 59A-C, 60A-B, 61; (b) providing a structure of a candidate polypeptide comprising an antigen-binding domain for binding to the peptide conjugate / MHC complex; (c) fitting the structure of the candidate polypeptide to said at least two atoms of the peptide conjugate / MHC complex, wherein fitting comprises determining interactions between one or more atoms of the antigen-binding domain of the candidate polypeptide and atoms of the peptide conjugate / MHC complex; and (d) selecting a candidate polypeptide that is predicted to bind to the peptide conjugate / MHC complex.

[0141] In another aspect, the present disclosure provides a computer-assisted method for designing potential polypeptides comprising an antigen-binding domain that binds to a peptide conjugate / MHC complex, wherein the peptide conjugate of the peptide conjugate / MHC complex is a peptide covalently linked to a target covalent inhibitor or fragment thereof, the method comprising: (a) providing a structure of a candidate polypeptide comprising an antigen-binding domain for binding to the peptide conjugate / MHC complex; (b) providing coordinates of at least two atoms of the peptide conjugate / MHC complex and coordinates of at least two atoms of the antigen-binding domain of a polypeptide bound to the peptide conjugate / MHC complex of Figures 57A-C, 58A-B, 59A-C, 60A-B, and 61; (c) providing a structure of a candidate polypeptide comprising an antigen-binding domain for binding to the peptide conjugate / MHC complex; (d) fitting the structure of the candidate polypeptide to at least two atoms of the peptide conjugate / MHC complex and at least two atoms of the antigen-binding domain of the polypeptide bound to the peptide conjugate / MHC complex, wherein fitting includes determining interactions between one or more atoms of the antigen-binding domain of the candidate polypeptide and atoms of the peptide conjugate / MHC complex; and

[0142] In another aspect, the present disclosure provides a computer-assisted method for designing potential polypeptides comprising an antigen-binding domain that binds to a peptide conjugate / MHC complex, wherein the peptide conjugate of the peptide conjugate / MHC complex is a peptide covalently linked to a target covalent inhibitor or fragment thereof, the method comprising: (a) providing a structure of a polypeptide comprising an antigen-binding domain bound to the peptide conjugate / MHC complex; (b) determining interactions between one or more atoms of the antigen-binding domain of the polypeptide and one or more atoms of the peptide conjugate / MHC complex; (c) providing a candidate polypeptide comprising an antigen-binding domain for binding to the peptide conjugate / MHC complex, wherein the antigen-binding domain of the candidate polypeptide comprises one or more amino acid substitutions relative to the polypeptide comprising the antigen-binding domain, wherein the one or more amino acid substitutions are of residues in the antigen-binding domain of the polypeptide that comprise one or more atoms that interact with or modulate an interaction with one or more atoms of the peptide conjugate / MHC complex; and (d) Selecting candidate polypeptides that bind or are predicted to bind to the peptide conjugate / MHC complex with an affinity higher than the affinity of the polypeptide comprising the antigen-binding domain for the peptide conjugate / MHC complex.

[0143] In another aspect, the present disclosure provides a computer-assisted method for identifying or designing potential polypeptides comprising an antigen-binding domain that binds to a peptide conjugate / MHC complex, wherein the peptide conjugate of the peptide conjugate / MHC complex is a peptide covalently linked to a target covalent inhibitor or fragment thereof, the method comprising using a computer system, e.g., using a programmed computer comprising a processor, a data storage system, an input device, and an output device, and comprising the following steps: (a) inputting data comprising three-dimensional coordinates of a subset of atoms derived from or related to the crystal structures of Figures 57A-C, 58A-B, 59A-C, 60A-B, 61 into the programmed computer via the input device, thereby generating a data set; (b) using the processor to compare the data set with a computer database of structures of polypeptides comprising an antigen-binding domain that bind, or are predicted to bind, or are desired to bind to a peptide conjugate / MHC complex, stored in the computer data storage system; (c) selecting from the database, using a computer method, a structure that has the potential to bind to a peptide conjugate / MHC complex; and (d) selecting from the database a structure that has the potential to bind to a peptide conjugate / MHC complex using a computer method. (e) using a computer method to construct a model of the selected structures; and (f) outputting the selected structures to the output device, and optionally synthesizing one or more selected structures; and further optionally testing the synthesized selected structures for binding to a peptide conjugate / MHC complex.

[0144] In another aspect, the present disclosure provides a computer-readable medium comprising: atomic coordinate data according to the structure of any one of Figures 57A-C, 58A-B, 59A-C, 60A-B, and 61, said data defining or providing a three-dimensional structure of a polypeptide comprising an antigen-binding domain bound to a peptide conjugate / MHC complex, wherein the peptide conjugate of the peptide conjugate / MHC complex is a peptide covalently linked to a targeted covalent inhibitor, or a fragment thereof, or at least one subdomain thereof; or structure factor data of a polypeptide comprising an antigen-binding domain bound to the peptide conjugate / MHC complex, wherein the structure factor data can be derived from the atomic coordinate data of any of Figures 57A-C, 58A-B, 59A-C, 60A-B, and 61.

[0145] In another aspect, the present disclosure provides a computer readable medium comprising atomic coordinate data according to the structure of any one of Figures 57A-C, 58A-B, 59A-C, 60A-B, and 61, said data defining or providing a three-dimensional structure of a peptide conjugate / MHC complex, wherein the peptide conjugate of the peptide conjugate / MHC complex is a peptide covalently linked to a targeted covalent inhibitor or a fragment thereof, or at least one subdomain thereof, or structure factor data of the peptide conjugate / MHC complex, wherein the structure factor data can be derived from the atomic coordinate data of the structure of any one of Figures 57A-C, 58A-B, 59A-C, 60A-B, and 61.

[0146] In another aspect, the present disclosure provides a method for identifying a T cell receptor (TCR) that recognizes a peptide conjugate / MHC complex disclosed herein, the method comprising: (a) contacting a plurality of candidate TCRs with the peptide conjugate / MHC complex; and (b) identifying at least one TCR that binds to the peptide conjugate / MHC complex.

[0147] In another aspect, the present disclosure provides a method for identifying a T cell receptor (TCR) that recognizes a peptide conjugate / MHC complex disclosed herein, the method comprising: (a) culturing T cells and antigen-presenting cells (APCs) treated with a targeted covalent inhibitor, thereby generating peptide conjugate / MHC complex-specific T cells having a T cell receptor (TCR) that binds to the peptide conjugate / MHC complex; and (b) identifying at least one TCR from the peptide conjugate / MHC complex-specific T cells that bind to the peptide conjugate / MHC complex. In some embodiments, the method disclosed herein further comprises selecting or isolating at least one TCR.

[0148] In some embodiments, the plurality of candidate TCRs are a plurality of soluble TCRs or a plurality of TCRs expressed on the cell surface of a plurality of cells. In some embodiments, the plurality of candidate TCRs are a plurality of TCRs expressed on the cell surface of a plurality of cells, and the identifying in (b) comprises isolating or selecting cells comprising at least one TCR based on an activation marker of the cells. In some embodiments, the activation marker is a T cell activation marker. In some embodiments, the T cell activation marker is CD26, CD27, CD28, CD30, CD154, CD40L, CD134, CD 25, CD44, CD69, CD137 or KLRG1.

[0149] In another aspect, the present disclosure provides a T cell receptor (TCR) comprising at least one TCR disclosed herein. In some embodiments, the TCR is a soluble TCR. In some embodiments, the TCR is a bispecific TCR.

[0150] In some aspects disclosed herein, methods of treating cancer are disclosed. In some embodiments, the methods include prophylactically administering a peptide conjugate to a subject in need thereof before the subject develops cancer, before the subject is administered a drug, or both, wherein the peptide conjugate comprises a peptide covalently linked to a targeted covalent inhibitor or fragment thereof. [Brief explanation of the drawings]

[0151] [Figure 1A-C] Graphs showing the results of phage enzyme-linked immunosorbent assay (ELISA) of phage-displayed antibody clones. G12C -GDP and KRAS G12C Binding to the G12C-GDP-ARS-1620 conjugate was measured. For each clone, the bars in the graph represent, from left to right, buffer, G12C-GDP-ARS, and G12C-GDP. The results for clones 1-32 are shown in Figure 1A, clones 33-64 in Figure 1B, and clones 65-96 in Figure 1C.

[0152] [Figure 2] KRAS in the GTPγS- or GDP-bound nucleotide state (with or without ARS-1620) G12C Graph showing the binding of 12C-ARS Fab59 to IgG1A, IgG2A, and IgG3A using a bead binding assay.

[0153] [Figure 3] 12C-ARS-Fab59 conjugated to ARS-1620. G12C- Graph showing specific binding to GDP.

[0154] [Figure 4A-D] Figure 4A shows the ARS-1620 / KRAS expression level measured by pull-down assay of lysates prepared from cell lines. G12C Data obtained using 12C-ARS-Fab59 to measure adducts are shown in Figure 4B. RAS-less MEFs (Figure 4A) and KCP (Kras) reconstituted with the indicated KRAS mutants were treated with or without ARS-1620. G12C; Tp53 R172H Immunoblots of whole-cell lysates and 12C-ARS Fab pulldowns (PD) from (Pdx-Cre) mouse pancreatic cancer cells (Figure 4B). Figure 4C shows whole-cell lysates and 12C-ARS Fab pulldowns (PD) from H358 and MIAPaCa-2 cells treated as indicated. Figure 4D shows ARS adduct formation in the samples from Figure 4C, quantified by LC / MS-MS assay. The concentrations of ARS-1620 and SHP099 were 10 μM in all panels.

[0155] [Figures 5A and 5B] Data demonstrating that the involvement of ARS-1620 in mutant KRAS can be measured using 12C-ARS-Fab59 by pull-down assay using lysates prepared from animal tissues. Figures 5A and 5B show the effect of ARS-1620 on mutant KRAS after 3 days of oral administration of ARS-1620 (200 mg / kg / d) alone or the SHP2 inhibitor SHP099 (75 mg / kg / d). G12C -Tp53 R270H (Figure 5A) and LSL-KRAS G12C (FIG. 5B) Anti-pan RAS and anti-ERK2 (loading control) immunoblots of lysates from tumors and 12C-ARS Fab pulldowns (PD).

[0156] Figure 6: Binding of antibody clones to AMG-510 conjugated to KRAS(G12C) peptide and poly-Ser (irrelevant) peptide. For each antibody clone, the bars represent, from left to right, no target, KRAS(G12C), KRAS(G12C)-AMG, and irrelevant peptide-AMG. The signals for the two negative controls, no target and KRAS(G12C) peptide without conjugated drug, were too low to be visible on the graph. Antibody clones were displayed on yeast cells, and binding of the target conjugated to fluorescently labeled streptavidin was detected by flow cytometry.

[0157] [Figure 7] Binding of the P2AMR-1 clone in human IgG1 format to AMG-510 conjugated to the KRAS(G12C) peptide (circles) and the same peptide without drug conjugation (squares). The peptide was immobilized on streptavidin-coated beads, and the antibody bound to the beads was detected with a fluorescently labeled secondary antibody. Apparent K D The values shown are from triplicate measurements. Error bars are within the symbol size.

[0158] [Figure 8] Recognition of AMG-510 presented on class I MHC molecules. AMG510-RAS(G12C) conjugate (circle) and unconjugated peptide (square) were loaded onto HLA-A*03:01 and immobilized on streptavidin-coated beads. Antibody binding was detected as in Figure 2. Apparent KDs are shown. Data shown are from triplicate determinations. Error bars are within the symbol size.

[0159] [Figure 9] HapImmune TM A cartoon-style representation of the disclosed concept called.

[0160] [Figures 10A-C] Data demonstrating the development of antibodies that bind to MHC / peptide-drug conjugate complexes. Figure 10A shows a multiplex bead binding assay (MBBA) of phage displaying different antibody clones. Figure 10B shows an MBBA assay of phage displaying different antibody clones against HLA-A*01:01 in complex with a BTK peptide conjugated to ibrutinib. Figure 10C shows an MBBA assay of phage displaying different antibody clones against HLA-A*02:01 in complex with an EGFR peptide conjugated to osimertinib.

[0161] [Figures 11A-C] Binding titration graphs using multiplex bead binding assays (MBBA) of purified antibodies targeting the KRAS(G12C)-AMG510 conjugate. Clone names are listed above each graph. Graphs for clones AMRA3-7 hIgG1, AMRA3-18 hIgG1, and AMRA3-22 hIgG1 are shown in Figure 11A, for clones AMRA11-2 hIgG1 and AMRA11-15 hIgG1 in Figure 11B, and for clones AMRA311-16 hIgG1, AMRA311-17 hIgG1, and AMRA311-18 hIgG1 in Figure 11C. Antigen nomenclature is described in Figure 10.

[0162]

[0023] FIG. 12 is a graph showing that antibody binding to HLA-complexed AMG510-peptide conjugates is not affected by the presence of free drug AMG-510.

[0163] Figure 13: Graphs of binding titration using multiplex bead binding assay (MBBA) of purified antibodies targeting BTK-ibrutinib conjugates. Clone names are listed above each graph. Antigen nomenclature is as in Figure 10.

[0164] [Figures 14A and 14B] Graphs of binding titration of purified antibodies to KRAS(G12C)-AMG510 conjugates presented by endogenous HLA molecules on the cell surface. Raji cells were first incubated with KRAS(G12C)-AMG510 conjugates or KRAS (wild-type) peptide, and excess conjugate and peptide were washed away. The amount of antibody detected using a fluorescently labeled secondary antibody is shown as a function of the IgG concentration used for staining. Apparent dissociation constant (KD) values were determined using nonlinear least-squares fitting of a 1:1 binding function. Data shown are from triplicate determinations. Graphs for antibodies AMRA3-7 hIgG1, AMRA3-22 hIgG1, AMRA3-18 hIgG1, and AMRA11-2 hIgG1 are shown in Figure 14A, and graphs for antibodies AMRA3-7 hIgG1, AMRA3-22 hIgG1, AMRA3-18 hIgG1, and AMRA11-2 hIgG1 are shown in Figure 14B.

[0165] [Figures 15A-C] Antibody binding to KRAS(G12C)-expressing cell lines pretreated with AMG-510. Figure 15A shows flow cytometry histograms. Figure 15B shows quantification of median fluorescence intensity of H358 cells treated with or without AMG-510. Figure 15C shows quantification of median fluorescence intensity of H358 cells and HEK293T cells (negative control) treated with or without AMG-510 and stained with AMRA3-7 antibody.

[0166] [Figure 16] Graph showing binding of P2AMR-1 IgG to cells preincubated with KRAS(G12C) peptide-AMG510 conjugate, KRAS(wild-type) peptide, or no peptide.

[0167] [Figures 17A and 17B] Graphs showing binding of purified antibodies in IgG format to the indicated drug-peptide / HLA complexes, measured using a multiplex bead-binding assay (MBBA). Figure 17A shows antibody clones identified using AMG-510 conjugated to the KRAS(G12C) peptide complexed with HLA-A*03:01 as the antigen. Figure 17B shows antibody clones identified using AMG-510 conjugated to the KRAS(G12C) peptide complexed with HLA-*11:01 as the antigen.

[0168] [Figures 18A and 18B] Graphs showing the cytotoxic effect of single-chain diabodies (scDbs) on cells pulsed with exogenous peptide-drug conjugates. Figure 18A shows Raji cells first pulsed with AMG-510 conjugated to a peptide corresponding to a fragment of KRAS (G12C) or a control peptide corresponding to KRAS (wild-type). The pulsed cells were co-cultured with human T cells (effector:target = 3:1) in the presence of the indicated concentrations of single-chain diabodies (scDbs). After incubation, dead cells were stained and detected by flow cytometry. Data shown are from triplicate measurements. Error bars indicate standard deviation. Where error bars are not visible, the error is smaller than the symbol. Figure 18B shows a comparable experiment using T2 cells and osimertinib conjugated to an EGFR peptide. A peptide conjugated to β-mercaptoethanol was used as a negative control.

[0169] [Figures 19A and 19B] Graphs showing the specific cytotoxicity of AMRA3-7_UCHT1 scDb against drug-treated lung cancer cell lines. Figure 19A shows lung cancer cell lines treated with 100 nM AMG-510 for 24 hours and then cocultured with human T cells (E:T = 5:1) in the presence of AMRA3-7_UCHT1 scDb. After incubation, cell viability was measured. The scDb antibody showed dose-dependent cytotoxicity only against AMG510-treated cells harboring the cognate KRAS mutation (G12C) and HLA (HLA-A3). Figure 19B shows the cytotoxicity of 0.1 nM of scDb. Data shown here are from quadruplicate assays.

[0170] Figure 20 shows the binding titration curves of AMR-A3-7 and AMR-A3-7D displayed on the yeast cell surface to HLA-A*03:01 presenting the 9-mer or 10-mer RAS(G12C) peptide (VVGACGVGK and VVVGACGVGK, respectively, with AMG-510 conjugated to their Cys residues).

[0171] [Figures 21A and 21B] Graphs showing the cytotoxicity of AMRA3-7D scDb. Figure 21A shows the dose-dependent cytotoxicity of AMRA3-7D scDb tested using Raji cells first pulsed with AMG-510 conjugated to a peptide corresponding to a fragment of KRAS (G12C) (RAS-AMG510) or a control peptide corresponding to KRAS (wild type, WTRAS). Figure 21B shows the cytotoxicity of the tested AMRA3-7D scDb against the H2122 non-small cell lung cancer cell line treated with AMG-510 or DMSO alone (negative control).

[0172] [Figures 22A-C] Cell binding and killing activity of AMRA3-7D crossMab. Figure 22A shows the binding of AMRA3-7D to Jurkat cells expressing CD3 and Raji cells not expressing CD3. Figure 22B shows the dose-dependent cell-killing activity of AMRA3-7D crossMab tested using Raji cells first pulsed with AMG-510 conjugated to a peptide corresponding to a fragment of KRAS (G12C) (RAS-AMG510) or a control peptide corresponding to KRAS (wild type, WTRAS). Figure 22C shows the cell-killing activity of AMRA3-7D crossMab tested on H2122 non-small cell lung cancer cell lines treated with AMG-510 or DMSO alone (negative control).

[0173] [Figures 23A and 23B] Deep mutational scanning of AMR-A3-7D. Figure 23A shows representative flow cytometry profiles of yeast cells displaying AMRA3-7D and its deep mutational scanning library derivatives. Figure 23B shows, in heat map format, the prevalence of mutations at each position in a sorted subset of the deep mutational scanning library.

[0174] [Figures 24A and 24B] Deep mutational scanning of OEA2-5. Figure 24A shows representative flow cytometry profiles of yeast cells displaying OEA2-5 and its deep mutational scanning library derivatives in single-chain Fv format. Figure 24B shows the prevalence of mutations at each position in a sorted subset of the deep mutational scanning library in heat map format.

[0175] [Figure 25] Binding of single-chain Fv antibody clones to the AMG510-KRAS(G12C) peptide conjugate complexed with HLA-A*11 was measured in a yeast display format. Binding intensity is expressed as median fluorescence intensity (MFI) in arbitrary units.

[0176] [Figure 26A-F] Binding titration of selected clones to the AMG510-KRAS(G12C) peptide conjugate complexed with HLA-A*03 (top row; Figures 26A-C) and HLA-A*11 (bottom row; Figures 26D-F) was tested in a yeast display format. Data using the 9-mer peptide (VVGAC*GVGK:C* indicates the Cys residue conjugated to AMG-510) are shown as filled circles, and data using the 10-mer peptide (VVVGAC*GVGK) are shown as open circles. Data for the wild-type 9-mer (VVGAGGVGK) are shown as filled triangles. Curves are best fits of a 1:1 binding model.

[0177] [Figure 27A-D] Figures 27A-C show a comparison of the binding of antibody clones to the AMG510-KRAS(G12C) peptide (9-mer) presented on HLA-A*03 (filled circles) with the binding of antibody clones to the AMG-510-KRAS(G12C) peptide in isolation, i.e., in the absence of HLA-A*03 (open squares). Figure 27D shows the binding of antibodies to the AMG510-KRAS(G12C) peptide conjugate presented on HLA in the presence and absence of 10 μM free AMG-510, demonstrating that these antibodies are not inhibited by the presence of excess concentrations of free AMG-510.

[0178] Figure 28: Cytotoxicity of scDb antibodies against lung cancer cells treated with AMG510. Luciferase-expressing NCI-H2122 cells were incubated with 1 μM AMG-510 or DMSO (vehicle) and then cocultured with human T cells (E:T = 10:1) in the presence of scDb. After incubation, target cell killing was assessed by measuring the luciferase activity released into the medium from dead cells. The affinity-matured antibodies exhibited potent cytotoxicity in a manner dependent on AMG-510 treatment. The concentration of scDb was 10 nM except for the anti-HLA-A3 scDb (positive control scDb targeting HLA-A*03), which was used at 5 nM. Data shown here are from quadruplicate assays.

[0179] [Figure 29] Cytotoxic effect of scDb antibodies on lung cancer cells treated with AMG510. The upper panel shows dose-escalation of three scDb antibodies. The lower panel shows the cytotoxic effect of 10 nM scDb compared to 5 nM control scDb. The experiment was performed in the same manner as in Figure 28.

[0180] [Figure 30] Table E shows mutation combinations that showed improved binding.

[0181] [Figure 31] Table F shows mutation combinations that showed improved binding.

[0182] [Figure 32] Binding titration of clones showing reduced affinity for the AMG510-KRAS(G12C) peptide conjugate complexed with HLA-A*03 compared to the affinity of the parent clone, AMR3-7-D, was tested in a yeast display format. The experiment was performed in the same manner as in Figure 26.

[0183] FIG. 33 shows a table showing the combinations of mutations that showed the reduced binding shown in FIG.

[0184] FIG. 34 shows a table showing the combinations of mutations that showed the reduced binding shown in FIG.

[0185] Figure 35: Biolayer interferometry (BLI) measurements of RA_D11 against AMG510-KRAS(G12C) peptide conjugates complexed with HLA-A*03 (left panel) and HLA-A*11 (right panel). Sensorgrams using the indicated concentrations of HLA complexes with the 9-mer peptide, VVGAC*GVGK (C* indicates the Cys residue conjugated to AMG-510), are shown in black in the top panel. Data for the 10-mer peptide, VVVGAC*GVGK, are shown in the bottom panel. Data using 64 nM HLA complexes with wild-type 9-mer (VVGAGGVGK) and 10-mer (VVVGAGGVGK) are shown in gray. RA_D11 in biotinylated Fab format was immobilized on a BLI chip and reacted with the HLA complexes in solution. D Values were estimated by global curve fitting of a 1:1 binding model.

[0186] [Figure 36] BLI sensorgram showing binding of RA_D11 Fab to AMG510-KRAS(G12C) peptide conjugate complexed with HLA-A*02. Sensorgram using the indicated concentrations of HLA complex with the 10-mer peptide, KLVVVGAC*GV (C* indicates a Cys residue conjugated to AMG-510). K D Values were estimated by global curve fitting of a 1:1 binding model.

[0187] [Figure 37] Specific cytotoxicity of the RA_D11 scDb antibody against KRAS(G12C)-expressing lung cancer cell lines H2122 (left panel) and SW1573 (right panel) pretreated with 1 μM AMG-510. Anti-pan HLA-A*03 scDb (A3-2) was used as a positive control. Note that it also binds to HLA-A02, albeit weaker than HLA-A*03. The experiment was performed in the same manner as in Figure 28.

