Anti-IL2RA antibodies and uses thereof

Abstract

Anti-IL2RA (Interleukin-2 Receptor Alpha Chain) antibodies, antigen binding fragments, and uses thereof are provided.

Description

[0001] Priority Declaration

[0002] This application claims priority to PCT / CN2023 / 077623, filed on February 22, 2023. The entire contents of the aforementioned application are incorporated herein by reference. Technical Field

[0003] The present disclosure relates to anti-IL2RA (interleukin-2 receptor alpha chain) antibodies and uses thereof. Background Art

[0004] Cancer is one of the diseases with the highest mortality rates in the world. According to the World Health Organization, in 2012, there were 14 million cancer cases and 8.2 million cancer deaths worldwide. In China, there were 3.07 million new cancer cases and 2.2 million cancer deaths.

[0005] The recent clinical and commercial success of anti-cancer antibodies has sparked significant interest in antibody-based therapeutics. There is a growing need to develop antibodies for various antibody-based therapies to treat cancer or autoimmune diseases. Summary of the Invention

[0006] The present disclosure relates to anti-IL2RA antibodies, antigen-binding fragments thereof, and uses thereof.

[0007] In one aspect, the present disclosure relates to an antibody or antigen-binding fragment thereof that binds to interleukin-2 receptor alpha chain (IL2RA), comprising: a heavy chain variable region (VH) comprising complementarity determining regions (CDRs) 1, 2, and 3, in some embodiments, the VH CDR1 region comprises an amino acid sequence at least 80% identical to a selected VH CDR1 amino acid sequence, the VH CDR2 region comprises an amino acid sequence at least 80% identical to a selected VH CDR2 amino acid sequence, and the VH CDR3 region comprises an amino acid sequence at least 80% identical to a selected VH CDR3 amino acid sequence; and a light chain variable region (VL) comprising CDRs 1, 2, and 3, in some embodiments, the VL CDR1 region comprises an amino acid sequence at least 80% identical to a selected VL CDR1 amino acid sequence, the VL CDR2 region comprises an amino acid sequence at least 80% identical to a selected VL CDR2 amino acid sequence, and the VL CDR3 region comprises an amino acid sequence at least 80% identical to a selected VL CDR3 amino acid sequence;

[0008] In some embodiments, the selected VH CDR 1, 2, and 3 amino acid sequence and the selected VL CDR 1, 2, and 3 amino acid sequence are one of the following:

[0009] (1) the selected VH CDR 1, 2, and 3 amino acid sequences are as shown in SEQ ID NOs: 1, 2, and 3, respectively, and the selected VLCDR 1, 2, and 3 amino acid sequences are as shown in SEQ ID NOs: 43, 44, and 45, respectively;

[0010] (2) the selected VH CDR 1, 2, and 3 amino acid sequences are shown in SEQ ID NOs: 4, 5, and 6, respectively, and the selected VLCDR 1, 2, and 3 amino acid sequences are shown in SEQ ID NOs: 46, 47, and 48, respectively;

[0011] (3) the selected VH CDR 1, 2, and 3 amino acid sequences are shown in SEQ ID NOs: 7, 8, and 9, respectively, and the selected VLCDR 1, 2, and 3 amino acid sequences are shown in SEQ ID NOs: 49, 50, and 51, respectively;

[0012] (4) the selected VH CDR 1, 2, and 3 amino acid sequences are shown in SEQ ID NOs: 10, 11, and 12, respectively, and the selected VL CDR 1, 2, and 3 amino acid sequences are shown in SEQ ID NOs: 49, 50, and 51, respectively;

[0013] (5) the selected VH CDR 1, 2, and 3 amino acid sequences are shown in SEQ ID NOs: 13, 14, and 15, respectively, and the selected VL CDR 1, 2, and 3 amino acid sequences are shown in SEQ ID NOs: 49, 50, and 51, respectively;

[0014] (6) the selected VH CDR 1, 2, and 3 amino acid sequences are shown in SEQ ID NOs: 16, 17, and 18, respectively, and the selected VL CDR 1, 2, and 3 amino acid sequences are shown in SEQ ID NOs: 49, 50, and 51, respectively;