[0188] [Figure 38] As expected, the AMRA3-7 and RA_D11 scDb antibodies lacked cytotoxicity against AMG510-treated Raji cells expressing wild-type KRAS. Raji cells were pretreated with 1 μM AMG-510 and stained with carboxyfluorescein succinimidyl ester (CSFE). Subsequently, the cells were cocultured with human T cells (E:T = 5:1) in the presence of the scDb for 18 hours, after which the cells were harvested, stained with Fixable Viability Dye eFluor660, and analyzed by flow cytometry.

[0189] [Figure 39] Binding titration of EO_Q16 and EO_Q17 against osimertinib-EGFR / HLA-A*02 (closed circles) and EGFR / HLA-A*02 (no drug conjugation; open circles) shows that these antibodies have high affinity for their antigens of interest, but not for the equivalent antigens without drug conjugation.

[0190] [Figure 40] Binding of EO_Q16 and EO_Q17 to osimertinib-EGFR / HLA-A*02 (10 nM) in the absence and presence of free osimertinib (1 μM). This shows that antigen binding of these antibodies is not inhibited by the presence of free drug.

[0191] [Figure 41] Cytotoxic effects of scDb antibodies against cells treated with drug-peptide conjugates. EO_Q16 and EO_Q17 scDbs exhibited potent cytotoxic effects against cells treated with osimertinib-EGFRb peptide conjugates, but not against cells treated with the same peptides without drug conjugation. BB7.2 scDb is an anti-pan HLA-A2 clone and was used as a positive control. Data are from triplicate measurements.

[0192] [Figure 42] Cytotoxicity of scDb antibodies against OCI-AML3 cells treated with osimertinib. As expected, the EO_Q16 and EO_Q17 scDb antibodies did not show cytotoxicity, since OCI-AML3 cells are not osimertinib targets. BB7.2 scDb is an anti-pan HLA-A2 clone and was used as a positive control. Data are from triplicate assays.

[0193] [Figures 43A-H] Cytotoxic effect of the RA_D11 scDb on sotorasib-treated tumor cells. Figure 43A shows the dose-response curves for the viability of H358 and H2122 cells exposed to sotorasib for 72 hours. Figure 43B shows the analysis of sotorasib conjugation to KRAS(G12C) in H2122 cells by Western blot. H2122 cells were incubated with 100 nM sotorasib for 24 hours. The arrow indicates KRAS(G12C) conjugated to sotorasib. Note that the anti-pan-RAS antibody detects KRAS, HRAS, and NRAS, so a complete shift of the original band is not expected. Figure 43C shows the cytotoxic effect of the indicated scDbs on H2122-Nluc cells treated with 1 μM sotorasib. The fold change in luminescence signal intensity is shown normalized to that from the control without scDb (i.e., H2122 and T cells only). The scDb concentration was 10 nM except for the A3-2 scDb (1 nM). DMSO was the sotorasib vehicle. Figure 43D shows the cell-killing titration curve of RA_D11 scDb against H2122-Nluc cells treated with 1 μM sotorasib. Figure 43E shows the dependence of cell killing on sotorasib concentration (the indicated scDb was used at 1 nM). Figure 43F shows the HLA-dependence of cell killing by RA_D11 scDb. Normalized luminescence intensity is shown for cell lines treated with 0.3 μM sotorasib and cocultured with T cells in the presence of 1 nM scDb and 0.3 μM sotorasib. The KRAS mutation status and HLA alleles of the cell lines are indicated. Because each cell line expresses different levels of Nluc, assays were performed separately to determine the relative luciferase levels for each cell line. Figures 43G and 43H show the cytotoxic effect of RA_D11 scDb (1 nM) on H2030-Nluc (G) and SW1573-Nluc (H) cells treated with sotorasib. Data are from quadruplicate determinations.

[0194] [Figures 44A and 44B] HLA expression of cell lines. Figure 44A shows the analysis of HLA-A*03 expression on Raji cells. Figure 44B shows the analysis of HLA-A*03 expression on H2122 cells and H2122(HLA-A*03KO) cells. The latter lack HLA-A*03, indicating successful deletion.

[0195] [Figures 45A-D] Cytotoxic effects of RA_D11 on tumor cells treated with sotorasib. Figure 45A shows the cytotoxic effects of the indicated scDbs on H2122-Nluc cells treated with 1 mM sotorasib. The scDb concentration was 10 nM except for the positive control A3-2 scDb (1 nM). Figure 45B shows the cytotoxic effects of the indicated scDbs on various NLuc-expressing tumor cell lines treated with 0.3 μM sotorasib. The scDb concentration was 1 nM. Figures 45C and 45D show the cytotoxic effects of RA_D11 scDb on H2030-Nluc cells (Figure 45C) and SW1573-Nluc cells (Figure 45D) treated with sotorasib. The scDb concentration was 1 nM. Positive controls included anti-pan HLA-A*03 / A*02 antibody (A3-2 scDb), anti-pan HLA-A*02 antibody (BB7.2 scDb), and anti-pan HLA-A*11 antibody (A11-1). Data are from quadruplicate determinations.

[0196] [Figure 46] Binding analysis of antibody phage clones to peptide / MHC complexes or sotorasib / peptide / MHC complexes by multiplex bead binding assay (MBBA). Phage MBBA signals indicate the binding specificity of antibody clones (A3-2 and A11-1) that is not specific for the identification of the bound peptide.

[0197] [Figure 47A-C] Structures of the soto-pMHC-RA_D11 complex (side and top views). Figure 47A shows the side-view (upper panel) and top-view (lower panel) structures of soto-p8 / A03-RA_D11. Figure 47B shows the side-view (upper panel) and top-view (lower panel) structures of soto-p7 / A03-RA_D11. Figure 47C shows the side-view (upper panel) and top-view (lower panel) structures of soto-p7 / A11-RA_D11. Proteins and peptides are represented as tubes, and sotolacib is represented as sticks. 4701 indicates the relevant region of RA_D11 VH, 4702 indicates the relevant region of RA_D11 VL, 4703 indicates the relevant region of β2m, 4704 indicates the relevant region of HLA, and 4705 indicates the relevant position of the drug-peptide conjugate soto-p. The angle between the axes of Fab RA_D11 and soto-pMHC is shown in the soto-p8 / A03 structure (Figure 47A).

[0198] [Figure 48] Residue surface area buried by the interaction of Fab RA_D11 with soto-pMHC, calculated with PDBePISA. Shades of gray indicate varying fractions of the solvent-accessible residue surface that are buried by the interaction of the indicated molecules (darker gray indicates a higher fraction of buried surface area [BSA: Buried Surface Area]), as shown in the scale bar on the right.

[0199] [Figure 49A-E] Details of the interaction between Fab RA_D11 and soto-p8 / A03. Figure 49A shows soto-p8 (stick; e.g., V8V9G) complexed with Fab RA_D11 (not shown). 10 A 11 C 12 G 13 V 14 G 15 K 16-Figure 49B shows a tube representation of HLA-A*03:01 (tube) presenting Fab RA_D11 (sotorasib). HLA-A*03:01 residues in contact with either Fab RA_D11 or sotorasib are represented as sticks. Figure 49B shows the contact areas between the HLA molecule and Fab RA_D11. The contact areas of Fab RA_D11 (4901 indicates the approximate area of contact with VL, 4902 indicates the approximate area of contact with VH) or sotorasib (4903 indicates the approximate area of contact with sotorasib), or both (4904), depicted on the surface of HLA-A*03:01, are labeled. Figure 49C shows a tube representation of Fab RA_D11 complexed with soto-p8 / A03 (showing only sotorasib; sticks). Fab RA_D11 residues in contact with either HLA-A*03:01 or sotorasib are represented as sticks. Figure 49D shows the contact areas of HLA-A*03:01 (4905), sotorasib (4906), or both (4907, the area surrounding sotorasib) depicted on the surface of Fab RA_D11. 27 , S 29 , S 30 , S 31 , Y 93 ) and HLA-A*03:01 residues (e.g., R 108 , D 161 , E 166 , W 167 , R 168 ) are shown in detail. Residues involved in H-bonds are shown as sticks, and H-bonds are shown as dashed lines.

[0200] [Figure 50A-E] Interaction details of Fab RA_D11 binding to soto-p7 / A03. Figure 50A shows soto-p7 (stick; e.g., V7V8V9G) complexed with Fab RA_D11 (not shown). 10 A 11 C 12 G 13 V 14 G 15 K 16Figure 50B shows the contact areas of Fab RA_D11 (5001 indicates the approximate region contacting the VH; 5002 indicates the VL; 5003 indicates the VH and VL), sotorasib (5004), or both (5005) depicted on the surface of HLA-A*03:01. Figure 50C shows a tube representation of Fab RA_D11 complexed with soto-p7 / A03 (showing only sotorasib; sticks). Fab RA_D11 residues contacting either HLA-A*03:01 or sotorasib are represented as sticks. Figure 50D shows the contact regions of HLA-A*03:01 (5006), sotorasib (5007), or both (5008) depicted on the surface of Fab RA_D11. Figure 50E shows the contact regions of Fab RA_D11 VH (e.g., S33) and RA_D11 VL (e.g., Y 93 , S 92 , S 28 , S 30 ) and HLA-A*03:01 (e.g., E 166 , W 167 , R 108 ) residues are shown in detail. Residues involved in H-bonds are shown as stick figures, and H-bonds are shown as dashed black lines.

[0201] [Figure 51A-F] Details of the interaction of Fab RA_D11 binding to soto-p7 / A11. Figure 51A shows the interaction of soto-p7 (stick; e.g., V8V9G) complexed with Fab RA_D11 (not shown). 10 A 11 C 12 G 13 V 14 G 15 K 16Figure 51B shows the contact areas of Fab RA_D11 (5101 indicates the approximate region of contact with VH; 5102 indicates VL; 5103 indicates VH and VL), sotorasib (5104), or both (5105) depicted on the surface of HLA-A*11:01. Figure 51C shows the tube representation of Fab RA_D11 complexed with soto-p7 / A11 (showing only sotorasib; sticks). Fab RA_D11 residues in contact with either HLA-A*11:01 or sotorasib are represented as sticks. Figure 51D shows the contact regions of HLA-A*11:01 (5106), sotorasib (5107), or both (5108) depicted on the surface of Fab RA_D11. Figures 51E and 51F show the contact regions of Fab RA_D11 VH (e.g., G 100 , A 103 ) and RA_D11 VL (e.g., K 96 , S 92 , R 95 , Q 27 , S 28 , G 68 , R 66 ) and HLA-A*11:01 (e.g., E 166 , R 169 , R 108 , D 106 ) residues are shown in detail. Residues involved in H-bonds are shown as stick figures, and H-bonds are shown as dashed black lines.

[0202] [Figure 52A-F] Comparison of different Fab RA_D11-soto-pMHC structures. Figures 52A and 52B show the structural superposition of soto-p8 / A03 and soto-p7 / A03. HLA-A*03:01 is represented as a tube. Side chains of HLA residues involved in RA_D11 interaction are represented as sticks. Figure 52B shows a detailed comparison of soto-p8 (dark) and soto-p7 (light) when loaded with HLA-A*03:01 (not shown) and bound to Fab RA_D11 (not shown). Figures 52C-F show the structural superposition of soto-p7 / A03 and soto-p7 / A11. soto-p7 is represented as a stick (light), and HLA-A*03:01 (light) and HLA-A*11:01 (dark) are represented as tubes. Side chains of HLA residues that differ between HLA-A*03 and A*11 are represented as sticks. Figure 52D shows a detailed comparison of soto-p7 peptides presented by HLA-A*03:01 (light sticks, HLA not shown) or HLA-A*11:01 (dark sticks, HLA not shown) and bound to Fab RA_D11 (not shown). Figure 52E shows a detailed comparison of soto-p7 presentation caused by the presence of T163 / R163 in HLA-A*03 / A*11. Figure 52F shows a detailed comparison of the different conformations of Fab RA_D11 CDRH3 when bound to soto-p7 / A03 or soto-p7 / A11.

[0203] [Figure 53] Flow cytometry profiles of binding of RA_D11 (left column) to drug-peptide / MHC complexes and to sorted deep mutational scanning libraries. Condition 1: 10 nM soto-p7 / A03; Condition 2: 10 nM soto-p7 / A11; Condition 3: 10 nM soto-p5 / A02 tetramer (on streptavidin-DyLight650).

[0204] [Figure 54] Heatmap representation of single point mutants in the VL of RA_D11 that retain antigen binding (left column) or that abolish binding (right column). Numbers represent the number of reads for the mutation of interest divided by the total number of reads at that position multiplied by 100. Condition 1: 10 nM soto-p7 / A03, Condition 2: 10 nM soto-p7 / A11, Condition 3: 10 nM soto-p5 / A02 tetramer (on streptavidin-DyLight650).

[0205] [Figure 55] Heatmap representation of single point mutants in the VH of RA_D11 that retain antigen binding (left column) or that abolish binding (right column). Numbers represent the number of reads for the mutation of interest divided by the total number of reads at that position, multiplied by 100. Condition 1: 10 nM soto-p7 / A03; Condition 2: 10 nM soto-p7 / A11; Condition 3: 10 nM soto-p5 / A02 tetramer (on streptavidin-DyLight650).

[0206] [Figure 56] Fab RA_D11 can specifically bind to all four soto-pMHC targets with high affinity, with minimal inhibition by free inhibitors. The minimal inhibition of RA_D11 binding by free sotorasib may be a fundamental requirement for co-administration with inhibitors.

[0207] [Figure 57A-C] CryoEM structures of RA_D11 bound to multiple drug-peptide / MHC complexes reveal the molecular basis of target recognition. RA_D11 similarly recognizes the soto-p / A03-A11 target, interacting primarily with the HLA surface and sotorasib (located in the pocket formed by the interface of the Fab variable chain), with minimal contact with the remainder of the peptide (p7 or p8).

[0208] [Figures 58A and 58B] RA_D11 binds to soto-pMHC in a non-canonical manner, distinct from typical TCR-pMHC engagement.

[0209] [Figure 59A-C] RA_D11 recognizes both soto-p7 / A03 and soto-p8 / A03 in almost the same way, because the peptide accommodation in the HLA-A*03 pocket is different for soto-p8 (stretch) and soto-p7 (bend). When the soto-p7 peptide is loaded into HLA-A*03 or A*11, the conformation of the Fab CDR (especially CDR-H3) during target binding is slightly different due to the difference in residues between the two HLAs.

[0210] [Figures 60A and 60B] RA_D11 can also bind to soto-p5 / A02, and therefore, RA_D11-based specific T cell engagers can bind to the most prevalent HLA-A*02-bearing KRAS (G12C) It can also selectively kill cancer cells. RA_D11 binds to soto-p5 / A02 in a non-unique manner, exhibiting multiple possible orientations and reduced surface contraction with HLA for soto-p / A03-A11 target binding.

[0211] [Figure 61] soto-p5 is soto-p 7-8 are very different peptides and should be presented in a different way.

[0212] [Figure 62] Establishment of a grid on the MHC backbone. Grid angles were set using the following α-carbons from the amino acids of the MHC complex: the origin set was Met98, the X axis was set using Val28, and the Y axis was set using Glu128, with the Y axis orthogonal. Incident angles were measured along the XY axis, and back angles were measured along the YZ axis.

[0213] [Figure 63] Binding angle between soto-p7 / A03 and RA_D11 antibody. The angle was measured from the axis to the center of the RA_D11 antibody (Met105 sulfur). The back angle obtained from the YZ axis was measured to be 64.7°. The entrance angle obtained from the XY axis was measured to be 77.3°.

[0214] [Figure 64] Binding angle between soto-p7 / All and RA_D11 antibody. The angle was measured from the axis to the center of the RA_D11 antibody (Met105 sulfur). The back angle obtained from the YZ axis was measured to be 67.2°. The incident angle obtained from the XY axis was measured to be 78.4°.

[0215] [Figure 65] Binding angle between soto-p8 / A03 and RA_D11 antibody. The angle was measured from the axis to the center of the RA_D11 antibody (Met105 sulfur). The back angle obtained from the YZ axis was measured to be 65.1°. The entrance angle obtained from the XY axis was measured to be 77.2°.

[0216] [Figure 66] Binding angles of a TCR-like antibody from PDB 3CHV. The angle was measured from the axis to the center of the TCR-like antibody (Phe107 C4). The back angle taken from the YZ axis was measured to be 92.5°. The entrance angle taken from the XY axis was measured to be 111.1°. Detailed Description

[0217] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs.

[0218] Every numerical range given throughout this specification includes its upper and lower limits, and every narrower numerical range that falls within that range, as if such narrower numerical ranges were all expressly written herein.

[0219] As used in this specification and the appended claims, the singular forms "a," "and," and "the" include plural references unless the context clearly dictates otherwise. Ranges may be expressed herein as from "about" one particular value and / or to "about" another particular value. When such a range is expressed, another embodiment includes from the one particular value and / or to the other particular value. Similarly, when values are expressed as approximations, it is understood that the particular value forms another embodiment by use of the antecedent "about." The term "about" with respect to numerical values encompasses variations of ±10%, ±5%, or ±1%.

[0220] The present disclosure includes all amino acid sequences described herein and all nucleotide sequences encoding said amino acid sequences. All antibody sequences and antigen-binding fragments thereof are included. Polynucleotide and amino acid sequences with 80-99% similarity (including upper and lower limits, and all numbers and ranges therebetween) to the sequences provided herein are encompassed by the present invention. All amino acid sequences described herein may contain amino acid substitutions (e.g., conservative substitutions) that do not adversely affect the function of the protein containing the amino acid sequence. In this regard, the present disclosure provides alternative residues for certain positions in the described binding partners, as shown below. In certain instances, the alternative residues were identified by deep mutational scanning, which demonstrated the binding functionality of each binding partner containing the described amino acid changes. The present disclosure includes each binding partner in which only the original residue is replaced with each alternative residue, as well as each binding partner with any combination of the described alternative residues. Thus, the binding partners described herein can have any single described residue change or a combination of the described changes. Representative changes for specific antibodies are shown in the table. The alterations may be in CDR1, CDR2, CDR3, or a combination thereof. Alterations may also include amino acid insertions. The present disclosure includes each amino acid sequence encompassed by the description of the specific sequence identifiers and alternative amino acids by reference to those listed in the preceding table.

[0221] As described above, the present disclosure provides antibodies and antigen-binding fragments thereof (collectively, "binding partners" and individually, "binding partners"). The term "antibody" encompasses each form of binding partner herein. A binding partner specifically binds to a protein or fragment thereof, including a covalently attached molecule, or a peptide provided in peptide form. The covalently attached molecule forms a peptide conjugate. As used herein, "peptide conjugate" refers to any protein or peptide that has been modified to be covalently conjugated to another molecule. Peptide conjugates are considered novel antigens, or neoantigens. The other molecule covalently conjugated to a protein or peptide to form the peptide conjugate is not particularly limited, except that the other molecule is not an additional amino acid added to the described peptide conjugate. In embodiments, the molecule covalently conjugated to a protein or peptide has or may have biological activity prior to conjugation, or may be biologically inactive prior to conjugation. In embodiments, the molecule is a drug, including, but not necessarily limited to, a small molecule drug. As used herein, a molecule that is covalently attached to a peptide to form a peptide conjugate is referred to as a "targeted covalent inhibitor (TCI)" or "covalent drug." In some cases, a targeted covalent inhibitor may be a molecule that inhibits the activity of a target protein and may be covalently attached to a peptide derived from the target protein. In some cases, a targeted covalent inhibitor may not inhibit the activity of the target protein but may be covalently attached to a peptide derived from the target protein. Representative, non-limiting examples of drugs that are covalently attached to a peptide or protein to form a peptide conjugate are described below. Peptide conjugates include, but are not limited to, covalently modified full-length proteins and fragments thereof.Peptide conjugates include fragments of full-length proteins containing covalent modifications, for example, produced by intracellular processing. In certain embodiments, full-length proteins can be covalently modified intracellularly and then processed to produce peptide conjugates that are fragments of the full-length protein. In certain embodiments, peptide conjugates comprise fragments of full-length proteins. As described further below, the produced peptide conjugates can be displayed on the cell surface. Cell surface display of peptide conjugates can be any form of cell surface display, including, but not limited to, by any receptor with an extracellular segment, or by any type of major histocompatibility complex (MHC) or human leukocyte antigen (HLA). Non-limiting examples of HLA types that display peptide conjugates and to which the described binding partners specifically bind are further described below.

[0222] As used herein, the term "peptide conjugate / MHC complex" refers to a peptide conjugate comprising a peptide and a chemical fragment of a target covalent inhibitor, which is presented by the major histocompatibility complex (MHC). For example, the peptide conjugate can be formed by a covalent reaction between the target covalent inhibitor and a residue (e.g., a cysteine residue) in the peptide. In some embodiments, the peptide conjugate is formed by a covalent reaction between AMG-510 and a KRASG12C peptide. In some embodiments, the peptide is exogenously introduced as a vaccine. In some embodiments, the peptide comprises a nucleophilic or electrophilic residue. In some embodiments, the residue comprises cysteine, lysine, tyrosine, histidine, serine, arginine, or threonine. In one embodiment, the MHC is a human leukocyte antigen (HLA). In embodiments, the HLA is HLA-A*01:01, HLA-A*02:01, HLA-A*03:01, HLA-A*11:01, HLA-A*24:02, HLA-A*26:01, HLA-B*07:02, HLA-B*08:01, HLA-B*27:05, HLA-B*39:01, HLA-B*40:01, HLA-B*58:01, or HLA-B*15:01.

[0223] As used herein, the term "CDR" or "complementarity determining region" refers to the non-contiguous antigen-binding sites found within the variable regions of heavy and light chain polypeptides. These particular regions are described, for example, in Kabat et al., J. Biol. Chem. 252, 6609-6616 (1977); Kabat et al., Sequences of proteins of immunological interest (1991); Chothia et al., J. Mol. Biol. 196:901-917 (1987); and MacCallum et al., J. Mol. Biol. 262:732-745 (1996), all of which are incorporated herein by reference in their entireties, where the definitions include overlapping or subsets of amino acid residues when compared against each other. In certain embodiments, the term "CDR" refers to a CDR as defined by MacCallum et al., J. Mol. Biol. 262:732-745 (1996) and Martin A. "Protein Sequence and Structure Analysis of Antibody Variable Domains," in Antibody Engineering, Kontermann and Dubel, eds., Chapter 31, pp. 422-439, Springer-Verlag, Berlin (2001). In certain embodiments, the term "CDR" refers to a CDR as defined by Kabat et al., J. Biol. Chem. 252, 6609-6616 (1977) and Kabat et al., Sequences of proteins of immunological interest (1991). In certain embodiments, the heavy chain CDRs and light chain CDRs of an antibody are defined using different conventions. In certain embodiments, the heavy and / or light chain CDRs are defined by performing a structural analysis of the antibody and identifying variable region residues that are predicted to contact the epitope region of the target molecule (e.g., a peptide conjugate).CDR-H1, CDR-H2, and CDR-H3 represent heavy chain CDRs, and CDR-L1, CDR-L2, and CDR-L3 represent light chain CDRs.

[0224] The determination of "percent identity" between two sequences (e.g., amino acid sequences or nucleic acid sequences) can be accomplished using a mathematical algorithm. A specific, non-limiting example of a mathematical algorithm utilized for comparing two sequences is the algorithm of Karlin S & Altschul SF (1990) PNAS 87: 2264-2268, modified as in Karlin S & Altschul SF (1993) PNAS 90: 5873-5877, each of which is incorporated herein by reference in its entirety. Such an algorithm is incorporated into the NBLAST and XBLAST programs of Altschul SF et al., (1990) J Mol Biol 215: 403, which is incorporated herein by reference in its entirety. BLAST nucleotide searches can be performed, for example, using the NBLAST nucleotide program parameters set to score=100 and wordlength=12 to obtain nucleotide sequences homologous to the nucleic acid molecules described herein. BLAST protein searches can be performed using the XBLAST program parameters, for example, set to score 50 and wordlength=3, to obtain amino acid sequences homologous to the protein molecules described herein. To obtain gapped alignments for comparison purposes, Gapped BLAST can be utilized as described in Altschul SF et al. (1997) Nuc Acids Res 25:3389-3402, which is incorporated herein by reference in its entirety. Alternatively, PSI BLAST can be used to perform an iterated search that detects distant relationships between molecules (Id.). When utilizing BLAST, Gapped BLAST, and PSI Blast programs, the default parameters of the respective programs (e.g., XBLAST and NBLAST) can be used (see, for example, the National Center for Biotechnology Information (NCBI) on the worldwide web at ncbi.nlm.nih.gov).Another specific, non-limiting example of a mathematical algorithm utilized for comparing sequences is the algorithm of Myers and Miller, 1988, CABIOS 4:11-17, which is incorporated herein by reference in its entirety. Such an algorithm is incorporated into the ALIGN program (version 2.0), which is part of the GCG sequence alignment software package. When utilizing the ALIGN program to compare amino acid sequences, a PAM120 weight residue table, a gap length penalty of 12, and a gap penalty of 4 can be used. The percent identity between two sequences can be determined using techniques similar to those described above, with or without allowing gaps. In calculating percent identity, typically, only exact matches are counted.

[0225] As used herein, the term "free target covalent inhibitor" or "free drug" refers to a target covalent inhibitor that is not covalently bound to a protein or peptide. Once a target covalent inhibitor is covalently bound to a protein or peptide, the target covalent inhibitor may be referred to as a portion or fragment of the free target covalent inhibitor or drug. For example, a protein or fragment thereof may be a chemical fragment that binds to a cysteine residue of a peptide by a covalent reaction of the free drug with the cysteine residue of the peptide. As used herein, "KRAS G12C The term "KRAS" refers to a KRAS protein having a G12C mutation, i.e., a cysteine at amino acid position 12 (UniProt Accession No. P01116). G12D " refers to a KRAS protein with a G12D mutation, i.e., aspartic acid at amino acid position 12 (UniProt Accession No. P01116). As used herein, the term "KRAS G12R " refers to a KRAS protein with a G12R mutation, i.e., arginine at amino acid position 12 (UniProt Accession No. P01116). As used herein, the term "KRAS G12S" refers to a KRAS protein with a G12S mutation, i.e., a serine at amino acid position 12 (UniProt Accession No. P01116).

[0226] In some embodiments, the binding partner preferentially binds to a protein or peptide covalently attached to a peptide conjugate, or to a complex containing the protein or peptide, compared to the same protein or peptide not conjugated to a drug. Thus, the binding partners described herein do not detectably bind, or bind with lower affinity, to the same protein or fragment thereof in the absence of the covalently attached molecule. In some embodiments, the binding partner binds to a protein or peptide containing a covalently attached drug with an affinity that is 10-10,000 times (including all numbers and ranges between 10 and 10,000) greater than the affinity for the protein or peptide without the covalently attached molecule. While not intending to be bound by any particular theory in this regard, it is believed that the presence of the covalently attached molecule contributes to the epitope to which the binding partner specifically binds. Similarly, the binding partners of the present disclosure preferentially bind to a peptide conjugate compared to binding to the free drug. In embodiments, the binding partner binds to a peptide (e.g., a peptide conjugate / MHC complex) containing a covalently attached drug with an affinity that is 10-10,000 times (including all numbers and ranges between 10-10,000) greater than the affinity for the free drug. In some embodiments, the interaction between the binding partner and the peptide conjugate / MHC complex is not inhibited by the free drug. For example, the interaction between the binding partner and the peptide conjugate / MHC complex is not inhibited by a 100-fold, 1,000-fold, 10,000-fold, 100,000-fold, or greater excess of free drug.

[0227] In some embodiments, the molecule covalently bonded to form the peptide conjugate is a drug, which may be any target covalent inhibitor (TCI), although the covalent drug need not necessarily inhibit the target peptide. In some embodiments, the molecule reacts with a specific residue within the target protein. In some embodiments, the molecule reacts at least partially with a protein or peptide segment containing a nucleophilic or electrophilic residue. In some embodiments, the protein or peptide segment with which the molecule reacts contains any of Cys, Lys, Tyr, His, Ser, Thr, or Arg, the latter of which is described in "Ziyang Zhang, Johannes Morstein, Andrew K. Ecker, Keelan Z. Guiley, and Kevan M. Shokat Journal of the American Chemical Society Article ASAP, DOI: 10.1021 / jacs.2c05377," the disclosure of which is incorporated herein by reference. In some embodiments, the protein or peptide contains selenocysteine. In some embodiments, the targeted covalent inhibitor reacts with selenocysteine, hi some embodiments, the molecule reacts at least partially with a segment of a protein or peptide containing wild-type Cys or a mutation of the residue to Cys, and thus can be covalently attached via a so-called sulfur tether.In embodiments, the drug is any drug described in Ghosh AK, Samanta I, Mondal A, Liu WR. Covalent Inhibition in Drug Discovery. ChemMedChem. 2019;14(9):889-906. doi:10.1002 / cmdc.201900107 or De Cesco, et al., European Journal of Medicinal Chemistry 138 (2017) 96e114 or Bauer, RA, Drug Discovery Today, Volume 20, Number 9, September 2015, the disclosures of compounds that covalently modify protein targets described therein are incorporated herein by reference.

[0228] In non-limiting embodiments, any of the Cys, Lys, Tyr, Ser, Thr, Arg, and His amino acids are present in the protein or peptide to which the molecule binds (because the gene encoding the wild-type protein has been mutated to encode a protein containing one or a combination of the listed residues). In non-limiting embodiments, the molecule binds to a protein or peptide that correlates with a disease or condition (such as cancer, autoimmune disease, or other disease or disorder treated with a targeted covalent inhibitor). In some embodiments, the target (e.g., the protein or peptide to which the molecule covalently binds) is a receptor, including, but not necessarily limited to, any receptor with a catalytically active segment. In some embodiments, the drug binds to an enzyme (not necessarily a receptor, including, but not limited to, a kinase). In some embodiments, the protein target includes a receptor with one or more activating mutations that promote ligand-independent enzymatic activity.

[0229] In some embodiments, the molecule targets and thus covalently binds to an amino acid sequence present in any of the following proteins and / or variants thereof (which may or may not contain mutations, e.g., mutations associated with a particular condition, including, but not limited to, any type of cancer): In some embodiments, the protein is any protein described in "Visscher M, et al., Covalent targeting of acquired cysteines in cancer. Curr Opin Chem Biol. 2016;30:61-67. doi:10.1016 / j.cbpa.2015.11.004," which is incorporated herein by reference. Visscher et al. also teach a method for identifying disease-associated mutant genes that introduce a Cys residue suitable for covalent modification. In embodiments, the protein is KRAS, Bruton's tyrosine kinase (BTK); any member of the epidermal growth factor receptor (EGFR) family, also known as the ERBB family, including but not limited to EGFR (ERBB1), HER2 / NEU (ERBB2), HER3 (ERBB3), and HER4 (ERBB4); fibroblast growth factor receptor (FGFR); MET, BRAF, receptor kinases known in the art as cyclin-dependent kinases (CDKs); acetylcholinesterase (ACHE); TP53, IDH1, GNAS, FBXW7, CTNNB1, DNMT3A, any cathepsin (including cathepsins B, C, F, H, K, L, O, S, V, W, and X); any caspase; any protein involved in obesity (e.g., pancreatic lipase and METAP2), or any cancer-testis antigen. In embodiments, the drug targets and thus covalently binds to any viral protein (including, but not limited to, any viral DNA polymerase, RNA polymerase, reverse transcriptase, or polymerase, including RNA-dependent RNA polymerase, or a viral protein required for, for example, viral cell entry, or a protein encoded by any transposable element).In some embodiments, the drug targets EGFR and may be selected from PD168393, PF00299804 (dacomitinib), EKB569 (pelitinib), afatinib, WZ4002, osimertinib (also known as AZD9291), PF-06459988, nazartinib, naquotinib, olmutinib, avitinib, and rociletinib, neratinib, pyrotinib, poziotinib, and derivatives thereof. In some embodiments, the drug targets Bruton's tyrosine kinase (BTK) and may be selected from ibrutinib, acalabrutinib, zanubrutinib, CHMFL-BTK-11, ONO / GS-405, PRN1008, and CC-292. In some embodiments, the drug targets any p90 ribosomal S6 kinase (RSK) and may be selected from fluoromethyl ketone (FMK) and dimethyl fumarate. In some embodiments, the drug targets any FGFR and may be selected from FIIN-1, FIIN-2, FIIN-3, BGJ398, AZD4547, PRN1371, and FGF401. In some embodiments, the targeted covalent inhibitor targets an E3 ligase such as RNF4, HOIP, RSP5, SMURF1, E6AP, HUWE1, and NEDD4-1. In some embodiments, the targeted covalent inhibitor targets a DDB1- and CUL4-associated factor (DCAF), such as DCAF1 or DCAF15. In some embodiments, the targeted covalent inhibitor targets any cancer-testis antigen, any endogenous retroviral protein, long interspersed nuclear element-1 (LINE-1), or short interspersed nuclear element (SINE). In some embodiments, the targeted covalent inhibitor targets a short interspersed nuclear element, which is optionally Alu.In certain embodiments, the targeted covalent inhibitor is iniparib, abiraterone, carfilzomib, afatinib, or neratinib.

[0230] In some embodiments, the molecule covalently linked to form the peptide conjugate targets any RAS oncogene protein product, including, but not limited to, HRAS, NRAS, KRAS4A, and KRAS4B. The amino acid sequences of RAS proteins are known in the art, and the residue numbering is the same for the relevant portions of all RAS isotypes discussed in this disclosure, which amino acid sequences are available, for example, from UniProt P01116, which amino acid sequence is incorporated herein as of the effective filing date of this application or patent. The G12 position is numbered according to the known amino acid sequence, regardless of whether G12 is the 12th amino acid in the expressed RAS peptide sequence of this disclosure.

[0231] In one embodiment, the molecule covalently binds to a KRAS protein or peptide that contains a mutation. In embodiments, the mutation is at least one of KRAS residues 12, 13, or 61. References herein to drugs include their names, regardless of case.

[0232] In some embodiments, the drug inhibits KRAS G12C In some embodiments, the drug targets the KRAS protein containing the mutation. G12D In some embodiments, the drug targets the KRAS protein containing the mutation. G12R In some embodiments, the drug targets the KRAS protein containing the mutation. G12STargeting KRAS protein that contains mutation.In a non-limiting embodiment, the drug that targets KRAS protein is selected from AMG-510 (sotorasib), ARS-853, ARS-1620, ARS-3248, MRTX849, JNJ74699157, LY3499446, LY3537982, MRTX-1257, JDQ443, MRTX-1133, RMC-6291, RMC-9805, GDC-6036, D-1553, 2E07, 6H05, SML-8-73-1 or BI 182391, or their derivatives. In other embodiments, the drug targeting the KRAS protein is selected from β-lactones G12Si-1, G12Si-2, G12Si-3, G12Si-4, and G12Si-5. In certain embodiments, the drug comprises a covalent drug-targeted proteolysis chimera (PROTAC) derivative, a non-limiting description of which is available at doi: 10.1021 / acscentsci.0c00411, which description of PROTACs is incorporated herein by reference. In some embodiments, the PROTAC is LC-1 or LC-2. In some embodiments, the present disclosure relates to autophagy-mediated degraders, referred to as AUTACs, as described in doi.org / 10.1080 / 15548627.2020.1718362, which description of AUTACs is incorporated herein by reference.

[0233] The peptide conjugates described herein can be formed by the covalent reaction of a free target covalent inhibitor with a KRAS peptide. G12C Peptides, KRAS G12D Peptides, KRAS G12R peptide, or KRAS G12SThe free covalent target inhibitor can be formed by covalent reaction with.The free covalent target inhibitor can be any free covalent target inhibitor described herein.For example, the free covalent target inhibitor can be osimertinib, ibrutinib, neratinib, AMG-510, ARS-853, ARS-1620, ARS-3248, MRTX849, JNJ74699157, LY3499446, LY3537982, MRTX-1257, JDQ443, MRTX-1133, RMC-6291, RMC-9805, GDC-6036, D-1553, 2E07, 6H05, SML-8-73-1 or BI 182391, or derivatives thereof. In other embodiments, the free target covalent inhibitor can be β-lactone G12Si-1, G12Si-2, G12Si-3, G12Si-4, or G12Si-5 (see Table R). In some examples, the peptide conjugate is a conjugate of AMG-510 and KRAS G12C It may be formed by a covalent bond reaction with a peptide, which may comprise or consist of the amino acid sequence VVVGACGVGK, VVGACGVGK or KLVVVGACGV.

[0234] The antigen-binding domain of a polypeptide described herein or a binding partner described herein can bind to a peptide conjugate / MHC complex presented by different HLAs. For example, in certain embodiments, the antigen-binding domain can have specificity for: (i) a peptide conjugate / MHC complex comprising VVVGACGVGK conjugated to a targeted covalent inhibitor or fragment thereof presented by HLA-A*11:01, (ii) a peptide conjugate / MHC complex comprising VVVGACGVGK conjugated to a targeted covalent inhibitor or fragment thereof presented by HLA-A*03:01, or (iii) both. In certain other embodiments, the antigen-binding domain may have specificity for: (i) a peptide conjugate / MHC complex comprising VVGACGVGK conjugated to a targeted covalent inhibitor or fragment thereof presented by HLA-A*03:01; (ii) a peptide conjugate / MHC complex comprising VVVGACGVGK conjugated to a targeted covalent inhibitor or fragment thereof presented by HLA-A*03:01; or (iii) both.In other embodiments, the antigen-binding domain may have specificity for: (i) a peptide conjugate / MHC complex comprising VVVGACGVGK conjugated to a targeted covalent inhibitor or fragment thereof presented by HLA-A*11:01; (ii) a peptide conjugate / MHC complex comprising VVGACGVGK conjugated to a targeted covalent inhibitor or fragment thereof presented by HLA-A*11:01; (iii) a peptide conjugate / MHC complex comprising VVGACGVGK conjugated to a targeted covalent inhibitor or fragment thereof presented by HLA-A*03:01; (iv) a peptide conjugate / MHC complex comprising VVVGACGVGK conjugated to a targeted covalent inhibitor or fragment thereof presented by HLA-A*03:01; (v) a peptide conjugate / MHC complex comprising KLVVVGACGV conjugated to a targeted covalent inhibitor or fragment thereof presented by HLA-A*02:01; or (vi) any combination of (i)-(v) (e.g., (i) and (ii); or (i), (ii) and (iv)), or (vii) all of (i)-(v). In certain specific embodiments, the polypeptide binds to: (i) a peptide conjugate / HLA-A*11:01 MHC complex comprising a peptide consisting of the amino acid sequence VVGACGVGK, (ii) a peptide conjugate / HLA-A*11:01 MHC complex comprising a peptide consisting of the amino acid sequence VVVGACGVGK, (iii) a peptide conjugate / HLA-A*03:01 MHC complex comprising a peptide consisting of the amino acid sequence VVVGACGVGK, (iv) a peptide conjugate / HLA-A*03:01 MHC complex comprising a peptide consisting of the amino acid sequence VVVGACGVGK, and / or (v) a peptide conjugate / HLA-A*02:01 MHC complex comprising a peptide consisting of the amino acid sequence KLVVVGACGV.In still other embodiments, the polypeptide binds to: (i) a peptide conjugate / HLA-A*03:01 MHC complex comprising a peptide consisting of the amino acid sequence VVVGACGVGK, and a peptide conjugate / HLA-A*11:01 MHC complex comprising a peptide consisting of the amino acid sequence VVVGACGVGK; (ii) a peptide conjugate / HLA-A*03:01 MHC complex comprising a peptide consisting of the amino acid sequence VVVGACGVGK, and a peptide conjugate / HLA-A*03:01 MHC complex comprising a peptide consisting of the amino acid sequence VVVGACGVGK; (iii) a peptide conjugate / HLA-A*02:01 MHC complex comprising a peptide consisting of the amino acid sequence KLVVVGACGV; or any combination of (i) to (iii) (e.g., (i) and (ii); or (i) and (iii)); or all of (i) to (iii). Covalent inhibitors that target peptide conjugate / MHC complexes can have a chemical structure comprising C-R1, where C is a chemical fragment linked to R1 of the compounds exemplified below. [ka] R1 in the C-R1 chemical structure of the covalent inhibitor can be any of the structures exemplified below. [ka]

[0235] A targeted covalent inhibitor having a structure including C-R1 can form a covalent bond to multiple different amino acid residues on a peptide. For example, a targeted covalent inhibitor can form a covalent bond with a cysteine residue in a peptide having the amino acid sequence VVVGACGVGK, VVGACGVGK, or KLVVVGACGV. In other embodiments, a targeted covalent inhibitor can form a covalent bond with an aspartic acid residue, a serine residue, or an arginine residue in a peptide having the amino acid sequence VVVGADGVGK, VVGADGVGK, or KLVVVGADGV. In some embodiments, the antigen-binding domain of the polypeptide recognizes the C portion of a targeted covalent inhibitor having a structure including C-R1 of a peptide conjugate / MHC complex. In certain other embodiments, the antigen-binding domain of the polypeptide recognizes the C portion of a targeted covalent inhibitor of a peptide conjugate / MHC complex, but not the R1 portion.

[0236] Small molecules (e.g., targeted covalent inhibitors) bearing electrophilic warhead groups can undergo covalent reactions with cysteine residues in peptides to form peptide-small molecule conjugates. This type of reaction is illustrated in the following scheme for AMG-510 (sotorasib). [ka]

[0237] The following table lists targeted covalent inhibitors along with the chemical structure of the fragment ("chemical fragment") that binds to a cysteine residue of the peptide via a covalent reaction of the drug with the cysteine. In one embodiment, the binding partners disclosed herein bind to a peptide conjugate / MHC complex comprising a peptide and MHC conjugated to a chemical fragment of Table K below.

[0238] Table K. Structures of target covalent inhibitors and their chemical fragments after covalent reaction with cysteine residues. [Table K1] [Table K2] [Table K3]

[0239] Table R. Structures of exemplary targeted covalent inhibitors targeting G12S [Table R1] [Table R2]

[0240] In non-limiting embodiments, the binding partner specifically binds to a site containing a neoantigen, including a covalently attached small molecule drug or other covalently attached molecule, as a component of an antigen associated with a particular MHC. In one aspect, provided herein are binding partners that specifically bind to a peptide conjugate / MHC complex, where the peptide conjugate is formed by the covalent reaction of a targeted covalent inhibitor with a peptide. In one embodiment, the binding partner binds to the peptide conjugate / MHC complex with higher affinity than to the peptide or the free targeted covalent inhibitor.

[0241] In certain embodiments, the affinity of the binding partner for the peptide conjugate / MHC complex is 100-10,000 times greater than the affinity of the binding partner for the peptide or free target covalent inhibitor. In certain embodiments, the affinity of the binding partner for the peptide conjugate / MHC complex is at least 100 times, at least 200 times, at least 300 times, at least 400 times, at least 500 times, at least 600 times, at least 700 times, at least 800 times, at least 900 times, at least 1,000 times, at least 2,500 times, at least 5,000 times, or at least 10,000 times greater than the affinity of the binding partner for the free target covalent inhibitor or free peptide conjugate. In some embodiments, the affinity of the binding partner for the free drug is measured by an IC 50 In some embodiments, the affinity of the binding partner for the free drug is reported as an IC of greater than 1 μM. 50 In some embodiments, the affinity of the binding partner for the free drug is reported as an IC of greater than 5 μM. 50 In some embodiments, the affinity of the binding partner for the free drug is reported as an IC of greater than 10 μM. 50 In some embodiments, the affinity of the binding partner for the free drug is reported as an IC of greater than 15 μM. 50 In some embodiments, the affinity of the binding partner for the free drug is reported as an IC of greater than 20 μM. 50 In some embodiments, the affinity of the binding partner for the free drug is reported as an IC of greater than 30 μM. 50 In some embodiments, the affinity of the binding partner for the free drug is reported as an IC of greater than 40 μM. 50 In some embodiments, the affinity of the binding partner for the free drug is reported as an IC of greater than 50 μM. 50 In some embodiments, the affinity of the binding partner for the free drug is reported as an IC of greater than 75 μM. 50In some embodiments, the affinity of the binding partner for the free drug is reported as an IC of greater than 100 μM. 50 In some embodiments, the affinity of the binding partner for the free drug is reported as an IC of greater than 200 μM. 50 It is reported as

[0242] In some embodiments, the antigen-binding domain does not detectably bind to the free target covalent inhibitor. In some embodiments, the antigen-binding domain exhibits an IC of at least about 50 nM. 50 In some embodiments, the antigen-binding domain binds to the free target covalent inhibitor with an IC of less than about 50 nM. 50 The antigen-binding domain has an IC of at least about 5 nM and does not bind to free target covalent inhibitors. 50 The antigen-binding domain can bind to the free target covalent inhibitor with an IC of at least about 25 nM. 50 The antigen-binding domain can bind to the free target covalent inhibitor with an IC of at least about 40 nM. 50 The antigen-binding domain can bind to the free target covalent inhibitor with an IC of at least about 60 nM. 50 The antigen-binding domain can bind to the free target covalent inhibitor with an IC of at least about 75 nM. 50 The antigen-binding domain can bind to the free target covalent inhibitor with an IC of at least about 100 nM. 50 The antigen-binding domain can bind to the free target covalent inhibitor with an IC of at least about 250 nM. 50 The antigen-binding domain can bind to the free target covalent inhibitor with an IC of at least about 500 nM. 50 The antigen-binding domain can bind to the free target covalent inhibitor with an IC of at least about 750 nM. 50 The antigen-binding domain can bind to the free target covalent inhibitor with an IC of at least about 1000 nM. 50 The antigen-binding domain can bind to the free target covalent inhibitor with an IC of at least about 2 μM. 50The antigen-binding domain can bind to the free target covalent inhibitor with an IC of at least about 5 μM. 50 The antigen-binding domain can bind to the free target covalent inhibitor with an IC of at least about 10 μM. 50 The antigen-binding domain can bind to the free target covalent inhibitor with an IC of at least about 15 μM. 50 The antigen-binding domain can bind to the free target covalent inhibitor with an IC of at least about 20 μM. 50 The antigen-binding domain can bind to the free target covalent inhibitor with an IC of at least about 25 μM. 50 The antigen-binding domain can bind to the free target covalent inhibitor with an IC of at least about 50 μM. 50 The covalent inhibitor can bind to the free target at the same time.