[0015] (7) the selected VH CDR 1, 2, and 3 amino acid sequences are shown in SEQ ID NOs: 19, 20, and 21, respectively, and the selected VL CDR 1, 2, and 3 amino acid sequences are shown in SEQ ID NOs: 49, 50, and 51, respectively;

[0016] (8) the selected VH CDR 1, 2, and 3 amino acid sequences are shown in SEQ ID NOs: 22, 23, and 24, respectively, and the selected VL CDR 1, 2, and 3 amino acid sequences are shown in SEQ ID NOs: 43, 44, and 45, respectively;

[0017] (9) the selected VH CDR 1, 2, and 3 amino acid sequences are shown in SEQ ID NOs: 25, 26, and 27, respectively, and the selected VL CDR 1, 2, and 3 amino acid sequences are shown in SEQ ID NOs: 46, 47, and 48, respectively;

[0018] (10) the selected VH CDR 1, 2, and 3 amino acid sequences are shown in SEQ ID NOs: 28, 29, and 30, respectively, and the selected VL CDR 1, 2, and 3 amino acid sequences are shown in SEQ ID NOs: 49, 50, and 51, respectively;

[0019] (11) the selected VH CDR 1, 2, and 3 amino acid sequences are shown in SEQ ID NOs: 31, 32, and 33, respectively, and the selected VL CDR 1, 2, and 3 amino acid sequences are shown in SEQ ID NOs: 49, 50, and 51, respectively;

[0020] (12) the selected VH CDR 1, 2, and 3 amino acid sequences are shown in SEQ ID NOs: 34, 35, and 36, respectively, and the selected VL CDR 1, 2, and 3 amino acid sequences are shown in SEQ ID NOs: 49, 50, and 51, respectively;

[0021] (13) the selected VH CDR 1, 2, and 3 amino acid sequences are shown in SEQ ID NOs: 37, 38, and 39, respectively, and the selected VL CDR 1, 2, and 3 amino acid sequences are shown in SEQ ID NOs: 49, 50, and 51, respectively; and

[0022] (14) The selected VH CDR 1, 2, and 3 amino acid sequences are shown in SEQ ID NOs: 40, 41, and 42, respectively, and the selected VL CDR 1, 2, and 3 amino acid sequences are shown in SEQ ID NOs: 49, 50, and 51, respectively.

[0023] In some embodiments, according to the Kabat definition, the VH comprises CDRs 1, 2, and 3 of the amino acid sequences set forth in SEQ ID NOs: 1, 2, and 3, respectively, and the VL comprises CDRs 1, 2, and 3 of the amino acid sequences set forth in SEQ ID NOs: 43, 44, and 45, respectively. In some embodiments, according to the Kabat definition, the VH comprises CDRs 1, 2, and 3 of the amino acid sequences set forth in SEQ ID NOs: 4, 5, and 6, respectively, and the VL comprises CDRs 1, 2, and 3 of the amino acid sequences set forth in SEQ ID NOs: 46, 47, and 48, respectively. In some embodiments, according to the Kabat definition, the VH comprises CDRs 1, 2, and 3 of the amino acid sequences set forth in SEQ ID NOs: 7, 8, and 9, respectively, and the VL comprises CDRs 1, 2, and 3 of the amino acid sequences set forth in SEQ ID NOs: 49, 50, and 51, respectively. In some embodiments, according to the Kabat definition, the VH comprises CDRs 1, 2, and 3 of the amino acid sequences set forth in SEQ ID NOs: 10, 11, and 12, respectively, and the VL comprises CDRs 1, 2, and 3 of the amino acid sequences set forth in SEQ ID NOs: 49, 50, and 51, respectively. In some embodiments, according to the Kabat definition, the VH comprises CDRs 1, 2, and 3 of the amino acid sequences set forth in SEQ ID NOs: 13, 14, and 15, respectively, and the VL comprises CDRs 1, 2, and 3 of the amino acid sequences set forth in SEQ ID NOs: 49, 50, and 51, respectively. In some embodiments, according to the Kabat definition, the VH comprises CDRs 1, 2, and 3 of the amino acid sequences set forth in SEQ ID NOs: 16, 17, and 18, respectively, and the VL comprises CDRs 1, 2, and 3 of the amino acid sequences set forth in SEQ ID NOs: 49, 50, and 51, respectively. In some embodiments, according to the Kabat definition, the VH comprises CDRs 1, 2, and 3 of the amino acid sequences shown in SEQ ID NOs: 19, 20, and 21, respectively, and the VL comprises CDRs 1, 2, and 3 of the amino acid sequences shown in SEQ ID NOs: 49, 50, and 51, respectively.