[0243] In some embodiments, the antigen-binding domain has an IC of less than about 50 nM. 50 The antigen-binding domain can bind to the free target covalent inhibitor with an IC of less than about 5 nM. 50 The antigen-binding domain can bind to the free target covalent inhibitor with an IC of less than about 25 nM. 50 The antigen-binding domain can bind to the free target covalent inhibitor with an IC of less than about 40 nM. 50 The antigen-binding domain can bind to the free target covalent inhibitor with an IC of less than about 60 nM. 50 The antigen-binding domain can bind to the free target covalent inhibitor with an IC of less than about 75 nM. 50 The antigen-binding domain can bind to the free target covalent inhibitor with an IC of less than about 100 nM. 50 The antigen-binding domain can bind to the free target covalent inhibitor with an IC of less than about 250 nM. 50 The antigen-binding domain can bind to the free target covalent inhibitor with an IC of less than about 500 nM. 50 The antigen-binding domain can bind to the free target covalent inhibitor with an IC of less than about 750 nM. 50The antigen-binding domain can bind to the free target covalent inhibitor with an IC of less than about 1000 nM. 50 The antigen-binding domain can bind to the free target covalent inhibitor with an IC of less than about 2 μM. 50 The antigen-binding domain can bind to the free target covalent inhibitor with an IC of less than about 5 μM. 50 The antigen-binding domain can bind to the free target covalent inhibitor with an IC of less than about 10 μM. 50 The antigen-binding domain can bind to the free target covalent inhibitor with an IC of less than about 15 μM. 50 The antigen-binding domain can bind to the free target covalent inhibitor with an IC of less than about 20 μM. 50 The antigen-binding domain can bind to the free target covalent inhibitor with an IC of less than about 25 μM. 50 The antigen-binding domain can bind to the free target covalent inhibitor with an IC of less than about 50 μM. 50 can be bonded with

[0244] The binding partners described herein may not bind to the peptide / MHC complex without the conjugate (e.g., a targeted covalent inhibitor). In some cases, the affinity of the binding partner for the peptide conjugate / MHC complex may be such that the binding partner has a dissociation constant (K) of 10 pM to 50 nM. D In some cases, the affinity of the binding partner for the peptide conjugate / MHC complex is reported as a dissociation constant (K) between 1 pM and 10 pM. D In some cases, the affinity of the binding partner for the peptide conjugate / MHC complex is reported as a dissociation constant (K) between 0.01 pM and 10 pM. D In some cases, the affinity of the binding partner for the peptide conjugate / MHC complex is reported as a dissociation constant (K) between 0.1 nM and 1 nM. D In some cases, the affinity of the binding partner for the peptide conjugate / MHC complex is reported as a dissociation constant (K) of 1 nM to 2 nM. DIn some cases, the affinity of the binding partner for the peptide conjugate / MHC complex is reported as a dissociation constant (K) of 2 nM to 5 nM. D In some cases, the affinity of the binding partner for the peptide conjugate / MHC complex is reported as a dissociation constant (K) of about 5 nM or less. D In some cases, the affinity of the binding partner for the peptide conjugate / MHC complex is reported as a dissociation constant (K) of about 4 nM or less. D In some cases, the affinity of the binding partner for the peptide conjugate / MHC complex is reported as a dissociation constant (K) of about 3 nM or less. D In some cases, the affinity of the binding partner for the peptide conjugate / MHC complex is reported as a dissociation constant (K) of about 2 nM or less. D In some cases, the affinity of the binding partner for the peptide conjugate / MHC complex is reported as a dissociation constant (K) of about 1 nM or less. D In some cases, the affinity of the binding partner for the peptide conjugate / MHC complex is reported as a dissociation constant (K) of about 0.1 nM or less. D In some cases, the affinity of the binding partner for the peptide conjugate / MHC complex is reported as a dissociation constant (K) of about 0.01 nM or less. D In some cases, the affinity of the binding partner for the peptide conjugate / MHC complex is reported as a dissociation constant (K) of about 1 pM or less. D In some cases, the affinity of the binding partner for the peptide conjugate / MHC complex is reported as a dissociation constant (K) of about 0.1 pM or less. D )

[0245] In some embodiments, the affinity of a binding partner for a peptide conjugate / MHC complex is measured by IC 50 or EC 50In some embodiments, the affinity of the binding partner for the peptide conjugate / MHC complex is reported as an IC of less than about 1 pm. 50 In some embodiments, the affinity of the binding partner for the peptide conjugate / MHC complex is reported as an IC of less than about 10 pm. 50 In some embodiments, the affinity of the binding partner for the peptide conjugate / MHC complex is reported as an IC of less than about 100 pm. 50 In some embodiments, the affinity of the binding partner for the peptide conjugate / MHC complex is reported as an IC of less than about 1000 pm. 50 In some embodiments, the affinity of the binding partner for the peptide conjugate / MHC complex is reported as an IC of less than about 1 nm. 50 In some embodiments, the affinity of the binding partner for the peptide conjugate / MHC complex is reported as an IC of less than about 10 nM. 50 In some embodiments, the affinity of the binding partner for the peptide conjugate / MHC complex is reported as an IC of less than about 100 nM. 50 In some embodiments, the affinity of the binding partner for the peptide conjugate / MHC complex is reported as an IC of less than about 1000 nm. 50 In some embodiments, the affinity of the binding partner for the peptide conjugate / MHC complex is reported as an IC of less than about 1 μM. 50 It is reported as

[0246] In some embodiments, the binding partner does not detectably bind to the peptide-MHC molecule complex, where the peptide is not covalently bound to a non-peptide molecule. In some embodiments, the binding partner binds to the peptide-MHC molecule complex with less than 10-fold affinity, where the peptide is not covalently bound to a non-peptide molecule. In some embodiments, the binding partner binds to the peptide-MHC molecule complex with less than 100-fold affinity, where the peptide is not covalently bound to a non-peptide molecule. In some embodiments, the binding partner binds to the peptide-MHC molecule complex with less than 1,000-fold affinity, where the peptide is not covalently bound to a non-peptide molecule.

[0247] In some embodiments, the binding partner can be a polypeptide. In some embodiments, the polypeptide can have an antigen-binding domain. The antigen-binding domain has a dissociation constant (K) of at most about 50 nM, at most about 40 nM, at most about 30 nM, at most about 20 nM, at most about 10 nM, at most about 1 nM, at most about 0.1 nM, at most about 10 pM, or at most about 1 pM. D The antigen-binding domain may bind to the peptide conjugate / MHC complex with a dissociation constant (K) of about 0.01 nM to about 20 nM. D ) can bind to the peptide conjugate / MHC complex.

[0248] The antigen-binding domain is the K of the antibody or antigen-binding fragment to the peptide conjugate / MHC complex. D higher dissociation constant (K D In some cases, the antigen-binding domain may bind to the free target covalent inhibitor with a dissociation constant (K) of at least about 100 nM, at least about 200 nM, at least about 300 nM, at least about 400 nM, at least about 500 nM, at least about 1 μM, at least about 10 μM, at least about 20 μM, at least about 30 μM, at least about 40 μM, at least about 50 μM, or at least about 100 μM. D) and can bind to the free target covalent inhibitor. The antigen-binding domain is the K of the antigen-binding domain that binds to the peptide conjugate / MHC complex. D at least about 2, 3, 4, 5, 10, 20, 50, 100, 200, 500, 1,000, 2,000, 5,000, 10,000, 20,000, 50,000, 100,000 times higher dissociation constant (K D ) and can bind to a free target covalent inhibitor. The antigen-binding domain is the K of the antibody or antigen-binding fragment to the peptide conjugate / MHC complex. D higher dissociation constant (K D The antigen-binding domain may bind to the free peptide conjugate with a dissociation constant (K) of greater than about 100 nM, greater than about 200 nM, greater than about 300 nM, greater than about 400 nM, greater than about 500 nM, greater than 1 μM, greater than 10 μM, greater than 20 μM, greater than 30 μM, greater than 40 μM, greater than 50 μM, or greater than 100 μM. D ) and can bind to the free peptide conjugate. The antigen-binding domain can be expressed as the K of the antigen-binding domain bound to the peptide conjugate / MHC complex. D a dissociation constant (K) that is at least about 10-fold, 100-fold, 10,000-fold, or 100,000-fold higher than D ) and can bind to the free peptide conjugate. The antigen-binding domain is the K Dthan at least 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3, 3.1, 3.2, 3.3, 3.4, 3.5, 3.6, 3.7, 3.8, 3.9, 4, 4.1, 4.2, 4.3, 4.4, 4.5, 4.6, 4.7, 4.8, 4.9, 5, 5.1, 5.2, 5.3, 5.4, 5.5, 5.6, 5.7, 5.8, 5.9, 6, 6.1, 6.2, 6.3, 6.4, 6.5, 6.6, 6.7, 6.8, 6.9, 7, 7.1, 7.2, 7.3, 7.4, 7.5, 7.6, 7.7, 7.8, 7.9, 8, 8.1, 8.2, 8.3, 8.4, 8.5, 8.6, 8.7, 8.8, 8.9, 9, 9.1, 9.2, 9.3, 9.4, 9.5, 9.6, 9.7, 9.8, 9.9 or a 10-fold higher dissociation constant (K D ) can bind to the free peptide conjugate. In some embodiments, the antigen-binding domain can bind to the free peptide conjugate at a K D at least 2.5-fold higher dissociation constant (K D ) to the free peptide conjugate. The antigen-binding domain may bind to the peptide conjugate / MHC complex with an affinity that is at least 100-fold, at least 200-fold, at least 300-fold, at least 400-fold, at least 500-fold, at least 600-fold, at least 700-fold, at least 800-fold, at least 900-fold, at least 1,000-fold, at least 2,500-fold, at least 5,000-fold, or at least 10,000-fold greater than the affinity of the antigen-binding domain for the free target covalent inhibitor or the free peptide conjugate. The MHC may be HLA-A02:01. The MHC may be HLA-A03:01.

[0249] In one embodiment, the MHC is a human leukocyte antigen (HLA). In one embodiment, the HLA is any HLA found in AFND (Allele Frequency Net Database) (allelefrequencies.net). In one embodiment, the HLA is HLA-A*02:01, HLA-A*03:01, HLA-A*01:01, HLA-A*02:01, HLA-A*03:01, HLA-A*11:01, HLA-A*24:02, HLA-A*26:01, HLA-B*07:02, HLA-B*08:01, HLA-B*27:05, HLA-B*39:01, HLA-B*40:01, HLA-B*58:01, HLA-B*15:01, and / or HLA-A*11:01.

[0250] In one aspect, provided herein is a cell-free peptide conjugate / MHC complex comprising: (a) a peptide conjugate formed by covalent reaction of a targeted covalent inhibitor with a peptide; and (b) MHC. In one embodiment, the peptide comprises: a segment of RAS (e.g., KRAS G12C , KRAS G12D , KRAS G12R , KRAS G12S , HRAS G12C , HRAS G12D , HRAS G12R , HRAS G12S NRAS G12C , NRAS G12D , NRAS G12R or NRAS G12S), EGFR, BTK, HER2 / NEU (ERBB2), HER3 (ERBB3), HER4 (ERBB4), MET (HGFR); FGFR, CDK, acetylcholinesterase (ACHE), p90 ribosomal S6 kinase (RSK), TP53, IDH1, GNAS, FBXW7, CTNNB1, DNMT3A, cathepsin B, cathepsin C, cathepsin F, cathepsin H, cathepsin K, cathepsin L, cathepsin O, cathepsin S, cathepsin V, cathepsin W, cathepsin X, caspase, pancreatic lipase, METAP2, any cancer-testis antigen, any endogenous retroviral protein, long interspersed nucleotide sequence-1 (LINE-1), or short interspersed nucleotide sequence (SINE). In one embodiment, the short interspersed nucleotide sequence is Alu. In certain embodiments, the targeted covalent inhibitor is AMG-510, ARS-853, ARS-1620, ARS-3248, MRTX849, JNJ74699157, LY3499446, LY3537982, MRTX-1257, JDQ443, MRTX-1133, RMC-6291, RMC-9805, GDC-6036, D-1553, 2E07, 6H05, SML-8-73-1 or BI 182391. Additional examples of compounds capable of covalently targeting KRAS peptides containing the G12C mutation can be found in PCT / IB2019 / 050993, PCT / EP2018 / 083853, and U.S. application US 16 / 917,128, each of which is incorporated herein by reference in its entirety. In other embodiments, the targeted covalent inhibitor is selected from β-lactones G12Si-1, G12Si-2, G12Si-3, G12Si-4, and G12Si-5.

[0251] In some embodiments, provided herein is a cell-free peptide conjugate / MHC complex comprising: (a) a compound selected from the group consisting of compounds 1 and 4-8, covalently attached to a cysteine residue in a peptide comprising the amino acid sequence VVVGACGVGK, VVGACGVGK, or KLVVVGACGV: [ka] [ka] and (b) MHC.

[0252] In some embodiments, provided herein is a cell-free peptide conjugate / MHC complex comprising: (a) Compound 2 covalently attached to a cysteine residue in a peptide comprising the amino acid sequence QLMPFGCLL, LMPFGCLLDY, or MPFGCLLDY: [ka] ; and (b) MHC.

[0253] In some embodiments, provided herein is a cell-free peptide conjugate / MHC complex comprising: (a) Compound 3 covalently attached to a cysteine residue in a peptide comprising the amino acid sequence YMANGCLLNY: [ka] ; and (b) MHC.

[0254] In one embodiment, the MHC is HLA, optionally wherein the HLA is HLA-A*02:01, HLA-A*03:01, HLA-A*01:01, HLA-A*02:01, HLA-A*03:01, HLA-A*11:01, HLA-A*24:02, HLA-A*26:01, HLA-B*07:02, HLA-B*08:01, HLA-B*27:05, HLA-B*39:01, HLA-B*40:01, HLA-B*58:01, HLA-B*15:01, or HLA-A*11:01.

[0255] In one aspect, provided herein is a combination of AMG-510 and HLA-presented KRAS G12Ca binding partner that specifically binds to a conjugate formed by a covalent reaction with the peptide, wherein the binding partner comprises a heavy chain variable region (VH) and a light chain variable region (VL), (a) VH comprises: (i) CDR-H1 containing the amino acid sequence DYSIH, or a variant thereof containing 1 to 5 amino acid changes; (ii) a CDR-H2 comprising the amino acid sequence SISSSSGSTSYADSVKG, or a variant thereof comprising 1 to 5 amino acid changes; and / or (iii) a CDR-H3 comprising the amino acid sequence GX1WX2X3AMDY, where X1 is G, R, H, S, or K, X2 is Y or I, and X3 is P or A; and / or (b) VL includes: (i) CDR-L1 containing the amino acid sequence RASQSVSSAVA, or a variant thereof containing 1 to 5 amino acid changes; (ii) a CDR-L2 comprising the amino acid sequence SASSLYS, or a variant thereof comprising 1 to 5 amino acid changes; and / or (iii) CDR-L3 comprising the amino acid sequence QQX1SYVX2X3X4IT, where X1 is I, A, P, V, or S, X2 is K, R, A, or H, X3 is K or R, and X4 is L, T, K, R, V, A, or E.

[0256] The CDR sequences of exemplary antibodies that bind to conjugates formed by the covalent reaction of AMG-510 with the KRAS(G12C) peptide presented on HLA-A*03:01 and HLA-A*11:01 are shown in Tables G and H below.

[0257] Table G: Heavy chain CDR sequences of exemplary antibodies that bind to conjugates formed by the covalent reaction of AMG-510 with KRAS(G12C) peptide presented on HLA-A*03:01 and HLA-A*11:01. [Table G]

[0258] Table H: Light chain CDR sequences of exemplary antibodies that bind to conjugates formed by the covalent reaction of AMG-510 with KRAS(G12C) peptide presented on HLA-A*03:01 and HLA-A*11:01. [Table H]

[0259] In one embodiment, the binding partner comprises the amino acid sequences CDR-H1, CDR-H2 and CDR-H3 of the heavy chain variable region of the binding partner, and / or CDR-L1, CDR-L2 and CDR-L3 of the light chain variable region of the binding partner, wherein the binding partner is selected from the group consisting of RA_D11, RA_D01-RA_D04, RA_D06-RA_D09, RA_D12-RA_D14, RA_D16, RA_D18-RA_D21, RA_D23 and RA_D24, or variants thereof having 1 to 5 amino acid changes in one or more of the CDR amino acid sequences.

[0260] In one embodiment, the binding partner comprises the CDR-H1, CDR-H2, CDR-H3, CDR-L1, CDR-L2 and / or CDR-L3 amino acid sequences of the binding partner, wherein the binding partner is selected from the group consisting of RA_D11, RA_D01-RA_D04, RA_D06-RA_D09, RA_D12-RA_D14, RA_D16, RA_D18-RA_D21, RA_D23 and RA_D24 (Tables G and H), or variants thereof having 1 to 5 amino acid changes in one or more of the CDR amino acid sequences.

[0261] In one embodiment, a binding partner may comprise a VH and / or VL amino acid sequence having at least about 40%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 100% sequence identity to the following VH and VL sequences described herein:

[0262] In one embodiment, a binding partner may comprise a VH and / or VL amino acid sequence having at least about 40%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 100% sequence identity to the VH and / or VL of RA_D11, RA_D01-RA_D04, RA_D06-RA_D09, RA_D12-RA_D14, RA_D16, RA_D18-RA_D21, RA_D23 and RA_D24 (Tables G and H).

[0263] AMG-510 and KRAS G12C A non-limiting example of a reference binding partner that is formed by covalent reaction with a peptide and binds to a conjugate presented by an HLA molecule is referred to herein as RA_D11.

[0264] The amino acid sequences of the light chain variable region (VL) and heavy chain variable region (VH) of RA_D11 are as follows (CDRs are shown in bold; underlined residues correspond to those shown in Tables E and F in the figure): TIFF2025526240000034.tif36158

[0265] A non-limiting example of a reference binding partner that binds to KRAS(G12C)-AMG-510 presented by HLA molecules is referred to herein as AMRA3-7D. L ) and heavy chain variable region (V H ) has the following amino acid sequence: TIFF2025526240000035.tif36158

[0266] Data including a comparison of AMRA3-7D and RA-D11 (and other binding partners) binding to the HLA-A*11 associated AMG510-G12C 9mer are shown in FIG.

[0267] In another embodiment, the present disclosure provides a binding partner (or antigen-binding domain) with improved binding affinity to a drug-peptide / MHC complex. The peptide can be a KRAS peptide. The drug can be sotorasib (e.g., AMG-510). The drug-peptide conjugate can be a KRAS(G12C) peptide conjugated to sotorasib. The binding partner can bind to KRAS(G12C)-sotorasib presented by an MHC molecule. In some embodiments, the MHC can be HLA-A*02:01, HLA-A*03:01, or HLA-A*11:01. The binding partners described herein can include various antigen-binding domains disclosed herein.

[0268] For example, in some embodiments, the antigen-binding domain may comprise a heavy chain variable region (VH), wherein the VH may comprise a heavy chain complementarity-determining region 3 (CDR-H3), wherein the CDR-H3 comprises the amino acid sequence GSWIHAMDY. The VH may further comprise a CDR-H2 comprising the amino acid sequence SISSSWGVTSYADSVKG. The VH may further comprise a CDR-H1 comprising the amino acid sequence FHWYSIH. In some embodiments, the antigen-binding domain may further comprise a light chain variable region (VL), wherein the VL may comprise a CDR-L3 comprising the amino acid sequence QQASYVRKTIT. The VL may further comprise a CDR-L2 comprising the amino acid sequence SASSLYS. The VL may further comprise a CDR-L1 comprising the amino acid sequence RASQSVSSAVA. The antigen-binding domain can comprise a CDR-H3 sequence of GSWIHAMDY; a CDR-H2 sequence of SISSSWGVTSYADSVKG; a CDR-H1 sequence of FHWYSIH; a CDR-L3 sequence of QQASYVRKTIT; a CDR-L2 sequence of SASSLYS; and a CDR-L1 sequence of RASQSVSSAVA. The VH can comprise a sequence having at least 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the following sequence: EVQLVESGGGLVQPGGSLRLSCAASGFTFHWYSIHWVRQAPGKGLEWVASISSSWGVTSYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARGSWIHAMDYWGQGTLVTVSS The VH may comprise a sequence having at least 80% sequence identity to the following sequence: EVQLVESGGGLVQPGGSLRLSCAASGFTFHWYSIHWVRQAPGKGLEWVASISSSWGVTSYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARGSWIHAMDYWGQGTLVTVSS The VL can comprise a sequence having at least 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the following sequence: DIQMTQSPSSLSASVGDRVTITCRASQSVSSAVAWYQQKPGKAPKLLIYSASSLYSGVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQASYVRKTITFGQGTKVEIKRTV The VL may comprise a sequence having at least 80% sequence identity to the following sequence: DIQMTQSPSSLSASVGDRVTITCRASQSVSSAVAWYQQKPGKAPKLLIYSASSLYSGVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQASYVRKTITFGQGTKVEIKRTV

[0269] In some embodiments, the antigen-binding domain comprises a heavy chain variable region (VH), wherein the VH may comprise a heavy chain complementarity-determining region 3 (CDR-H3), wherein the CDR-H3 comprises the amino acid sequence GHWIAAMDY. The VH may further comprise a CDR-H2 comprising the amino acid sequence SIASSSGSTGYADSVKG. The VH may further comprise a CDR-H1 comprising the amino acid sequence FSWYSIH. In some embodiments, the antigen-binding domain may further comprise a light chain variable region (VL), wherein the VL may comprise a CDR-L3 comprising the amino acid sequence QQASYVRKTIT. The VL may further comprise a CDR-L2 comprising the amino acid sequence SASSLYS. The VL may further comprise a CDR-L1 comprising the amino acid sequence RASQSVSSAVA. The antigen-binding domain can comprise a CDR-H3 sequence of GHWIAAMDY; a CDR-H2 sequence of SIASSSGSTGYADSVKG; a CDR-H1 sequence of FSWYSIH; a CDR-L3 sequence of QQASYVRKTIT; a CDR-L2 sequence of SASSLYS; and a CDR-L1 sequence of RASQSVSSAVA. The VH can comprise a sequence having at least 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the following sequence: EVQLVESGGGLVQPGGSLRLSCAASGFTFSWYSIHWVRQAPGKGLEWVASIASSSGSTGYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARGHWIAAMDYWGQGTLVTVSS. The VH may comprise a sequence having at least 80% sequence identity to the following sequence: EVQLVESGGGLVQPGGSLRLSCAASGTFSWYSIHWVRQAPGKGLEWVASIASSSGSTGYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARGHWIAAMDYWGQGTLVTVSS The VL can comprise a sequence having at least 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the following sequence: DIQMTQSPSSLSASVGDRVTITCRASQSVSSAVAWYQQKPGKAPKLLIYSASSLYSGVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQASYVRKTITFGQGTKVEIKRTV The VL may comprise a sequence having at least 80% sequence identity to the following sequence: DIQMTQSPSSLSASVGDRVTITCRASQSVSSAVAWYQQKPGKAPKLLIYSASSLYSGVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQASYVRKTITFGQGTKVEIKRTV

[0270] In some embodiments, the antigen-binding domain comprises a heavy chain variable region (VH), wherein the VH may comprise a heavy chain complementarity-determining region 3 (CDR-H3), wherein the CDR-H3 comprises the amino acid sequence GGVIHAMDY. The VH may further comprise a CDR-H2 comprising the amino acid sequence SILSRWGVTSYADSVKG. The VH may further comprise a CDR-H1 comprising the amino acid sequence FSPYSIH. In some embodiments, the antigen-binding domain may further comprise a light chain variable region (VL), wherein the VL may comprise a CDR-L3 comprising the amino acid sequence QQASYVRKTIT. The VL may further comprise a CDR-L2 comprising the amino acid sequence SASSLYS. The VL may further comprise a CDR-L1 comprising the amino acid sequence RASQSVSSAVA. The antigen-binding domain can comprise the CDR-H3 sequence of GGVIHAMDY; the CDR-H2 sequence of SILSRWGVTSYADSVKG; the CDR-H1 sequence of FSPYSIH; the CDR-L3 sequence of QQASYVRKTIT; the CDR-L2 sequence of SASSLYS; and the CDR-L1 sequence of RASQSVSSAVA. The VH can comprise a sequence having at least 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the following sequence: EVQLVESGGGLVQPGGSLRLSCAASGFTFSPYSIHWVRQAPGKGLEWVASILSRWGVTSYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARGGVIHAMDYWGQGTLVTVSS The VH may comprise a sequence having at least 80% sequence identity to the following sequence: EVQLVESGGGLVQPGGSLRLSCAASGFTFSPYSIHWVRQAPGKGLEWVASILSRWGVTSYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARGGVIHAMDYWGQGTLVTVSS The VL can comprise a sequence having at least 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the following sequence: DIQMTQSPSSLSASVGDRVTITCRASQSVSSAVAWYQQKPGKAPKLLIYSASSLYSGVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQASYVRKTITFGQGTKVEIKRTV The VL may comprise a sequence having at least 80% sequence identity to the following sequence: DIQMTQSPSSLSASVGDRVTITCRASQSVSSAVAWYQQKPGKAPKLLIYSASSLYSGVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQASYVRKTITFGQGTKVEIKRTV

[0271] In some embodiments, the antigen-binding domain comprises a heavy chain variable region (VH), wherein the VH may comprise a heavy chain complementarity-determining region 3 (CDR-H3), wherein the CDR-H3 comprises the amino acid sequence GSWIAAMDY. The VH may further comprise a CDR-H2 comprising the amino acid sequence SISSWHGETGYADSVKG. The VH may further comprise a CDR-H1 comprising the amino acid sequence FSPYSIH. In some embodiments, the antigen-binding domain may further comprise a light chain variable region (VL), wherein the VL may comprise a CDR-L3 comprising the amino acid sequence QQASYVRKTIT. The VL may further comprise a CDR-L2 comprising the amino acid sequence SASSLYS. The VL may further comprise a CDR-L1 comprising the amino acid sequence RASQSVSSAVA. The antigen-binding domain can comprise the CDR-H3 sequence of GSWIAAMDY; the CDR-H2 sequence of SISSWHGETGYADSVKG; the CDR-H1 sequence of FSPYSIH; the CDR-L3 sequence of QQASYVRKTIT; the CDR-L2 sequence of SASSLYS; and the CDR-L1 sequence of RASQSVSSAVA. The VH can comprise a sequence having at least 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the following sequence: EVQLVESGGGLVQPGGSLRLSCAASGFTFSPYSIHWVRQAPGKGLEWVASISSWHGETGYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARGSWIAAMDYWGQGTLVTVSS The VH may comprise a sequence having at least 80% sequence identity to the following sequence: EVQLVESGGGLVQPGGSLRLSCAASGFTFSPYSIHWVRQAPGKGLEWVASISSWHGETGYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARGSWIAAMDYWGQGTLVTVSS The VL can comprise a sequence having at least 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the following sequence: DIQMTQSPSSLSASVGDRVTITCRASQSVSSAVAWYQQKPGKAPKLLIYSASSLYSGVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQASYVRKTITFGQGTKVEIKRTV The VL may comprise a sequence having at least 80% sequence identity to the following sequence: DIQMTQSPSSLSASVGDRVTITCRASQSVSSAVAWYQQKPGKAPKLLIYSASSLYSGVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQASYVRKTITFGQGTKVEIKRTV

[0272] In some embodiments, the antigen-binding domain comprises a heavy chain variable region (VH), which may comprise a heavy chain complementarity-determining region 3 (CDR-H3), which comprises the amino acid sequence GGWIAAMDY. The VH may further comprise a CDR-H2 comprising the amino acid sequence SISSLQGDTGYADSVKG. The VH may further comprise a CDR-H1 comprising the amino acid sequence FSWYSIH. In some embodiments, the antigen-binding domain may further comprise a light chain variable region (VL), which may comprise a CDR-L3 comprising the amino acid sequence QQASYVRKTIT. The VL may further comprise a CDR-L2 comprising the amino acid sequence SASSLYS. The VL may further comprise a CDR-L1 comprising the amino acid sequence RASQSVSSAVA. The antigen-binding domain can comprise a CDR-H3 sequence of GGWIAAMDY; a CDR-H2 sequence of SISSLQGDTGYADSVKG; a CDR-H1 sequence of FSWYSIH; a CDR-L3 sequence of QQASYVRKTIT; a CDR-L2 sequence of SASSLYS; and a CDR-L1 sequence of RASQSVSSAVA. The VH can comprise a sequence having at least 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the following sequence: EVQLVESGGGLVQPGGSLRLSCAASGTFSWYSIHWVRQAPGKGLEWVASISSLQGDTGYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARGGWIAAMDYWGQGTLVTVSS The VH may comprise a sequence having at least 80% sequence identity to the following sequence: EVQLVESGGGLVQPGGSLRLSCAASGTFSWYSIHWVRQAPGKGLEWVASISSLQGDTGYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARGGWIAAMDYWGQGTLVTVSS The VL can comprise a sequence having at least 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the following sequence: DIQMTQSPSSLSASVGDRVTITCRASQSVSSAVAWYQQKPGKAPKLLIYSASSLYSGVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQASYVRKTITFGQGTKVEIKRTV The VL may comprise a sequence having at least 80% sequence identity to the following sequence: DIQMTQSPSSLSASVGDRVTITCRASQSVSSAVAWYQQKPGKAPKLLIYSASSLYSGVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQASYVRKTITFGQGTKVEIKRTV

[0273] In some embodiments, the antigen-binding domain comprises a heavy chain variable region (VH), wherein the VH may comprise a heavy chain complementarity-determining region 3 (CDR-H3), wherein the CDR-H3 comprises the amino acid sequence GSWIAAMDY. The VH may further comprise a CDR-H2 comprising the amino acid sequence SIASWYGDTGYADSVKG. The VH may further comprise a CDR-H1 comprising the amino acid sequence FHYYSIH. In some embodiments, the antigen-binding domain may further comprise a light chain variable region (VL), wherein the VL may comprise a CDR-L3 comprising the amino acid sequence QQASYVRKTIT. The VL may further comprise a CDR-L2 comprising the amino acid sequence SASSLYS. The VL may further comprise a CDR-L1 comprising the amino acid sequence RASQSVSSAVA. The antigen-binding domain can comprise a CDR-H3 sequence of GSWIAAMDY; a CDR-H2 sequence of SIASWYGDTGYADSVKG; a CDR-H1 sequence of FHYYSIH; a CDR-L3 sequence of QQASYVRKTIT; a CDR-L2 sequence of SASSLYS; and a CDR-L1 sequence of RASQSVSSAVA. The VH can comprise a sequence having at least 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the following sequence: EVQLVESGGGLVQPGGSLRLSCAASGFTFHYYSIHWVRQAPGKGLEWVASIASWYGDTGYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARGSWIAAMDYWGQGTLVTVSS The VH may comprise a sequence having at least 80% sequence identity to the following sequence: EVQLVESGGGLVQPGGSLRLSCAASGFTFHYYSIHWVRQAPGKGLEWVASIASWYGDTGYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARGSWIAAMDYWGQGTLVTVSS The VL can comprise a sequence having at least 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the following sequence: DIQMTQSPSSLSASVGDRVTITCRASQSVSSAVAWYQQKPGKAPKLLIYSASSLYSGVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQASYVRKTITFGQGTKVEIKRTV The VL may comprise a sequence having at least 80% sequence identity to the following sequence: DIQMTQSPSSLSASVGDRVTITCRASQSVSSAVAWYQQKPGKAPKLLIYSASSLYSGVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQASYVRKTITFGQGTKVEIKRTV

[0274] In some embodiments, the antigen-binding domain comprises a heavy chain variable region (VH), which may comprise a heavy chain complementarity-determining region 3 (CDR-H3), which comprises the amino acid sequence GGRIEAMDY. The VH may further comprise a CDR-H2 comprising the amino acid sequence SISSWYGKTGYADSVKG. The VH may further comprise a CDR-H1 comprising the amino acid sequence FGYYSIH. In some embodiments, the antigen-binding domain may further comprise a light chain variable region (VL), which may comprise a CDR-L3 comprising the amino acid sequence QQASYVRKTIT. The VL may further comprise a CDR-L2 comprising the amino acid sequence SASSLYS. The VL may further comprise a CDR-L1 comprising the amino acid sequence RASQSVSSAVA. The antigen-binding domain can comprise a CDR-H3 sequence of GGRIEAMDY; a CDR-H2 sequence of SISSWYGKTGYADSVKG; a CDR-H1 sequence of FGYYSIH; a CDR-L3 sequence of QQASYVRKTIT; a CDR-L2 sequence of SASSLYS; and a CDR-L1 sequence of RASQSVSSAVA. The VH can comprise a sequence having at least 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the following sequence: EVQLVESGGGLVQPGGSLRLSCAASGFTFGYYSIHWVRQAPGKGLEWVASISSWYGKTGYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARGGRIEAMDYWGQGTLVTVSS The VH may comprise a sequence having at least 80% sequence identity to the following sequence: EVQLVESGGGLVQPGGSLRLSCAASGFTFGYYSIHWVRQAPGKGLEWVASISSWYGKTGYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARGGRIEAMDYWGQGTLVTVSS The VL can comprise a sequence having at least 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the following sequence: DIQMTQSPSSLSASVGDRVTITCRASQSVSSAVAWYQQKPGKAPKLLIYSASSLYSGVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQASYVRKTITFGQGTKVEIKRTV The VL may comprise a sequence having at least 80% sequence identity to the following sequence: DIQMTQSPSSLSASVGDRVTITCRASQSVSSAVAWYQQKPGKAPKLLIYSASSLYSGVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQASYVRKTITFGQGTKVEIKRTV

[0275] In some embodiments, the antigen-binding domain comprises a heavy chain variable region (VH), wherein the VH may comprise a heavy chain complementarity-determining region 3 (CDR-H3), wherein the CDR-H3 comprises the amino acid sequence GYWIEAMDY. The VH may further comprise a CDR-H2 comprising the amino acid sequence SIASSYGSTGYADSVKG. The VH may further comprise a CDR-H1 comprising the amino acid sequence FSKYSIH. In some embodiments, the antigen-binding domain may further comprise a light chain variable region (VL), wherein the VL may comprise a CDR-L3 comprising the amino acid sequence QQASYVRKTIT. The VL may further comprise a CDR-L2 comprising the amino acid sequence SASSLYS. The VL may further comprise a CDR-L1 comprising the amino acid sequence RASQSVSSAVA. The antigen-binding domain can comprise a CDR-H3 sequence of GYWIEAMDY; a CDR-H2 sequence of SIASSYGSTGYADSVKG; a CDR-H1 sequence of FSKYSIH; a CDR-L3 sequence of QQASYVRKTIT; a CDR-L2 sequence of SASSLYS; and a CDR-L1 sequence of RASQSVSSAVA. The VH can comprise a sequence having at least 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the following sequence: EVQLVESGGGLVQPGGSLRLSCAASGFTFSKYSIHWVRQAPGKGLEWVASIASSYGSTGYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARGYWIEAMDYWGQGTLVTVSS The VH may comprise a sequence having at least 80% sequence identity to the following sequence: EVQLVESGGGLVQPGGSLRLSCAASGFTFSKYSIHWVRQAPGKGLEWVASIASSYGSTGYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARGYWIEAMDYWGQGTLVTVSS The VL can comprise a sequence having at least 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the following sequence: DIQMTQSPSSLSASVGDRVTITCRASQSVSSAVAWYQQKPGKAPKLLIYSASSLYSGVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQASYVRKTITFGQGTKVEIKRTV The VL may comprise a sequence having at least 80% sequence identity to the following sequence: DIQMTQSPSSLSASVGDRVTITCRASQSVSSAVAWYQQKPGKAPKLLIYSASSLYSGVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQASYVRKTITFGQGTKVEIKRTV

[0276] In some embodiments, the antigen-binding domain comprises a heavy chain variable region (VH), which may comprise a heavy chain complementarity-determining region 3 (CDR-H3), which comprises the amino acid sequence GSWIAAMDY. The VH may further comprise a CDR-H2 comprising the amino acid sequence SIHSSIGTTGYADSVKG. The VH may further comprise a CDR-H1 comprising the amino acid sequence FGLYSIH. In some embodiments, the antigen-binding domain may further comprise a light chain variable region (VL), which may comprise a CDR-L3 comprising the amino acid sequence QQASYVRKTIT. The VL may further comprise a CDR-L2 comprising the amino acid sequence SASSLYS. The VL may further comprise a CDR-L1 comprising the amino acid sequence RASQSVSSAVA. The antigen-binding domain can comprise the CDR-H3 sequence of GSWIAAMDY; the CDR-H2 sequence of SIHSSIGTTGYADSVKG; the CDR-H1 sequence of FGLYSIH; the CDR-L3 sequence of QQASYVRKTIT; the CDR-L2 sequence of SASSLYS; and the CDR-L1 sequence of RASQSVSSAVA. The VH can comprise a sequence having at least 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the following sequence: EVQLVESGGGLVQPGGSLRLSCAASGFTFGLYSIHWVRQAPGKGLEWVASIHSSIGTTGYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARGSWIAAMDYWGQGTLVTVSS The VH may comprise a sequence having at least 80% sequence identity to the following sequence: EVQLVESGGGLVQPGGSLRLSCAASGFTFGLYSIHWVRQAPGKGLEWVASIHSSIGTTGYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARGSWIAAMDYWGQGTLVTVSS The VL can comprise a sequence having at least 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the following sequence: DIQMTQSPSSLSASVGDRVTITCRASQSVSSAVAWYQQKPGKAPKLLIYSASSLYSGVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQASYVRKTITFGQGTKVEIKRTV The VL may comprise a sequence having at least 80% sequence identity to the following sequence: DIQMTQSPSSLSASVGDRVTITCRASQSVSSAVAWYQQKPGKAPKLLIYSASSLYSGVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQASYVRKTITFGQGTKVEIKRTV

[0277] In some embodiments, the antigen-binding domain comprises a heavy chain variable region (VH), wherein the VH may comprise a heavy chain complementarity-determining region 3 (CDR-H3), wherein the CDR-H3 comprises the amino acid sequence GSVIHAMDY. The VH may further comprise a CDR-H2 comprising the amino acid sequence SILSWIGKTSYADSVKG. The VH may further comprise a CDR-H1 comprising the amino acid sequence FSPYSIH. In some embodiments, the antigen-binding domain may further comprise a light chain variable region (VL), wherein the VL may comprise a CDR-L3 comprising the amino acid sequence QQASYVRKTIT. The VL may further comprise a CDR-L2 comprising the amino acid sequence SASSLYS. The VL may further comprise a CDR-L1 comprising the amino acid sequence RASQSVSSAVA. The antigen-binding domain may comprise the CDR-H3 sequence of GSVIHAMDY; the CDR-H2 sequence of SILSWIGKTSYADSVKG; the CDR-H1 sequence of FSPYSIH; the CDR-L3 sequence of QQASYVRKTIT; the CDR-L2 sequence of SASSLYS; and the CDR-L1 sequence of RASQSVSSAVA. The VH can comprise a sequence having at least 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the following sequence: EVQLVESGGGLVQPGGSLRLSCAASGFTFSPYSIHWVRQAPGKGLEWVASILSWIGKTSYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARGSVIHAMDYWGQGTLVTVSS The VH may comprise a sequence having at least 80% sequence identity to the following sequence: EVQLVESGGGLVQPGGSLRLSCAASGFTFSPYSIHWVRQAPGKGLEWVASILSWIGKTSYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARGSVIHAMDYWGQGTLVTVSS The VL can comprise a sequence having at least 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the following sequence: DIQMTQSPSSLSASVGDRVTITCRASQSVSSAVAWYQQKPGKAPKLLIYSASSLYSGVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQASYVRKTITFGQGTKVEIKRTV The VL may comprise a sequence having at least 80% sequence identity to the following sequence: DIQMTQSPSSLSASVGDRVTITCRASQSVSSAVAWYQQKPGKAPKLLIYSASSLYSGVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQASYVRKTITFGQGTKVEIKRTV

[0278] In some embodiments, the antigen-binding domain comprises a heavy chain variable region (VH), which may comprise a heavy chain complementarity-determining region 3 (CDR-H3), which comprises the amino acid sequence GGWIAAMDY. The VH may further comprise a CDR-H2 comprising the amino acid sequence SIASRWGHTGYADSVKG. The VH may further comprise a CDR-H1 comprising the amino acid sequence FSPYHIH. In some embodiments, the antigen-binding domain may further comprise a light chain variable region (VL), which may comprise a CDR-L3 comprising the amino acid sequence QQASYVRKTIT. The VL may further comprise a CDR-L2 comprising the amino acid sequence SASSLYS. The VL may further comprise a CDR-L1 comprising the amino acid sequence RASQSVSSAVA. The antigen-binding domain can comprise a CDR-H3 sequence of GGWIAAMDY; a CDR-H2 sequence of SIASRWGHTGYADSVKG; a CDR-H1 sequence of FSPYHIH; a CDR-L3 sequence of QQASYVRKTIT; a CDR-L2 sequence of SASSLYS; and a CDR-L1 sequence of RASQSVSSAVA. The VH can comprise a sequence having at least 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the following sequence: EVQLVESGGGLVQPGGSLRLSCAASGFTFSPYHIHWVRQAPGKGLEWVASIASRWGHTGYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARGGWIAAMDYWGQGTLVTVSS The VH may comprise a sequence having at least 80% sequence identity to the following sequence: EVQLVESGGGLVQPGGSLRLSCAASGFTFSPYHIHWVRQAPGKGLEWVASIASRWGHTGYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARGGWIAAMDYWGQGTLVTVSS The VL can comprise a sequence having at least 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the following sequence: DIQMTQSPSSLSASVGDRVTITCRASQSVSSAVAWYQQKPGKAPKLLIYSASSLYSGVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQASYVRKTITFGQGTKVEIKRTV The VL may comprise a sequence having at least 80% sequence identity to the following sequence: DIQMTQSPSSLSASVGDRVTITCRASQSVSSAVAWYQQKPGKAPKLLIYSASSLYSGVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQASYVRKTITFGQGTKVEIKRTV

[0279] In some embodiments, the antigen-binding domain comprises a heavy chain variable region (VH), wherein the VH may comprise a heavy chain complementarity-determining region 3 (CDR-H3), wherein the CDR-H3 comprises the amino acid sequence GSWIAAMDY. The VH may further comprise a CDR-H2 comprising the amino acid sequence SIASLQGITGYADSVKG. The VH may further comprise a CDR-H1 comprising the amino acid sequence FHEYSIH. In some embodiments, the antigen-binding domain may further comprise a light chain variable region (VL), wherein the VL may comprise a CDR-L3 comprising the amino acid sequence QQASYVRKTIT. The VL may further comprise a CDR-L2 comprising the amino acid sequence SASSLYS. The VL may further comprise a CDR-L1 comprising the amino acid sequence RASQSVSSAVA. The antigen-binding domain can comprise the CDR-H3 sequence of GSWIAAMDY; the CDR-H2 sequence of SIASLQGITGYADSVKG; the CDR-H1 sequence of FHEYSIH; the CDR-L3 sequence of QQASYVRKTIT; the CDR-L2 sequence of SASSLYS; and the CDR-L1 sequence of RASQSVSSAVA. The VH can comprise a sequence having at least 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the following sequence: EVQLVESGGGLVQPGGSLRLSCAASGFTFHEYSIHWVRQAPGKGLEWVASIASLQGITGYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARGSWIAAMDYWGQGTLVTVSS The VH may comprise a sequence having at least 80% sequence identity to the following sequence: EVQLVESGGGLVQPGGSLRLSCAASGFTFHEYSIHWVRQAPGKGLEWVASIASLQGITGYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARGSWIAAMDYWGQGTLVTVSS The VL can comprise a sequence having at least 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the following sequence: DIQMTQSPSSLSASVGDRVTITCRASQSVSSAVAWYQQKPGKAPKLLIYSASSLYSGVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQASYVRKTITFGQGTKVEIKRTV The VL may comprise a sequence having at least 80% sequence identity to the following sequence: DIQMTQSPSSLSASVGDRVTITCRASQSVSSAVAWYQQKPGKAPKLLIYSASSLYSGVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQASYVRKTITFGQGTKVEIKRTV

[0280] In some embodiments, the antigen-binding domain comprises a heavy chain variable region (VH), which may comprise a heavy chain complementarity-determining region 3 (CDR-H3), which comprises the amino acid sequence GGVIHAMDY. The VH may further comprise a CDR-H2 comprising the amino acid sequence SILSRWGVTSYADSVKG. The VH may further comprise a CDR-H1 comprising the amino acid sequence FSDYSIH. In some embodiments, the antigen-binding domain may further comprise a light chain variable region (VL), which may comprise a CDR-L3 comprising the amino acid sequence QQASYVRKTIT. The VL may further comprise a CDR-L2 comprising the amino acid sequence SASSLYS. The VL may further comprise a CDR-L1 comprising the amino acid sequence RASQSVSSAVA. The antigen-binding domain can comprise the CDR-H3 sequence of GGVIHAMDY; the CDR-H2 sequence of SILSRWGVTSYADSVKG; the CDR-H1 sequence of FSDYSIH; the CDR-L3 sequence of QQASYVRKTIT; the CDR-L2 sequence of SASSLYS; and the CDR-L1 sequence of RASQSVSSAVA. The VH can comprise a sequence having at least 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the following sequence: EVQLVESGGGLVQPGGSLRLSCAASGFTFSDYSIHWVRQAPGKGLEWVASILSRWGVTSYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARGGVIHAMDYWGQGTLVTVSS The VH may comprise a sequence having at least 80% sequence identity to the following sequence: EVQLVESGGGLVQPGGSLRLSCAASGFTFSDYSIHWVRQAPGKGLEWVASILSRWGVTSYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARGGVIHAMDYWGQGTLVTVSS The VL can comprise a sequence having at least 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the following sequence: DIQMTQSPSSLSASVGDRVTITCRASQSVSSAVAWYQQKPGKAPKLLIYSASSLYSGVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQASYVRKTITFGQGTKVEIKRTV The VL may comprise a sequence having at least 80% sequence identity to the following sequence: DIQMTQSPSSLSASVGDRVTITCRASQSVSSAVAWYQQKPGKAPKLLIYSASSLYSGVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQASYVRKTITFGQGTKVEIKRTV

[0281] In some embodiments, the antigen-binding domain comprises a heavy chain variable region (VH), which may comprise a heavy chain complementarity-determining region 3 (CDR-H3), which comprises the amino acid sequence GGVIHAMDY. The VH may further comprise a CDR-H2 comprising the amino acid sequence SISSRWGVTSYADSVKG. The VH may further comprise a CDR-H1 comprising the amino acid sequence FSDYSIH. In some embodiments, the antigen-binding domain may further comprise a light chain variable region (VL), which may comprise a CDR-L3 comprising the amino acid sequence QQASYVRKTIT. The VL may further comprise a CDR-L2 comprising the amino acid sequence SASSLYS. The VL may further comprise a CDR-L1 comprising the amino acid sequence RASQSVSSAVA. The antigen-binding domain can comprise the CDR-H3 sequence of GGVIHAMDY; the CDR-H2 sequence of SISSRWGVTSYADSVKG; the CDR-H1 sequence of FSDYSIH; the CDR-L3 sequence of QQASYVRKTIT; the CDR-L2 sequence of SASSLYS; and the CDR-L1 sequence of RASQSVSSAVA. The VH can comprise a sequence having at least 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the following sequence: EVQLVESGGGLVQPGGSLRLSCAASGFTFSDYSIHWVRQAPGKGLEWVASISSRWGVTSYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARGGVIHAMDYWGQGTLVTVSS The VH may comprise a sequence having at least 80% sequence identity to the following sequence: EVQLVESGGGLVQPGGSLRLSCAASGFTFSDYSIHWVRQAPGKGLEWVASISSRWGVTSYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARGGVIHAMDYWGQGTLVTVSS The VL can comprise a sequence having at least 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the following sequence: DIQMTQSPSSLSASVGDRVTITCRASQSVSSAVAWYQQKPGKAPKLLIYSASSLYSGVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQASYVRKTITFGQGTKVEIKRTV The VL may comprise a sequence having at least 80% sequence identity to the following sequence: DIQMTQSPSSLSASVGDRVTITCRASQSVSSAVAWYQQKPGKAPKLLIYSASSLYSGVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQASYVRKTITFGQGTKVEIKRTV

[0282] The CDRs of the binding partners or antigen-binding domains described herein can be designated using the Kabat numbering scheme. In some cases, light chain (LC) CDRs can be designated using the Kabat numbering scheme. In some cases, LC CDRs can be designated using a modified Kabat numbering scheme. In some cases, heavy chain (HC) CDRs can be designated using the Kabat numbering scheme. In some cases, HC CDRs can be designated using a modified Kabat numbering scheme. For example, a CDR may contain one or more extra amino acids than the CDR designated using the Kabat numbering scheme.