[0024] In some embodiments, the antibody or antigen-binding fragment thereof specifically binds to human IL2RA or monkey IL2RA. In some embodiments, the antibody or antigen-binding fragment thereof is a humanized or human antibody or antigen-binding fragment thereof (e.g., a human IgG1 antibody). In some embodiments, the antibody or antigen-binding fragment thereof is a single-chain variable region fragment (scFv) or a multispecific antibody (e.g., a bispecific antibody).

[0025] In one aspect, the disclosure relates to a nucleic acid comprising a polynucleotide encoding a polypeptide comprising:

[0026] (1) an immunoglobulin heavy chain or fragment thereof comprising a heavy chain variable region (VH), wherein the VH comprises complementarity determining regions (CDRs) 1, 2, and 3, wherein the CDRs comprise the amino acid sequences set forth in SEQ ID NOs: 1, 2, and 3, respectively; and in some embodiments, when the VH is paired with a light chain variable region (VL) comprising the amino acid sequence set forth in SEQ ID NO: 53, it binds to IL2RA;

[0027] (2) an immunoglobulin light chain or fragment thereof comprising a VL comprising CDRs 1, 2, and 3, wherein the CDRs comprise the amino acid sequences set forth in SEQ ID NOs: 43, 44, and 45, respectively; in some embodiments, when the VL is paired with a VH comprising the amino acid sequence set forth in SEQ ID NO: 52, it binds to IL2RA;

[0028] (3) an immunoglobulin heavy chain or fragment thereof comprising a VH comprising CDRs 1, 2, and 3, wherein the CDRs comprise the amino acid sequences set forth in SEQ ID NOs: 4, 5, and 6, respectively; in some embodiments, when the VH is paired with a VL comprising the amino acid sequence set forth in SEQ ID NO: 55, it binds to IL2RA;

[0029] (4) an immunoglobulin light chain or fragment thereof comprising a VL comprising CDRs 1, 2, and 3, wherein the CDRs comprise the amino acid sequences set forth in SEQ ID NOs: 46, 47, and 48, respectively; in some embodiments, when the VL is paired with a VH comprising the amino acid sequence set forth in SEQ ID NO: 54, it binds to IL2RA;

[0030] (5) an immunoglobulin heavy chain or fragment thereof comprising a VH comprising CDRs 1, 2, and 3, wherein the CDRs comprise the amino acid sequences set forth in SEQ ID NOs: 7, 8, and 9, respectively; in some embodiments, when the VH is paired with a VL comprising the amino acid sequence set forth in SEQ ID NO: 61, it binds to IL2RA;

[0031] (6) an immunoglobulin light chain or fragment thereof comprising a VL, wherein the VL comprises CDRs 1, 2, and 3, wherein the CDRs comprise the amino acid sequences set forth in SEQ ID NOs: 49, 50, and 51, respectively; in some embodiments, when the VL is paired with a VH comprising the amino acid sequence set forth in SEQ ID NO: 56, it binds to IL2RA;

[0032] (7) an immunoglobulin heavy chain or fragment thereof comprising a VH comprising CDRs 1, 2, and 3, wherein the CDRs comprise the amino acid sequences shown in SEQ ID NOs: 10, 11, and 12, respectively; in some embodiments, when the VH is paired with a VL comprising the amino acid sequence shown in SEQ ID NO: 61, it binds to IL2RA;

[0033] (8) an immunoglobulin light chain or fragment thereof comprising a VL, wherein the VL comprises CDRs 1, 2, and 3, wherein the CDRs comprise the amino acid sequences shown in SEQ ID NOs: 49, 50, and 51, respectively; in some embodiments, when the VL is paired with a VH comprising the amino acid sequence shown in SEQ ID NO: 57, it binds to IL2RA;