[0283] In one aspect, provided herein is a binding partner that specifically binds to a conjugate formed by the covalent reaction of osimertinib with an HLA-presented EGFR peptide, wherein the binding partner comprises a heavy chain variable region (VH) and a light chain variable region (VL), wherein (a) VH comprises: (i) CDR-H1 containing the amino acid sequence SSYIH, or a variant thereof containing 1 to 5 amino acid changes; (ii) a CDR-H2 comprising the amino acid sequence YISPSYGSTSYADSVKG, or a variant thereof comprising 1 to 5 amino acid changes; and / or (iii) a CDR-H3 comprising the amino acid sequence EX1X2X3MX4X5DY, where X1 is Y, L, S, or E, X2 is V, T, or I, X3 is T or I, X4 is A, T, or S, and X5 is L, A, I, K, P, or T; and / or (b) VL includes: (i) CDR-L1 containing the amino acid sequence RASQSVSSAVA, or a variant thereof containing 1 to 5 amino acid changes; (ii) a CDR-L2 comprising the amino acid sequence SASSLYS, or a variant thereof comprising 1 to 5 amino acid changes; and / or (iii) A CDR-L3 comprising the amino acid sequence QQYX1X2WPX3T, where X1 is S or A or S; X2 is Y, H, A, D, E, K, S or G; and X3 is I or E.

[0284] The CDR sequences of exemplary antibodies that bind to the conjugate formed by the covalent reaction of osimertinib with an EGFR peptide presented on HLA are shown in Tables I and J below.

[0285] Table I: Heavy chain CDR sequences of exemplary antibodies that bind to conjugates formed by the covalent reaction of osimertinib with EGFR peptides presented on HLA. [Table I]

[0286] Table J: Light chain CDR sequences of exemplary antibodies that bind to conjugates formed by the covalent reaction of osimertinib with an EGFR peptide presented on HLA. [Table J]

[0287] In one embodiment, the binding partner comprises CDR-H1, CDR-H2, and CDR-H3 of the heavy chain variable region amino acid sequences and / or CDR-L1, CDR-L2, and CDR-L3 of the light chain variable region of the binding partner, wherein the binding partner is selected from the group consisting of OEA2-5, EO_Q01-EO_Q18, and EO_Q20-EO_Q24, or variants thereof comprising 1 to 5 amino acid changes in one or more of the CDR amino acid sequences.

[0288] In one embodiment, the binding partner comprises the CDR-H1, CDR-H2, CDR-H3, CDR-L1, CDR-L2 and / or CDR-L3 amino acid sequences of the binding partner, wherein the binding partner is selected from the group consisting of OEA2-5, EO_Q01-EO_Q18 and EO_Q20-EO_Q24 (Table I or J), or variants thereof comprising 1 to 5 amino acid changes in one or more of the CDR amino acid sequences.

[0289] In one embodiment, the binding partner may comprise a VH and / or VL amino acid sequence having at least about 40%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 100% sequence identity to the VH and / or VL amino acid sequence of an antibody disclosed in Table I and Table J.

[0290] A non-limiting example of a binding partner that binds to an epidermal growth factor receptor (EGFR)-osimertinib conjugate is referred to herein as OEA2-5. The present disclosure includes all derivatives of OEA2-5 described herein, which contain alternative residues as described below by deep mutational analysis, and are in the form of scDbs (representative amino acid sequences of which are shown). The light chain variable region of OEA2-5 (V L) and heavy chain variable region (V H ) has the following amino acid sequence: TIFF2025526240000038.tif36156

[0291] In embodiments, the binding partner binds to the protein in its native form, except for the covalent attachment of a drug or other molecule thereto. "Native form" refers to an intact protein (prior to the covalent attachment of a drug or other molecule) that retains its biological function. In embodiments, the native form or protein is the form prior to fragmentation, such as by intracellular processing. Thus, in embodiments, the binding protein binds to a full-length polypeptide that is covalently attached to a drug or other molecule, where the covalently attached drug or other molecule at least partially permits preferential binding of the binding partner. Generally, a polypeptide, which is used interchangeably herein with the term "protein," comprises more than 50 contiguous amino acids. In embodiments, the binding partner specifically binds to an intact protein that is covalently attached to a drug or other molecule. In other embodiments, the binding partner specifically binds to a peptide that includes the covalently attached molecule. In embodiments, the binding partner specifically binds to a peptide having a particular amino acid sequence and that is covalently attached to another molecule, such as a drug. In some embodiments, the binding partner preferentially binds to a peptide covalently bound to a molecule such as a drug, regardless of the sequence of the peptide. This preferential binding is compared to the binding to the same peptide that is not conjugated to a drug. In some embodiments, the binding partner preferentially binds to a peptide that contains a KRAS(G12) mutation, or its variant, where the variant is at least 50% similar to the KRAS(G12)-containing peptide. This preferential binding is compared to the binding to the KRAS(G12)-containing peptide or its variant that is not covalently bound to a drug or other molecule.

[0292] In some embodiments, the described binding partners specifically bind to peptide conjugates of a length suitable for presentation to the major histocompatibility complex (MHC), which in humans is called human leukocyte antigen (HLA) and in non-human animals (including but not limited to non-human mammals) is called MHC or equivalent complexes.

[0293] Generally, peptide conjugates contain fewer than 50 consecutive amino acids. Thus, in embodiments, peptide conjugates containing the described epitopes can be 2-49 amino acids in length. In embodiments, peptides to which drugs or other molecules are covalently attached, where one or more residues of the attached drug and peptide may be included in the epitope, contain 4-12 consecutive amino acids, which may or may not be derived from a longer protein during protein processing (e.g., an antigen processed for presentation by an MHC molecule). In embodiments, drugs are conjugated to peptides containing or consisting of 7-30 amino acids. In embodiments, drugs or other molecules are conjugated to peptides containing or consisting of 7, 8, 9, 10, 11, 12, 13, 14, or 15 amino acids and capable of being presented in the context of MHC class I. In several embodiments, a drug or other molecule is conjugated to a peptide consisting of 9 to 30 amino acids (including the upper and lower limits, and all numbers and ranges therebetween) and capable of being presented in the context of MHC class II. In several embodiments, a drug or other molecule is conjugated to a peptide comprising at least 7 amino acids. Non-classical MHC class I molecules function to mediate inhibitory or activating stimuli in natural killer (NK) cells. Non-classical MHC class I molecules can be expressed by immune cells and tumor cells. For example, expression of non-classical MHC complexes in malignant cells interferes with the cytotoxic activity of effector cells in the immune system. Overexpression of non-classical MHC class I molecules, including but not limited to HLA-E, HLA-F, and HLA-G, can be observed in cancer cells. In some embodiments, a drug or other molecule is conjugated to a peptide consisting of 9 to 30 amino acids (including the upper and lower limits, and all numbers and ranges therebetween) and capable of being presented in the context of non-classical MHC class I.In some embodiments, the peptide conjugate forms a complex with a non-classical MHC class I molecule. In some embodiments, the non-classical MHC class I molecule is selected from the group consisting of HLA-E, HLA-F, and HLA-G. In some embodiments, the non-classical MHC class I molecule is selected from HLA-E, HLA-F, HLA-G, or a combination thereof.

[0294] In some embodiments, the binding partner specifically binds to a peptide conjugate that is covalently conjugated to a drug or other molecule, regardless of MHC presentation. In some embodiments, a non-limiting example of which is described below in Example 3, the binding partner specifically binds to a peptide conjugate only when the peptide conjugate is presented by an MHC molecule. In some embodiments, the binding partner can specifically bind to a peptide conjugate in both MHC-independent and MHC-presented contexts. In some embodiments, the MHC-peptide conjugate complex comprises an antigen (the antigen to which the described binding partner specifically binds).

[0295] In some embodiments, the described binding partner exhibits at least one improved property compared to the same property of a reference binding partner. In some embodiments, the reference binding partner is AMRA3-7D. In one embodiment, the described binding partner exhibits higher affinity for its target compared to the reference binding partner.

[0296] In some embodiments, the binding partner can bind to cells via any MHC capable of presenting the peptide conjugate. In some embodiments, the HLA is expressed by cells capable of class I, class II, or class III MHC presentation. In some embodiments, the binding partner can bind to cells expressing class I MHC that presents the peptide conjugate. Those skilled in the art will recognize that class I MHC contains, among other components, a polymorphic α chain and β2 microglobulin, where the peptide conjugate binds to the polymorphic chain.

[0297] In some embodiments, the cells are antigen-presenting cells (APCs). In some embodiments, the cells are so-called professional antigen-presenting cells, and thus may include, but are not limited to, macrophages and dendritic cells that present class II MHC. Those skilled in the art will recognize that class II MHC comprises, among other components, the MHC polymorphic α and β chains, and the presented peptide conjugate binds to both chains. In other embodiments, class II MHC may be presented with peptide conjugates by other cell types, such as cancer / tumor cells; thus, the present disclosure provides for the direct recognition of such cells using the described binding partners, without the need for professional APCs.

[0298] In several embodiments, the peptide conjugates are presented by non-classical MHC complexes (which may include CD1d, MR1, MHC-E, -F, -G and / or other emerging family members that will be recognized by those skilled in the art).

[0299] The present disclosure provides binding partners that include those that specifically bind peptide conjugates presented only by specific MHC types, and thus discriminate between MHC types. Representative examples of such binding partners are described herein at least by Figure 17.

[0300] In some embodiments, the binding partners of the present disclosure can specifically bind to peptide conjugates containing covalently conjugated drugs or other molecules that are presented by two or more specific MHC types. In some embodiments, a single binding partner of the present disclosure is suitable for use with various HLA types. Furthermore, a single binding partner can specifically bind to multiple combinations of (a) peptide conjugates containing covalently conjugated drugs or other molecules and (b) HLA types, as shown in Figures 35 and 36. In particular, Figures 35 and 36 demonstrate that a single antibody can bind to different peptides conjugated to AMG-510 that are presented by different HLAs with different preferences for peptide binding. For example, comparing the HLA-A*03 / 11 peptides VVVGAC*GVGK and KLVVVGAC*GV, antibody RA_D11 still selectively binds to the hapten-peptide-HLA complex compared to binding to the free drug. In some embodiments, the binding partners of the present disclosure can specifically bind to a peptide conjugate comprising a covalently conjugated drug only when associated with a particular MHC. In some embodiments, the peptide conjugate is presented by an MHC class I type selected from HLA-A, -B, -C, and combinations thereof. In certain aspects, the peptide conjugate is presented in the context of any MHC class I type that is A*02 / B*35 / C*04. In some embodiments, the peptide conjugate is presented by any of the following MHC class II types: DR*01 / DR*04 / DR*07 / DP*04. In embodiments, the HLA comprises A*01:01, A*02:01, A*03:01, A*11:01, A*24:02, A*26:01, B*07:02, B*08:01, B*27:05, B*39:01, B*40:01, B*58:01 or B*15:01.Specific examples of antibodies include antibodies that bind to the KRAS(G12C)-AMG510 conjugate presented on HLA-A*02:01, HLA-A*03:01, and HLA-A*11:01, the BTK-ibrutinib conjugate presented on HLA-A*01:01, and the EGFR-osimertinib conjugate presented on HLA-A*02:01. In non-limiting embodiments, the present disclosure provides scDbs specific for a particular drug covalently bound to a described peptide present on a specific HLA, or for the same drug covalently bound to a described peptide present on two different HLAs. Representative scDbs are described in Example 4. Data obtained using the scDb are shown in Figures 18 and 19. Data obtained using the CrossMab format are shown in Example 5 and its associated figures.

[0301] In some embodiments, the present disclosure includes selecting an individual based on the individual's HLA type and selecting an antibody described herein to treat the individual. In some embodiments, the binding partner is selected based at least in part on the degree of HLA restriction that the selected binding partner exhibits with respect to the individual's HLA type.

[0302] In certain embodiments, such as KRAS(G12C) binding partners, a representative example is provided below, which bind to two different peptides derived from KRAS(G12C) conjugated to the drug (AMG-510, also known as sotorasib).

[0303] In embodiments, the peptide conjugate is presented by a cell that participates in or can be the target of a cell-mediated immune response. In embodiments, the peptide conjugate, presented in the context of any appropriate MHC, is presented by a cell that is recognized by a leukocyte (including, but not limited to, a T cell or a natural killer (NK) cell). In embodiments, the T cell is a CD4+ T cell, a CD8+ T cell, a double-positive CD4+ / CD8+ T cell, a CD4+ / CD8+ double-negative T cell, or a γδ T cell. Thus, as described further below, the present disclosure provides binding partners configured to interact with both the presented peptide conjugate and a cell involved in a cell-mediated immune response. In embodiments, a particular described binding partner is capable of binding to a complex of 1) a particular MHC and 2) a particular peptide conjugate. In embodiments, a particular described binding partner is capable of binding to a particular peptide conjugate presented by at least two different MHCs.

[0304] In embodiments, any binding partner of the present disclosure comprises at least one chain comprising a complementarity-determining region (CDR) that is a CDR1, CDR2, or CDR3 derived from any heavy or light chain amino acid sequence described herein. In certain examples herein, the CDRs are shown in bold. The amino acid sequences of the CDR sequences are separately encompassed in the present disclosure by their position in the described heavy and light chain amino acid sequences. The present disclosure includes binding partners comprising the described heavy chain CDR1, CDR2, and CDR3. The present disclosure also includes binding partners comprising the described light chain CDR1, CDR2, and CDR3. The present disclosure also includes binding partners comprising the described heavy chain CDR1, CDR2, and CDR3 and the described light chain CDR1, CDR2, and CDR3. For amino acid sequences of the present disclosure that include amino acids that comprise a purification or protein production tag (such as a HIS tag and / or an AVI tag), the present disclosure includes the proviso that the sequence of the described tag may be excluded from the amino acid sequence. Amino acids between the listed tags may also be excluded.

[0305] The binding partners of the present disclosure can be provided as intact immunoglobulins or as immunoglobulin fragments, including, but not necessarily limited to, antigen-binding (Fab) fragments, Fab' fragments, (Fab')2 fragments, Fd (N-terminal portion of the heavy chain) fragments, Fv fragments (two variable domains), diabodies (Db), dAb fragments, single domain fragments or single monomeric variable antibody domains, single-chain diabodies (scDb), isolated complementarity-determining regions (CDRs), single-chain variable fragments (scFv), and other antibody fragments that retain antigen-binding function. In several embodiments, the one or more binding partners are provided as components of a bispecific T cell engager (BiTE), a bispecific killer cell engager (BiKE), a CrossMab (e.g., a binding partner comprising four different chains; immunoglobulin crossover (also known as Fab domain exchange or CrossMab format) technology (see, e.g., WO2009 / 080253; Schaefer et al., Proc. Natl. Acad. Sci. USA, 108:11187-11192 (2011))), or a chimeric antigen receptor (CAR), e.g., for producing chimeric antigen receptor T cells (e.g., CAR T cells) and CAR natural killer (NK) cells, neutrophils, and macrophages. The present disclosure includes binding partners comprising the described heavy and light chain variable regions.

[0306] The binding partner of the present disclosure can be an scFv. In the present disclosure, the VH of the polypeptide can be linked to the VL via a linker. The linker can be a peptide linker consisting of amino acids such as glycine and / or serine residues, used alone or in combination, to link the variable heavy chain region and the variable light chain region together. In one embodiment, the flexible polypeptide linker is a Gly / Ser linker and comprises the amino acid sequence (Gly-Gly-Gly-Ser)n, where n is a positive integer equal to or greater than 1. For example, n=1, n=2, n=3, n=4, n=5, n=6, n=7, n=8, n=9, and n=10. In one embodiment, the flexible polypeptide linker includes, but is not limited to, (Gly4Ser)4 or (Gly4Ser)3. In another embodiment, the linker includes multiple repeats of (Gly2Ser), (GlySer) or (Gly3Ser). In some cases, the linker sequence is (G4S) n where n=2 to 4. In some cases, the linker sequence comprises (G4S) n where n=1-3. In some cases, the linker comprises (G4S) n or (S4G) n where n is an integer from 1 to 10. In some cases, the linker comprises the glycine-serine-alanine linker G4SA 3、 Alternatively, it may contain a glycine-serine linker (G4S)4.

[0307] The binding partner of the present disclosure can be a bispecific T cell engager (BiTE). BiTE therapy can be used to connect a subject's endogenous T cells to cancer cells. The BiTE molecule can comprise two Fv fragments from a monoclonal antibody linked by a peptide linker. The BiTE molecule can comprise a first antigen-binding domain and a second antigen-binding domain. The first antigen-binding domain can be specific for and capable of binding to a T cell antigen. The second antigen-binding domain can bind to a tumor antigen (e.g., a tumor-associated antigen) expressed on the surface of cancer cells. In some embodiments, the BiTE molecule can specifically bind to a peptide conjugate / MHC complex and a T cell surface antigen. In some embodiments, the T cell surface antigen is CD3 (e.g., CD3 epsilon, CD3 delta, or CD3 gamma), TCR alpha chain, TCR beta chain, TCR gamma chain, TCR delta chain, ICOS, CD28, CD27, HVEM, LIGHT, CD40, 4-1BB, OX40, DR3, GITR, CD30, TIM1, SLAM, CD2, CD4, CD8, or CD226. In some embodiments, the tumor-associated antigen comprises a peptide conjugate formed by the covalent reaction of a targeted covalent inhibitor with a peptide in tumor cells. In some embodiments, the tumor-associated antigen comprises a peptide that is a segment of a protein associated with cancer, optionally encoded by a gene mutated in cancer. In some embodiments, the protein is overexpressed in cancer. In some embodiments, a segment of the protein is overexpressed in cancer. Cancer-testis antigens are a type of tumor-associated antigen expressed in human tumors. In some embodiments, the protein is a member of the cancer-testis antigen (CTA) family. In some embodiments, the cancer-associated protein comprises an endogenous retrovirus (ERV). In some embodiments, the protein is derived from a retrotransposon family, such as a long interspersed nucleotide sequence (LINE) or a short interspersed nucleotide sequence (SINE).In some embodiments, the tumor-associated antigen is an antigen associated with renal cell carcinoma. In some embodiments, the tumor-associated antigen is an antigen associated with breast cancer. In some embodiments, the tumor-associated antigen is an antigen associated with prostate cancer. In some embodiments, the tumor-associated antigen is an antigen associated with pancreatic cancer. In some embodiments, the tumor-associated antigen is an antigen associated with lung cancer. In some embodiments, the tumor-associated antigen is an antigen associated with liver cancer. In some embodiments, the tumor-associated antigen is an antigen associated with ovarian cancer. In some embodiments, the tumor-associated antigen is an antigen associated with cervical cancer. In some embodiments, the tumor-associated antigen is an antigen associated with colon cancer (or colorectal cancer). In some embodiments, the tumor-associated antigen is an antigen associated with esophageal cancer. In some embodiments, the tumor-associated antigen is an antigen associated with glioma. In some embodiments, the tumor-associated antigen is an antigen associated with glioblastoma or another brain tumor. In some embodiments, the tumor-associated antigen is an antigen associated with gastric cancer. In some embodiments, the tumor-associated antigen is an antigen associated with bladder cancer. In some embodiments, the tumor-associated antigen is an antigen associated with testicular cancer. In some embodiments, the tumor-associated antigen is an antigen associated with head and neck cancer. In some embodiments, the tumor-associated antigen is an antigen associated with melanoma or another skin cancer. In some embodiments, the tumor-associated antigen is an antigen associated with sarcoma, including but not limited to fibrosarcoma, angiosarcoma, osteosarcoma, and rhabdomyosarcoma. In some embodiments, the tumor-associated antigen is an antigen associated with any blood cancer, including all types of leukemia, lymphoma, and myeloma.

[0308] The binding partner can comprise a first antigen-binding domain that specifically binds to the peptide conjugate / MHC complex and a second antigen-binding domain that specifically binds to a T cell surface marker. The binding partner can comprise two polypeptide chains. For example, the polypeptide can comprise a first polypeptide chain comprising a first antigen-binding domain and a second polypeptide chain comprising a second antigen-binding domain. In some cases, the two polypeptide chains can comprise an Fc region. The binding partner of the present disclosure can be a bispecific or multivalent antibody or antibody fragment comprising a first antigen-binding domain that specifically binds to the peptide conjugate / MHC complex and a second antigen-binding domain that specifically binds to a T cell surface marker. In some embodiments, the bispecific or multivalent molecule can comprise an Fc region selected from the heavy chain constant regions of human IgM, IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2. The Fc region can be a modified version of a wild-type Fc region. For example, the Fc region may be a silenced version of the Fc region, including, but not limited to, LALA Fc, LALAPG Fc, LALAKA Fc, LAGA Fc, LGGR Fc, or LALE Fc. The Fc region may contain one or more mutations of a wild-type Fc region. The Fc region may be linked to one or more tumor-targeting moieties, T cell engagers, or cytokine molecules. In some embodiments, the interface between the first and second Fc regions may be altered to increase or decrease dimerization. Dimerization of the Fc region may be enhanced by providing paired-cavity protuberances, such as knob-in-holes, at the Fc interface of the first and second Fc regions.Knob-in-hole synthesis, as described in U.S. Pat. No. 5,731,116, U.S. Pat. No. 7,476,724, and Ridgway, J. et al. (1996) Prot. Engineering 9(7): 617-621, generally involves (1) mutating the CH3 domain of one or both antibodies to promote heterodimerization, and (2) binding the mutated antibodies under conditions that promote heterodimerization. A "knob" or "protrusion" can be created by replacing a small amino acid in the parent antibody with a larger one (e.g., T366Y or T366W). A "hole" or "cavity" can be created by replacing a large residue in the parent antibody with a smaller one (e.g., Y407T, T366S, L368A, and / or Y407V). Exemplary knobs-in-hole mutations include S354C or T366W in the "knob" heavy chain and Y349C, T366S, L368A, or Y407V in the "hole" heavy chain. In bispecific antibodies containing an Fc domain, specific mutations can be introduced into the constant region of the heavy chain to promote correct heterodimerization of the Fc portion. Such techniques include the knobs-in-hole approach, which introduces bulky residues into one of the CH3 domains of one of the antibody heavy chains. These bulky residues fit into a complementary "hole" in the CH3 domain of the other paired heavy chain to promote correct pairing of the heavy chains (see, e.g., US7642228). In some cases, two polypeptide chains can be combined using the knobs-in-hole approach described herein. In some cases, two polypeptide chains cannot be combined using the knobs-in-hole approach. In some cases, the first antigen-binding domain and the second antigen-binding domain are connected by a linker. The linker may be a peptide linker composed of amino acids (eg, glycine and / or serine residues used alone or in combination).The linker can comprise at least about 1, 2, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 150, 200, 250 or more amino acid residues. In some cases, the linker is a nucleotide sequence similar to (G4S). n (n is any integer from 1 to 10). In one embodiment, the flexible polypeptide linker is a Gly / Ser linker and comprises the amino acid sequence (Gly-Gly-Gly-Ser)n, where n is a positive integer equal to or greater than 1. For example, n=1, n=2, n=3, n=4, n=5, n=6, n=7, n=8, n=9, and n=10. In one embodiment, the flexible polypeptide linker comprises, but is not limited to, (Gly4Ser)4 or (Gly4Ser)3. In another embodiment, the linker comprises multiple repeats of (Gly2Ser), (GlySer) or (Gly3Ser). In some cases, the linker sequence is (G4S) n where n=2 to 4. In some cases, the linker sequence comprises (G4S) n where n=1-3. In some cases, the linker comprises (G4S) n or (S4G) n where n is any integer from 1 to 10. In some cases, the linker can include a glycine-serine-alanine linker G4SA3 or a glycine-serine linker (G4S)4.