[0034] (9) an immunoglobulin heavy chain or fragment thereof comprising a VH comprising CDRs 1, 2, and 3, wherein the CDRs comprise the amino acid sequences shown in SEQ ID NOs: 13, 14, and 15, respectively; in some embodiments, when the VH is paired with a VL comprising the amino acid sequence shown in SEQ ID NO: 61, it binds to IL2RA;

[0035] (10) an immunoglobulin light chain or fragment thereof comprising a VL, wherein the VL comprises CDRs 1, 2, and 3, wherein the CDRs comprise the amino acid sequences shown in SEQ ID NOs: 49, 50, and 51, respectively; in some embodiments, when the VL is paired with a VH comprising the amino acid sequence shown in SEQ ID NO: 58, it binds to IL2RA;

[0036] (11) an immunoglobulin heavy chain or fragment thereof comprising a VH, wherein the VH comprises CDRs 1, 2, and 3, wherein the CDRs comprise the amino acid sequences shown in SEQ ID NOs: 16, 17, and 18, respectively; in some embodiments, when the VH is paired with a VL comprising the amino acid sequence shown in SEQ ID NO: 61, it binds to IL2RA;

[0037] (12) an immunoglobulin light chain or fragment thereof comprising a VL, wherein the VL comprises CDRs 1, 2, and 3, wherein the CDRs comprise the amino acid sequences shown in SEQ ID NOs: 49, 50, and 51, respectively; in some embodiments, when the VL is paired with a VH comprising the amino acid sequence shown in SEQ ID NO: 59, it binds to IL2RA;

[0038] (13) an immunoglobulin heavy chain or fragment thereof comprising a VH comprising CDRs 1, 2, and 3, wherein the CDRs comprise the amino acid sequences shown in SEQ ID NOs: 19, 20, and 21, respectively; in some embodiments, when the VH is paired with a VL comprising the amino acid sequence shown in SEQ ID NO: 61, it binds to IL2RA;

[0039] (14) an immunoglobulin light chain or fragment thereof comprising a VL, wherein the VL comprises CDRs 1, 2, and 3, wherein the CDRs comprise the amino acid sequences shown in SEQ ID NOs: 49, 50, and 51, respectively; in some embodiments, when the VL is paired with a VH comprising the amino acid sequence shown in SEQ ID NO: 60, it binds to IL2RA;

[0040] (15) an immunoglobulin heavy chain or fragment thereof comprising a VH comprising CDRs 1, 2, and 3, wherein the CDRs comprise the amino acid sequences shown in SEQ ID NOs: 22, 23, and 24, respectively; in some embodiments, when the VH is paired with a VL comprising the amino acid sequence shown in SEQ ID NO: 53, it binds to IL2RA;

[0041] (16) an immunoglobulin heavy chain or fragment thereof comprising a VH comprising CDRs 1, 2, and 3, wherein the CDRs comprise the amino acid sequences shown in SEQ ID NOs: 25, 26, and 27, respectively; in some embodiments, when the VH is paired with a VL comprising the amino acid sequence shown in SEQ ID NO: 55, it binds to IL2RA;

[0042] (17) an immunoglobulin heavy chain or fragment thereof comprising a VH, wherein the VH comprises CDRs 1, 2, and 3, wherein the CDRs comprise the amino acid sequences shown in SEQ ID NOs: 28, 29, and 30, respectively; in some embodiments, when the VH is paired with a VL comprising the amino acid sequence shown in SEQ ID NO: 61, it binds to IL2RA;

[0043] (18) An immunoglobulin heavy chain or fragment thereof comprising a VH comprising CDRs 1, 2, and 3, wherein the CDRs comprise the amino acid sequences shown in SEQ ID NOs: 31, 32, and 33, respectively; in some embodiments, when the VH is paired with a VL comprising the amino acid sequence shown in SEQ ID NO: 61, it binds to IL2RA;

[0044] (19) An immunoglobulin heavy chain or fragment thereof comprising a VH, wherein the VH comprises CDRs 1, 2, and 3, wherein the CDRs comprise the amino acid sequences shown in SEQ ID NOs: 34, 35, and 36, respectively; in some embodiments, when the VH is paired with a VL comprising the amino acid sequence shown in SEQ ID NO: 61, it binds to IL2RA;