[0309] The first or second polypeptide chain may be further fused to a cytokine or a fragment thereof. The cytokine may include IL-2, IL-7, IL-15, IL-12, IL-18, or IL-21 or an interferon (IFN). In some embodiments, the cytokine may be selected from the group consisting of IFNγ, IL-2, IL-4, IL-6, IL-7, IL-10, IL-12, IL-13, IL-15, IL-18, IL-23, IL-24, IL-27, IL-28a, IL-28b, IL-29, KGF, IFNα (e.g., IFNα2b), IFNβ, IFNλ, and GM-CSF. In some embodiments, the cytokine is an engineered cytokine. For example, the engineered cytokine may be a variant or mutant of any of the cytokines described herein. The artificial cytokine may be a mutein, superkine, or PEGylated cytokine. In some embodiments, the cytokine is a single-chain cytokine. In some embodiments, the cytokine is a multi-chain cytokine. In some embodiments, the two polypeptides may comprise two moieties. In some embodiments, the two polypeptide chains may comprise at least three moieties. In some cases, a first moiety can bind to a peptide conjugate / MHC complex, a second moiety can comprise a binding partner for a T cell surface antigen (e.g., a T cell engager), and a third moiety can comprise a cytokine or a fragment thereof. In some cases, the first polypeptide chain can comprise an antigen-binding domain for the peptide conjugate / MHC complex, and the second polypeptide chain can comprise an antigen-binding domain for the T cell surface antigen. In some cases, the first polypeptide chain can comprise an antigen-binding domain for the T cell surface antigen, and the second polypeptide chain can comprise an antigen-binding domain for the peptide conjugate / MHC complex.In some cases, the first polypeptide chain can comprise an antigen binding domain for the peptide conjugate / MHC complex and a cytokine or fragment thereof, and the second polypeptide chain can comprise an antigen binding domain for a T cell surface antigen. In some cases, the first polypeptide chain can comprise an antigen binding domain for the T cell surface antigen and a cytokine or fragment thereof, and the second polypeptide chain can comprise an antigen binding domain for the peptide conjugate / MHC complex. In some cases, the first polypeptide chain can comprise an antigen binding domain for the peptide conjugate / MHC complex, and the second polypeptide chain can comprise an antigen binding domain for the T cell surface antigen and a cytokine or fragment thereof. In some embodiments, the first polypeptide chain can comprise an antigen binding domain for a T cell surface marker, and the second polypeptide chain can comprise an antigen binding domain for the peptide conjugate / MHC complex and a cytokine or fragment thereof. In various embodiments, the antigen binding domain for the peptide conjugate / MHC complex and the antigen binding domain for the T cell surface antigen can be present on the same polypeptide chain, and the cytokine or fragment thereof is fused to a different polypeptide chain. In some cases, the antigen binding domain for the peptide conjugate / MHC complex and the cytokine or fragment thereof are on the same polypeptide chain, and the antigen binding domain for the T cell surface antigen are on different polypeptide chains. In some cases, the antigen binding domain for the T cell surface antigen and the cytokine or fragment thereof are on the same polypeptide chain, and the antigen binding domain for the peptide conjugate / MHC complex are on different polypeptide chains.

[0310] The polypeptide may be an intact antibody, a bispecific antibody, a multispecific antibody, an antigen-binding (Fab) fragment, a Fab' fragment, a (Fab')2 fragment, an Fd, an Fv, a dAb, a single domain fragment or a single monomeric variable antibody domain, a dual affinity retargeting (DART) molecule, a diabody (Db), a single-chain diabody (scDb), a single-chain variable fragment (scFv), a bispecific T-cell engager (BiTE), a bispecific killer cell engager (BiKE), a CrossMab, a camelid antibody, a trispecific binding partner, a chimeric antigen receptor (CAR), a monobody (also known as an adnectin), a DARPin, an anticalin, an affibody, a nanobody, or an affimer. In some embodiments, the nanobody is derived from the heavy chain variable domain of an antibody found in members of the Camelidae family (e.g., camel, llama, alpaca). In some cases, the polypeptide may be a bispecific antibody. The bispecific antibody may be a bispecific T-cell engager (BiTE). The polypeptide can comprise a first antigen-binding domain and a second antigen-binding domain. The second antigen-binding domain can bind to a T cell surface marker. The T cell surface marker can be CD3 epsilon, CD3 gamma, CD3 delta, TCR alpha, TCR beta, TCR gamma, TCR delta, ICOS, CD28, CD27, HVEM, LIGHT, CD40, 4-1BB, OX40, DR3, GITR, CD30, TIM1, SLAM, CD2, CD4, CD8, or CD226. In some cases, the antigen-binding domain and the second antigen-binding domain of the polypeptide can be linked by a linker. As discussed above, the linker can comprise at least about 2, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 150, 200, 250, or more amino acid residues. n or (S4G) n where n is any integer from 1 to 10.

[0311] In various cases, the linker connecting the antigen-binding domain and the second antigen-binding domain of the polypeptide can influence the angle between the axis of the MHC and the axis of the polypeptide.

[0312] The binding geometry of the TCR to a peptide-MHC complex can be used as a descriptor of binding affinity. Diagonal binding of the TCR to the MHC complex can be quantified using the crossing angle, e.g., the angle between the vector between the MHC pocket and the TCR domain. This angle corresponds to the torsion of the receptor with respect to the peptide-MHC complex. The incidence angle refers to the angle between the normal vector of the MHC peptide-binding groove and the axis of rotation between the TCR domains. This angle corresponds to the tilt of the TCR with respect to the peptide-MHC complex and is usually measured along the XY axis. The binding geometries between the MHC complexes and polypeptides disclosed herein can also be measured, and methods for quantifying these binding geometries are known to those of skill in the art (Rudolph et al. How TCRs bind MHCs, peptides, and coreceptors. Annu Rev Immunol. 2006;24:419-66; Singh et al. Geometrical characterization of T cell receptor binding modes reveals class-specific binding to maximize access to antigen. Proteins. 2020 Mar;88(3):503-513).

[0313] The binding geometry between a peptide conjugate / MHC complex and a binding partner can be characterized using the incidence angle measured along the XY axis and / or the back angle measured along the YZ axis. In some embodiments, the binding geometry between a polypeptide and a peptide conjugate / MHC complex can be given as the incidence angle and / or the back angle. In some embodiments, a polypeptide can bind to a peptide conjugate / MHC complex at a back angle between the axis of the peptide conjugate / MHC complex and the axis of the polypeptide of about 10° to about 70° (e.g., about 10°, about 20°, about 30°, about 40°, about 50°, about 60°, or about 70°). In some embodiments, a polypeptide can bind to a peptide conjugate / MHC complex at a back angle between the axis of the peptide conjugate / MHC complex and the axis of the polypeptide of about 60° to about 65°. In some embodiments, the polypeptide is capable of binding to the peptide conjugate / MHC complex at a back angle of about 64.7°, 67.2°, or 65.1° between the axis of the peptide conjugate / MHC complex and the axis of the polypeptide.

[0314] In some embodiments, a polypeptide can bind to a peptide conjugate / MHC complex at an angle of incidence between about 10° and about 100° (e.g., about 10°, about 20°, about 30°, about 40°, about 50°, about 60°, about 70°, about 80°, about 90°, or about 100°) between the axis of the peptide conjugate / MHC complex and the axis of the polypeptide. In some embodiments, a polypeptide can bind to a peptide conjugate / MHC complex at an angle of incidence between 70° and 80° between the axis of the peptide conjugate / MHC complex and the axis of the polypeptide. In some embodiments, a polypeptide can bind to a peptide conjugate / MHC complex at an angle of incidence between the axis of the peptide conjugate / MHC complex and the axis of the polypeptide of about 77.2°, 77.3°, or 78.4°.

[0315] In various embodiments described herein, the peptide conjugate / MHC complex can be presented on the surface of a cell. The cell can express a low copy number of the peptide conjugate / MHC complex, where the low copy number is at most about 1000, 900, 800, 700, 600, 500, 400, 300, 200, 100, 90, 80, 70, 60, 50, 40, 30, 20, 10, 5, 3, or 1 copy per single cell. The peptide of the peptide conjugate / MHC complex can be derived from an intracellular protein. For example, the peptide can be processed from an endogenous or intracellular protein and presented on the MHC complex.

[0316] The binding partners described herein can be T cell receptors (TCRs) or functional fragments thereof (e.g., antigen-binding fragments, variable regions, or extracellular domains of TCRs). The binding partners described herein can be TCR mimetic binders. The T cell receptors or functional fragments thereof can specifically bind to the peptide conjugate / MHC complexes described herein. In some cases, the K of the TCR mimetic binder for the peptide conjugate / MHC complex is D may be less than about 1 μM, less than about 500 nM, less than about 400 nM, less than about 300 nM, less than about 200 nM, less than about 100 nM, less than about 50 nM, less than about 40 nM, less than about 30 nM, less than about 20 nM, less than about 10 nM, less than about 5 nM, less than about 1 nM, less than about 0.1 nM, less than about 50 pM, less than about 20 pM, less than about 10 pM, less than about 1 pM, less than about 0.1 pM, or even less. In some cases, the K of the TCR for the peptide conjugate / MHC complex Dmay be less than about 1 μM, less than about 500 nM, less than about 400 nM, less than about 300 nM, less than about 200 nM, less than about 100 nM, less than about 50 nM, less than about 40 nM, less than about 30 nM, less than about 20 nM, less than about 10 nM, less than about 5 nM, less than about 1 nM, or less than about 0.1 nM, less than about 50 pM, less than about 20 pM, less than about 10 pM, less than about 1 pM, less than about 0.1 pM, or less. In one aspect, the present disclosure provides a method for identifying a T cell receptor (TCR) that recognizes a peptide conjugate / MHC complex described herein. In some embodiments, the method of identifying a TCR includes contacting a plurality of candidate TCRs with the peptide conjugate / MHC complex and identifying at least one TCR that binds to the peptide conjugate / MHC complex. In some embodiments, the method for identifying a TCR that recognizes a peptide conjugate / MHC complex described herein includes selecting or isolating at least one TCR. Isolation of the TCR can be achieved by analyzing binding affinity to the peptide conjugate / MHC complex. In some embodiments, the peptide-drug conjugate can be a vaccine that enhances TCR isolation. In some embodiments, the vaccine can be used to enhance G12C-inhibition therapy. In some embodiments, the vaccine can be an RNA vaccine. For example, the vaccine can be an RNA vaccine encoding a KRAS peptide comprising a G12C mutation. In some embodiments, the vaccine can be an RNA vaccine encoding a KRAS peptide comprising a G12C mutation that is co-administered with a KRAS peptide comprising a G12C mutation. In some embodiments, the vaccine can be used to enhance, for example, G12C-inhibition therapy or HapImmune TM Responsiveness can be enhanced in patients treated with antibodies.

[0317] In some examples, the plurality of candidate TCRs are a plurality of soluble TCRs. In certain embodiments, the plurality of candidate TCRs are a plurality of TCRs expressed on the cell surface of a plurality of cells. In some embodiments, the method for identifying a TCR that recognizes a peptide conjugate / MHC complex described herein comprises isolating or selecting cells comprising at least one TCR based on a cellular activation marker. In some embodiments, the activation marker is a T cell marker. In some embodiments, the T cell activation marker is CD26, CD27, CD28, CD30, CD154, CD40L, CD134, CD25, CD44, CD69, CD137, or KLRG1. In another aspect, the present disclosure provides a TCR comprising at least one identified T cell receptor (TCR) that recognizes a peptide conjugate / MHC complex described herein. In some embodiments, the TCR is a soluble TCR. In some embodiments, the TCR is a bispecific TCR. In some embodiments, the TCR is expressed on CD4+ T cells. In some embodiments, the TCR is expressed on a CD8+ T cell.

[0318] In some embodiments, the binding partner is multivalent. In some embodiments, a trispecific binding partner is provided. In some embodiments, the cell expresses at least one segment of one or more binding partners in the form of a CAR. In some embodiments, the binding partner of the present disclosure can be provided as a complex with a polynucleotide (such as an RNA polynucleotide) to form an aptamer. In some embodiments, the multivalent binding partner comprises one binding moiety (such as a paratope) that confers specificity for a particular target on a desired cell type, such as any cancer cell marker. In some embodiments, a trispecific leukocyte engager is provided. In embodiments, the binding partner may be a portion of a molecule that is activated only in the presence of a protease or other enzyme present in the tumor microenvironment; such embodiments relate, for example, to probodies, examples of which are known in the art and described, for example, in doi: 10.1126 / scitranslmed.3006682, doi: 10.1038 / s41467-020-16838-w and doi: 10.1038 / s41587-019-0135-x, the descriptions of which are incorporated herein by reference. In one embodiment, the present disclosure provides a universal hapten that can be grafted onto an inhibitor.

[0319] In some embodiments, the CAR of the present disclosure comprises an scFv comprising a heavy chain and a light chain variable region as described herein. As known in the art for previously described CARs, the scFv is present in a contiguous polypeptide further comprising a CD3 zeta chain and a costimulatory domain. In some embodiments, the contiguous polypeptide further comprises a CD3 gamma chain or a CD3 epsilon chain. In some embodiments, the costimulatory domain comprises a 4-1BB costimulatory domain or a CD28 costimulatory domain. The CAR may also comprise a co-receptor hinge sequence (e.g., a C8 co-receptor hinge sequence).

[0320] In some embodiments, the binding partner of the present disclosure may comprise a constant region, such as an Fc region. Any isotype of constant region may be included. Binding partners comprising a constant region may be specifically adapted for antibody-dependent cell-mediated cytotoxicity (ADCC) or antibody-dependent cellular phagocytosis (ADCP), and thus may function to kill target cells through a cell-mediated response via any of a variety of effector cells. Similarly, the constant region may be specifically adapted to enhance complement-mediated responses.

[0321] In embodiments, the binding partners of the present disclosure may be modified to be present in a fusion protein. In embodiments, the antigen-binding segment of the binding partner may be present in the fusion protein and / or the constant region may be a component of the fusion protein. In embodiments, the fusion protein comprises amino acids from at least two different proteins. Fusion proteins may be produced using any of a variety of standard molecular biology approaches, including, but not limited to, expression from any suitable expression vector. In embodiments, the binding partners described herein may be present in a fusion protein with a detectable protein (e.g., green fluorescent protein (GFP), enhanced GFP (eGFP), mCherry, etc.). In embodiments, as an alternative to an expression vector, mRNA or chemically modified mRNA encoding any of the binding partners described herein may be delivered to a cell so that the binding partner is translated by the cell. In embodiments, the fusion protein comprises a binding partner disclosed herein and a cytokine. In embodiments, the cytokine is IL-2, IL-7, IL-15, IL-12, IL-18, or IL-21. In embodiments, the cytokine is modified to increase at least one therapeutic property, including, but not limited to, bioavailability, efficacy, increased half-life, or other desirable properties. Examples of suitable cytokine modifications are described in "Front. Immunol., 14 October 2021, doi.org / 10.3389 / fimmu.2021," the disclosure of which is incorporated herein by reference.

[0322] In some embodiments, a binding partner of the present disclosure may be fused to another antibody, for example, a binding partner of the present disclosure may be fused to an antibody or fragment thereof that is an immune checkpoint inhibitor. In some embodiments, the antibody or fragment thereof that is an immune checkpoint inhibitor may be an anti-PD-1 antibody, an antagonistic anti-PD-L1 antibody, an antagonistic anti-PD-L2 antibody, an antagonistic anti-CTLA-4 antibody, an antagonistic anti-BTLA antibody, an antagonistic anti-TREMR antibody, an antagonistic anti-TIGIT antibody, an antagonistic anti-VISTA antibody, an antagonistic anti-TIM-3 antibody, an antagonistic anti-LAG-3 antibody, an antagonistic anti-CEACAM1 antibody, an agonist anti-GITR antibody, an agonist anti-OX40 antibody, and an agonist anti-CD137 antibody, an agonist anti-DR3 antibody, an agonist anti-TNFSF14 antibody, an agonist anti-CD27 antibody, an agonist anti-ICOS antibody, or an agonist anti-CD28 antibody.

[0323] In embodiments, the binding partners described herein are used to deliver drugs or toxins, and thus the binding partners may be provided as immunotoxins or in the form of antibody-drug conjugates (ADCs).

[0324] In embodiments, agents useful for generating immunotoxins include enzymatically active toxins and enzymatically active fragments thereof. Suitable enzymatically active toxins include, but are not limited to, diphtheria A chain, nonbinding active fragments of diphtheria toxin, exotoxin A chain (from Pseudomonas aeruginosa), ricin A chain, abrin A chain, modeccin A chain, alpha-sarcin, Aleurites fordii protein, dianthin protein, Phytolacca americana proteins (PAPI, PAPII, and PAP-S), Momordica charantia inhibitor, curcin, crotin, soapwort (sapaonaria) officinalis inhibitors, gelonin, mitogellin, restrictocin, phenomycin, enomycin, and trichothecenes, which can be provided as components of a fusion protein or can be covalently attached to a binding partner by any suitable conjugation approach.

[0325] The binding partner can be linked to the chemotherapeutic agent using any suitable linker to form an antibody-drug conjugate (ADC). In some embodiments, the linker comprises a disulfide, hydrazine, or thioether. In some embodiments, the ADC comprises two payloads (e.g., two different chemotherapeutic agents), each linked to the binding partner by a linker. In some embodiments, the ADC comprises two payloads (e.g., two different chemotherapeutic agents), each linked to the binding partner by a linker using a site-specific aldehyde tag. The chemotherapeutic agent can be reversibly or irreversibly attached to the binding partner.

[0326] Cleavable linkers may be particularly useful for killing bystander cells. In some embodiments, a protease recognition site may be included to release the chemotherapeutic agent from the binding partner by the action of a protease that recognizes and cleaves the protease recognition site. Thus, ADCs are considered to include prodrugs.

[0327] In some embodiments, the binding partner of the present disclosure may include a linking sequence. As a non-limiting example, an ScFv may include a linker connecting the paratope-containing segments. Suitable amino acid linkers may be composed primarily of relatively small, neutral amino acids, such as glycine, serine, and alanine, and may include multiple copies of glycine- and serine-rich sequences. In specific, non-limiting embodiments, the linker includes 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 amino acids. In one example, the linker may be a glycine-serine-alanine linker G4SA3 or a glycine-serine linker (G4S)4. In some embodiments, the binding partner may include a cellular localization signal or a secretion signal. In some embodiments, the binding partner may comprise a transmembrane domain and thus be transported to and anchored in the cell membrane. For secretion, any suitable secretion signal may be used, many of which are known in the art.

[0328] In some embodiments, the binding partner can be part of an ADC and thus includes a drug. The drug can include, but is not limited to, any suitable chemotherapeutic agent. In some embodiments, the ADC includes a binding partner and a chemotherapeutic agent, where the chemotherapeutic agent is a microtubule inhibitor, an alkylating agent, or a DNA minor groove binding agent. In some embodiments, the chemotherapeutic agent includes a maytansinoid, a dolastatin, an auristatin drug analog, or a cryptophycin. In some embodiments, the chemotherapeutic agent is a duocarmycin derivative, or an antibiotic, such as an enediyne antibiotic, or a pyrolobenodiazepine (PBD), including dimers thereof. In some embodiments, the chemotherapeutic agent is an enzyme inhibitor, such as a topoisomerase or polymerase inhibitor. In embodiments, the chemotherapeutic agent comprises doxorubicin or a metal-containing compound such as a platinum-containing compound, non-limiting examples of which include cisplatin, carboplatin, or oxaliplatin. In some embodiments, the ADC comprises a binding partner described herein and a drug described in Barf and Kaptein, dx.doi.org / 10.1021 / jm3003203, J. Med. Chem. 2012, 55, 6243-6262; Wilson et al., dx.doi.org / 10.1021 / jm400224q, J. Med. Chem. 2013, 56, 7463-7476; Lambert and Morris, Adv Ther (2017) 34:1015-1035; and Tarantino and Tolaney, Cancer Res (2022) 82 (20): 3659-3661, the descriptions of which regarding drugs used as components of ADCs are incorporated herein by reference.In embodiments, the binding partner is conjugated to or includes a cytokine, thereby providing a cytokine conjugate, where the cytokine includes, but is not necessarily limited to, an interleukin (including, but not limited to, IL-2, IL-7, IL-8, IL-15, IL-12, IL-18, or IL-21) or an interferon (IFN). In embodiments, the binding partner includes a toxin conjugate. The toxin conjugate may include, but is not necessarily limited to, Pseudomonas exotoxin A (PE), diphtheria toxin (DT), or recombinant variants thereof. In some embodiments, the binding partner may include an immunotoxin agent derived from PE or DT (including, but not limited to, BL22, LMB-2, CAT-8015, SS1P, MR1-1, or Zemab).

[0329] Any suitable expression system can be used to produce the binding partner. Generally, a polynucleotide encoding the binding partner is used to express the binding partner in any suitable cell system, non-limiting examples of which include NS0 mouse myeloma cells, human cell lines, and Chinese hamster ovary (CHO) cells. In embodiments, the present disclosure provides polynucleotides that can selectively hybridize to polynucleotides encoding any one or combination of CDRs described herein. In embodiments, the polynucleotides selectively hybridize to polynucleotides encoding the heavy chain CDR1, CDR2, and CDR3 of any described binding partner. In embodiments, the polynucleotides selectively hybridize to polynucleotides encoding the light chain CDR1, CDR2, and CDR3 of any described binding partner. In embodiments, the polynucleotides selectively hybridize to polynucleotides encoding the heavy and light chain CDR1, CDR2, and CDR3 of any described binding partner.

[0330] In some embodiments, the binding partners described herein may be components of a fusion protein. In some embodiments, for example, for binding partners produced as fusion proteins, a peptide linker may be used. In some embodiments, the peptide linker includes an optional self-cleaving signal. In some embodiments, the self-cleaving signal may be present in the same open reading frame (ORF) as the binding partner. The self-cleaving amino acid sequence is typically about 18-22 amino acids in length. Any suitable sequence may be used, non-limiting examples of which include: T2A (EGRGSLLTCGDVEENPGP); P2A (ATNFSLKQAGDVENPGP); E2A (QCTNYALKLAGDVESNPGP) and F2A (VKQTLNFDLKLAGDVESNPGP).

[0331] To the extent that any segment of a protein comprising a binding partner described herein is a component of a library, including, but not necessarily limited to, a phage display library or a yeast surface display library, the present disclosure includes the proviso that the binding partner may not include any segment of the library (including bacteriophage or yeast amino acid sequences, including, but not limited to, a phage coat protein or a yeast host protein (including, but not limited to, Aga2)). Thus, in certain embodiments, the binding partner may be present in a fusion protein, but the fusion protein does not include a bacteriophage coat protein. In embodiments, any binding partner described herein may not include any of the pIII phage coat proteins, or any of the M1, fd filamentous phage, T4, T7, or λ phage proteins.

[0332] In some embodiments, the binding partners of the present disclosure include a detectable label, which can be used for diagnostic or therapeutic purposes. For example, the detectable label can be used to localize the binding partner for pathology and / or in vivo imaging approaches. In some embodiments, the binding partner is conjugated to any of a variety of radioactive agents, including, but not limited to, highly radioactive atoms such as In111, At211, I131, I125, Y90, Re186, Re188, Sm153, Bi212, P32, Pb212, and radioactive isotopes of Lu. In certain embodiments, such as for imaging, the binding partner may be conjugated to a radioactive atom for scintigraphic approaches, such as Tc99m (metastable technetium-99), 1123, or to a spin label for nuclear magnetic resonance (NMR) imaging (also known as magnetic resonance imaging, or "MRI"), such as 1123, 1131, 1124, F19, C13, N15, O17, or gadolinium(III) or manganese(II). In some embodiments, the radiopharmaceutical is suitable for use in CAT scans or PET imaging. In some embodiments, indium-111, technetium-99, or iodine-131 can be used for planar scans or single photon emission computed tomography (SPECT). Positron-emitting labels, such as fluorine-19, iodine-123, and iodine-124, can be used for positron emission tomography. Paramagnetic ions such as gadolinium (III) or manganese (II) can be used in magnetic resonance imaging (MRI). In embodiments, the described radioisotopes attached to the described binding partners can also be used in therapeutic approaches. In embodiments, the radiopharmaceutical or isotope comprises an alpha-emitting radionuclide. In embodiments, the radiopharmaceutical or isotope comprises a beta-emitting radionuclide. In some embodiments, the present disclosure provides an antibody of the present technology conjugated to a diagnostic or therapeutic agent.Diagnostic agents can include non-radioactive labels, contrast agents (e.g., for magnetic resonance imaging, computed tomography, ultrasound), and / or radioactive labels which can be gamma-, beta-, alpha-, Auger electron-, or positron-emitting isotopes. Diagnostic agents are molecules conjugated to antibody moieties (i.e., antibodies, antibody fragments, or antibody subfragments) and administered, which are useful in diagnosing or detecting disease by locating cells containing antigens.

[0333] The binding partner may be a polypeptide. The polypeptide is capable of binding to the peptide conjugate / MHC complex. The three-dimensional structure of the polypeptide that binds to the peptide conjugate / MHC complex can be determined by various methods, such as crystallography, cryoEM, or NMR. The polypeptide may have a three-dimensional structure of at least about 500, 600, 700, 800, 900, 1,000, 1,200, 1,500, 2,000, or more. 2The polypeptide can contact the peptide conjugate / MHC complex with an interfacial area of 1000 nm. The surface area of residues buried upon interaction between the polypeptide and the peptide conjugate / MHC complex can be calculated using PDBePISA. The polypeptide can contact the MHC of the peptide conjugate / MHC complex with an interfacial area greater than the interfacial area of the peptide and the targeted covalent inhibitor. The polypeptide can form a binding pocket at the interface between the VH domain and the VL domain to accommodate the targeted covalent inhibitor of the peptide conjugate / MHC complex. The polypeptide may not bind to the peptide conjugate / MHC complex via a head-to-head coaxial interaction. In some cases, the polypeptide may bind to the peptide conjugate / MHC complex via a head-to-head coaxial interaction. For example, the polypeptide may bind to the peptide conjugate / MHC complex in a manner similar to that of a TCR binding to the peptide / MHC complex. The polypeptide may bind to the peptide conjugate / MHC complex at an angle between the axis of the MHC and the axis of the polypeptide of about 10° to 60°, about 10° to 70°, or about 10° to 80°. The polypeptide may bind to the peptide conjugate / MHC complex at an angle between the axis of the MHC and the axis of the polypeptide of about 40°.

[0334] The polypeptide can contact the α1 and α2 domains of the heavy chain of MHC. The polypeptide can bind to an epitope of MHC. The epitope can include one or more residues from the region including residues 62-66, 106-109, and / or 150-170 of MHC. The polypeptide can bind to an epitope of MHC, and the epitope can include one or more residues from the region including residues 62-66, 106-109, and / or 150-170 of HLA-A*03:01 or HLA-A*11:01. The polypeptide can bind to an epitope of MHC, the epitope can comprise one or more residues selected from the group consisting of residues 62, 106, 108, 109, 158, 161, 162, 162, 165, 166, 167, 169 and 170 of HLA-A*03:01. The polypeptide can bind to an epitope of MHC, the epitope can comprise one or more residues selected from the group consisting of residues 62, 106, 108, 109, 158, 161, 162, 162, 165, 166, 167, 169 and 170 of HLA-A*03:01. The polypeptide can bind to an epitope of MHC, the epitope can comprise one or more residues selected from the group consisting of residues 62, 65, 66, 150, 151, 154, 155, 157 and 158 of HLA-A*03:01. The polypeptide can bind to an epitope of MHC, the epitope can comprise one or more residues selected from the group consisting of residues 62, 65, 66, 150, 151, 154, 155, 157 and 158 of HLA-A*03:01. The polypeptide can bind to an epitope of an MHC, and the epitope can include one or more residues selected from the group consisting of residues 62, 106, 108, 109, 154, 157, 158, 161, 162, 163, 165, 166, 167, 169 and 170 of HLA-A*11:01.The polypeptide can bind to an epitope of MHC, the epitope can comprise one or more residues selected from the group consisting of residues 62, 106, 108, 109, 154, 157, 158, 161, 162, 163, 165, 166, 167, 169 and 170 of HLA-A*11:01. The polypeptide can bind to an epitope of MHC, the epitope can comprise one or more residues selected from the group consisting of residues 62, 65, 66, 151, 154, 155 and 158 of HLA-A*11:01. The polypeptide can bind to an epitope of an MHC, and the epitope can include one or more residues selected from the group consisting of residues 62, 65, 66, 151, 154, 155 and 158 of HLA-A*11:01.

[0335] In some embodiments, the present disclosure provides methods for identifying or designing polypeptides capable of binding to peptide-conjugate / MHC complexes based on the three-dimensional structure(s) of the peptide-conjugate / MHC complex. The three-dimensional structure of the peptide-conjugate / MHC complex can be obtained from an existing complex structure of a binding partner that binds to the peptide-conjugate / MHC complex (e.g., a structure provided herein). Computer-assisted methods can be used to identify or design polypeptides capable of linking to a target covalent inhibitor, or a fragment thereof, that is covalently linked to a peptide of the peptide conjugate / MHC complex. In some embodiments, the method for identifying or designing candidate polypeptides capable of linking to a target covalent inhibitor comprises: (a) (i) providing coordinates of at least two atoms of the peptide conjugate / MHC complex of Figures 57A-C, 58A-B, 59A-C, 60A-B, and 61; and (b) providing a structure of a candidate polypeptide comprising an antigen-binding domain for binding to the peptide conjugate / MHC complex. The method may further include (c) fitting the structure of the candidate polypeptide to at least two atoms of the peptide conjugate / MHC complex. In some cases, the fitting includes determining interactions between one or more atoms of the antigen-binding domain of the candidate polypeptide and atoms of the peptide conjugate / MHC complex. The method may further include (d) selecting the candidate polypeptide if the candidate polypeptide is predicted to bind to the peptide conjugate / MHC complex.

[0336] Steps (a) and (b) in the above computer-assisted method for designing a polypeptide capable of linking to a target covalent inhibitor may be performed in any order or simultaneously. For example, in some embodiments, a computer-assisted method for designing a polypeptide capable of linking to a target covalent inhibitor comprises the following steps: (a) providing a structure of a candidate polypeptide comprising an antigen-binding domain for binding to a peptide conjugate / MHC complex; (b) providing coordinates of at least two atoms of the peptide conjugate / MHC complex and coordinates of at least two atoms of the antigen-binding domain of a polypeptide that binds to the peptide conjugate / MHC complex (Figures 57A-C, 58A-B, 59A-C, 60A-B and 61); (c) fitting the structure of the candidate polypeptide to at least two atoms of the peptide conjugate / MHC complex and at least two atoms of the antigen-binding domain of the polypeptide that binds to the peptide conjugate / MHC complex, wherein fitting comprises determining interactions between one or more atoms of the antigen-binding domain of the candidate polypeptide and atoms of the peptide conjugate / MHC complex; and (d) selecting the candidate polypeptide if the candidate polypeptide is predicted to bind to the peptide conjugate / MHC complex.

[0337] In some embodiments of computer-assisted methods for designing a polypeptide that can be linked to a target covalent inhibitor, interactions between atoms of the antigen-binding domain and atoms of the peptide are determined. For example, such methods can include the following steps: (a) providing a structure of a polypeptide comprising an antigen-binding domain bound to a peptide conjugate / MHC complex; (b) determining interactions between one or more atoms of the antigen-binding domain of the polypeptide and one or more atoms of the peptide conjugate / MHC complex; (c) providing a candidate polypeptide comprising an antigen-binding domain for binding to a peptide conjugate / MHC complex, wherein the antigen-binding domain of the candidate polypeptide comprises one or more amino acid substitutions relative to the polypeptide comprising the antigen-binding domain, wherein the one or more amino acid substitutions are substitutions of residues in the antigen-binding domain of the polypeptide that comprise one or more atoms that interact with or modulate the interaction of one or more atoms of the peptide conjugate / MHC complex; and d) selecting the candidate polypeptide if the candidate polypeptide binds or is predicted to bind to the peptide conjugate / MHC complex with a higher affinity (higher affinity than the affinity of the polypeptide comprising the antigen-binding domain for the peptide conjugate / MHC complex).

[0338] In some instances, computer-assisted methods for identifying or designing polypeptides capable of linking to target covalent inhibitors involve the use of a computer system, which may be, for example, a programmed computer including a processor, a data storage system, input devices, and output devices. In some embodiments, the steps of such methods include: (a) inputting data including three-dimensional coordinates of atoms from the crystal structures of Figures 57A-C, 58A-B, 59A-C, 60A-B, and 61, or a subset of atoms related thereto, into a programmed computer via the input device, thereby generating a data set; (b) using the processor to compare the data set with a computer database of structures of polypeptides comprising antigen-binding domains that bind, or are suspected to bind, or are desired to bind to peptide conjugate / MHC complexes, stored in the computer data storage system; (c) using a computer method to select from the database structure(s) structure(s) that are likely to bind to a particular peptide conjugate / MHC complex; (d) using a computer method to construct a model of the selected structure(s); and (e) outputting the selected structure(s) to the output device. Optionally, one or more of the selected structures can be synthesized and tested for binding to the peptide conjugate / MHC complex.

[0339] A computer-readable medium can be used to identify or design candidate polypeptides comprising an antigen-binding domain bound to a peptide conjugate / MHC complex. For example, in some embodiments, such a computer-readable medium can include: (a) atomic coordinate data according to the structure of any one of Figures 57A-C, 58A-B, 59A-C, 60A-B, and 61, which data can define or provide a three-dimensional structure of a polypeptide comprising an antigen-binding domain bound to a peptide conjugate / MHC complex, wherein the peptide conjugate of the peptide conjugate / MHC complex is a peptide covalently linked to a target covalent inhibitor, or a fragment thereof, or at least one subdomain thereof; or (b) structure factor data of a polypeptide comprising an antigen-binding domain bound to a peptide conjugate / MHC complex, which structure factor data can be derived from the atomic coordinate data of any one of Figures 57A-C, 58A-B, 59A-C, 60A-B, and 61. In other embodiments, the computer readable medium can include: (a) atomic coordinate data from the structure of any one of Figures 57A-C, 58A-B, 59A-C, 60A-B, and 61, wherein the data can define or provide a three-dimensional structure of a peptide conjugate / MHC complex, wherein the peptide conjugate of the peptide conjugate / MHC complex is a peptide covalently linked to a target covalent inhibitor, or a fragment thereof, or at least one subdomain thereof; (b) structure factor data of the peptide conjugate / MHC complex, wherein the structure factor data can be derived from the atomic coordinate data of the structure of any one of Figures 57A-C, 58A-B, 59A-C, 60A-B, 61.

[0340] Any of the binding partners described herein may be fully or partially humanized. Techniques for antibody humanization are known in the art and can be adapted for use in the present disclosure. In some embodiments, humanization may be performed, for example, by CDR-grafting. In some embodiments, one or more amino acids in the variable region may be altered for humanization or to improve the properties of the binding partner. In some embodiments, one or more amino acids in the framework region may be altered.

[0341] The present disclosure includes binding partners for use in diagnostic and therapeutic approaches. For therapeutic approaches, in certain embodiments, the binding partner may be delivered as an mRNA or DNA polynucleotide encoding the binding partner. Administering DNA or RNA encoding any of the binding partners described herein is also considered a method of delivering such binding partner to an individual or one or more cells. In some embodiments, the mRNA comprises synthetic mRNA containing modified nucleotides. In some embodiments, the modifications may include N1-methyl-pseudouridine (1mΨ) nucleotide substitutions and / or 5-methylcytidine (m5C) substitutions. Methods for delivering DNA and RNA encoding proteins are known in the art and can be adapted for delivering binding partners given the benefit of this disclosure. In several embodiments, one or more expression vectors are used, including viral vectors. Thus, in several embodiments, viral expression vectors are used. Viral expression vectors may be used as naked polynucleotides or may comprise any viral particle, including, but not limited to, defective interfering particles or other replication-defective viral constructs and virus-like particles. In embodiments, the expression vector comprises a modified viral polynucleotide (e.g., from an adenovirus, herpesvirus, or retrovirus). In embodiments, retroviral vectors adapted from the murine Moloney leukemia virus (MLV) or lentiviral vectors (such as lentiviral vectors adapted from human immunodeficiency virus type 1 (HIV-1)) may be used.

[0342] In one embodiment, an oncolytic viral vector is used. Oncolytic viruses (OVs), including vaccinia (OVV), mediate anticancer effects through both direct oncolysis and stimulation of the innate immune response through the production of damage-associated molecular patterns (DAMPs) and the presence of virus-derived pathogen-associated molecular patterns (PAMPs), leading to increased production of type I interferon. Furthermore, OVV-mediated oncolysis may promote direct acquisition of tumor-derived antigens by host antigen-presenting cells within the tumor microenvironment, thereby improving T cell priming and modulating the effector phase of the antitumor immune response. In another embodiment, a recombinant adeno-associated viral (AAV) vector can be used. In a specific embodiment, the expression vector is a self-complementary adeno-associated virus (scAAV).

[0343] Pharmaceutical formulations containing binding partners are included in the present disclosure and can be prepared by mixing the binding partners with one or more pharmaceutically acceptable carriers. Pharmaceutically acceptable carriers include solvents, dispersion media, isotonicity agents, and the like. The carrier can be a liquid, semi-solid (e.g., paste), or solid carrier. Examples of carriers include water, saline or other buffers (e.g., phosphate buffer, citrate buffer, etc.), oil, alcohol, proteins (e.g., serum albumin, gelatin, etc.), carbohydrates (e.g., monosaccharides, disaccharides, other carbohydrates, glucose, sucrose, trehalose, mannose, mannitol, sorbitol, dextrin), gels, lipids, liposomes, resins, porous matrices, binders, fillers, coatings, stabilizers, preservatives, liposomes, antioxidants, chelating agents such as EDTA, salt-forming counterions such as sodium; non-ionic surfactants such as TWEEN, PLURONICS, or polyethylene glycol (PEG), or combinations thereof. In several embodiments, liposomal formulations containing one or more binding partners are provided. Liposomal formulations include, but are not limited to, liposomal nanoparticles.

[0344] In some embodiments, an effective amount of one or more binding partners is administered to an individual in need thereof. In some embodiments, an effective amount is an amount that alleviates one or more signs or symptoms of a disease and / or reduces the severity of the disease. An effective amount may also inhibit or prevent the onset or recurrence of a disease. The exact dosage can be selected by an individual physician in consideration of the patient being treated. Dosage and administration can be adjusted to provide a sufficient level of binding partner to maintain the desired effect. Additional factors that may be considered include the severity and type of disease state, the patient's age, weight, and sex, the desired duration of treatment, the method of administration, the time and frequency of administration, drug combinations, reaction sensitivities, and / or tolerance / response to treatment.

[0345] The binding partner and pharmaceutical composition comprising the binding partner can be administered to an individual in need thereof by any suitable route (e.g., intravenous, intramuscular, intraperitoneal, intraspinal, subcutaneous, intraarticular, intrasynovial, oral, topical, or inhalation), depending on the specific condition being treated. The composition can be administered parenterally or enterally. The composition can be administered as a single dose or multiple doses, or continuously over a period of time. For example, administration can be a pre-specified number of times, or daily, weekly, or monthly, and can be continuous or intermittent as therapeutically indicated.

[0346] In some embodiments, the individual in need of a composition of the present disclosure has been diagnosed with or is suspected of having cancer. In some embodiments, the cancer is a solid tumor or a hematological malignancy. In some embodiments, the cancer is renal cell carcinoma, breast cancer, prostate cancer, pancreatic cancer, lung cancer (e.g., non-small cell lung cancer), liver cancer, ovarian cancer, cervical cancer, colon cancer (or colorectal cancer), esophageal cancer, glioma, glioblastoma or other brain cancer, gastric cancer, bladder cancer, testicular cancer, head and neck cancer, thyroid cancer, adrenal cancer, melanoma or other skin cancer, sarcoma including, but not limited to, fibrosarcoma, angiosarcoma, osteosarcoma, and rhabdomyosarcoma, and any type of hematological cancer, including leukemia, lymphoma, and myeloma. In some embodiments, the individual is in need of treatment for a neuroendocrine tumor. In some embodiments, the individual is in need of treatment for any pre-neoplastic disorder, including myelodysplastic syndrome or myeloproliferative neoplasm. In some embodiments, the described binding partners are used prophylactically against any of the described types of cancer. In some embodiments, the individual in need thereof has undergone a prior treatment and the individual is refractory (resistant) to the prior treatment. In some embodiments, the cancer is a relapsed or refractory cancer.

[0347] In some embodiments, one or more binding partners (which may, but need not, be in a pharmaceutical formulation) when administered to an individual in need thereof exhibit improved activity compared to a control. In some embodiments, the control comprises a different antibody, a different form of the same antibody / binding partner, or an antibody / binding partner delivered without the addition of an additional agent. In some embodiments, the binding partners described herein provide improved antibody-dependent cellular cytotoxicity (ADCC) or internalization (such as ADC) compared to the control. In some embodiments, the control protein or peptide does not comprise a covalently attached molecule. The control peptide may comprise the same sequence as the experimental peptide, or if the experimental peptide contains a mutation, the control peptide may comprise a wild-type sequence.

[0348] Compositions of the disclosure, such as pharmaceutical formulations, can include only one binding partner or more than one binding partner, and thus include combinations of different binding partners.

[0349] In one aspect, provided herein is a method of treating a disease or disorder in a subject, the method comprising administering to the subject (a) a targeted covalent inhibitor and (b) a binding partner disclosed herein. In certain embodiments, the disease or disorder is cancer, a fibrotic disease, or an autoimmune disease (e.g., rheumatoid arthritis). In certain embodiments, the targeted covalent inhibitor targets KRAS, Bruton's tyrosine kinase (BTK), EGFR (ERBB1), HER2 / NEU (ERBB2), HER3 (ERBB3), HER4 (ERBB4), fibroblast growth factor receptor (FGFR), MET, BRAF, cyclin-dependent kinase (CDK), acetylcholinesterase (ACHE), TP53, IDH1, GNAS, FBXW7, CTNNB1, DNMT3A, cathepsin, caspase, pancreatic lipase, METAP2, cancer-testis antigen, E3 ligase, DCAF, ERV, LINE, or translocation element protein.

[0350] In one aspect, provided herein is a method for killing cancer cells in a subject, the method comprising administering to the subject (a) a targeted covalent inhibitor and (b) a binding partner disclosed herein. In one embodiment, the targeted covalent inhibitor targets KRAS, Bruton's tyrosine kinase (BTK), EGFR (ERBB1), HER2 / NEU (ERBB2), HER3 (ERBB3), HER4 (ERBB4), fibroblast growth factor receptor (FGFR), MET, BRAF, cyclin-dependent kinase (CDK), acetylcholinesterase (ACHE), TP53, IDH1, GNAS, FBXW7, CTNNB1, DNMT3A, cathepsin, caspase, pancreatic lipase, METAP2, cancer-testis antigen, E3 ligase, DCAF, ERV, LINE, or translocation element protein.

[0351] In one aspect, provided herein is a method of targeting cells expressing EGFR, BTK, or RAS in a subject treated with an EGFR-, BTK-, or RAS-targeted covalent inhibitor, the method comprising administering to the subject a binding partner disclosed herein. In one embodiment, the subject has cancer.

[0352] In one aspect, provided herein is a method of treating a disease or disorder in a subject treated with a targeted covalent inhibitor, the method comprising administering to the subject a binding partner disclosed herein. In one embodiment, the disease or disorder is an autoimmune disease or a fibrotic disease.

[0353] In one aspect, provided herein is a method of treating cancer in a subject treated with a targeted covalent inhibitor, the method comprising administering to the subject a binding partner disclosed herein. In one embodiment, the targeted covalent inhibitor targets RAS, Bruton's tyrosine kinase (BTK), EGFR (ERBB1), HER2 / NEU (ERBB2), HER3 (ERBB3), HER4 (ERBB4), fibroblast growth factor receptor (FGFR), MET, BRAF, cyclin-dependent kinase (CDK), acetylcholinesterase (ACHE), TP53, IDH1, GNAS, FBXW7, CTNNB1, DNMT3A, cathepsin, caspase, pancreatic lipase, METAP2, cancer-testis antigen, E3 ligase, or DCAF.

[0354] The targeted covalent inhibitor can be a non-peptide molecule described herein. The non-peptide molecule can be administered to a subject in need thereof before administering a binding partner described herein. The subject described herein can be a cancer patient. In some cases, the binding partner can be administered to a subject who has already been previously treated with a non-peptide molecule. In some cases, the binding partner can be administered to a subject simultaneously with or after administration of a non-peptide molecule (e.g., a targeted covalent inhibitor or a drug described herein). In some cases, the binding partner can be administered to a subject before administering a non-peptide molecule.

[0355] In some cases, the subjects described herein are refractory to the first-line treatment. For example, the subjects described herein may be refractory to the first-line treatment, which is chemotherapy. For example, the subjects described herein may be refractory to a non-peptide molecule such as AMG-510, osimertinib, ARS-1620, MRTX849, JNJ74699157, LY3499446, MRTX-1257, JDQ443, MRTX-1133, RMC-6291, or RMC-9805. In some cases, the subject may be further treated or administered with a binding partner (e.g., an antibody or fragment thereof) described herein after relapse from the previous treatment. In some cases, the binding partners described herein can be used to treat relapsed or refractory cancer.

[0356] In some cases, the peptide conjugate described herein is administered prophylactically before the subject develops cancer, or before the subject is administered a drug, or both.Cancer can be caused by RAS mutation...

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