[0045] (20) an immunoglobulin heavy chain or fragment thereof comprising a VH comprising CDRs 1, 2, and 3, wherein the CDRs comprise the amino acid sequences shown in SEQ ID NOs: 37, 38, and 39, respectively; in some embodiments, when the VH is paired with a VL comprising the amino acid sequence shown in SEQ ID NO: 61, it binds to IL2RA; or

[0046] (21) An immunoglobulin heavy chain or fragment thereof, comprising a VH, wherein the VH comprises CDRs 1, 2, and 3, wherein the CDRs comprise the amino acid sequences shown in SEQ ID NOs: 40, 41, and 42, respectively; in some embodiments, when the VH is paired with a VL comprising the amino acid sequence shown in SEQ ID NO: 61, it binds to IL2RA.

[0047] In some embodiments, the nucleic acid comprises a polynucleotide encoding a polypeptide comprising an immunoglobulin heavy chain or a fragment thereof, the fragment comprising a heavy chain variable region (VH), the heavy chain variable region comprising complementarity determining regions 1, 2, and 3 (CDR1, CDR2, CDR3), the CDR1, CDR2, and CDR3 comprising the amino acid sequences set forth in SEQ ID NOs: 1, 2, and 3, respectively, or the amino acid sequences set forth in SEQ ID NOs: 22, 23, and 24, respectively. In some embodiments, the nucleic acid comprises a polynucleotide encoding a polypeptide comprising an immunoglobulin light chain or a fragment thereof, the fragment comprising a light chain variable region (VL), the light chain variable region comprising CDR1, CDR2, and CDR3, the CDR1, CDR2, and CDR3 comprising the amino acid sequences set forth in SEQ ID NOs: 43, 44, and 45, respectively.

[0048] In some embodiments, the nucleic acid comprises a polynucleotide encoding a polypeptide comprising an immunoglobulin heavy chain or a fragment thereof, wherein the fragment comprises a VH, wherein the VH comprises CDR1, CDR2, and CDR3, wherein the CDR1, CDR2, and CDR3 comprise the amino acid sequences set forth in SEQ ID NOs: 4, 5, and 6, respectively, or comprise the amino acid sequences set forth in SEQ ID NOs: 25, 26, and 27, respectively. In some embodiments, the nucleic acid comprises a polynucleotide encoding a polypeptide comprising an immunoglobulin light chain or a fragment thereof, wherein the fragment comprises a VL, wherein the VL comprises CDR1, CDR2, and CDR3, wherein the CDR1, CDR2, and CDR3 comprise the amino acid sequences set forth in SEQ ID NOs: 46, 47, and 48, respectively.

[0049] In some embodiments, the nucleic acid comprises a polynucleotide encoding a polypeptide comprising an immunoglobulin heavy chain or a fragment thereof, the fragment comprising a VH, the VH comprising CDR1, CDR2, and CDR3, wherein the CDR1, CDR2, and CDR3 comprise the amino acid sequences of SEQ ID NOs: 7, 8, and 9, respectively, or comprise the amino acid sequences of SEQ ID NOs: 28, 29, and 30, respectively. In some embodiments, the nucleic acid comprises a polynucleotide encoding a polypeptide comprising an immunoglobulin heavy chain or a fragment thereof, the fragment comprising a VH, the VH comprising CDR1, CDR2, and CDR3, wherein the CDR1, CDR2, and CDR3 comprise the amino acid sequences of SEQ ID NOs: 10, 11, and 12, respectively, or comprise the amino acid sequences of SEQ ID NOs: 31, 32, and 33, respectively. In some embodiments, the nucleic acid comprises a polynucleotide encoding a polypeptide comprising an immunoglobulin heavy chain or a fragment thereof, the fragment comprising a VH, the VH comprising CDR1, CDR2, and CDR3, wherein the CDR1, CDR2, and CDR3 comprise the amino acid sequences of SEQ ID NOs: 13, 14, and 15, respectively, or comprise the amino acid sequences of SEQ ID NOs: 34, 35, and 36, respectively. In some embodiments, the nucleic acid comprises a polynucleotide encoding a polypeptide comprising an immunoglobulin heavy chain or a fragment thereof, the fragment comprising a VH, the VH comprising CDR1, CDR2, and CDR3, wherein the CDR1, CDR2, and CDR3 comprise the amino acid sequences of SEQ ID NOs: 16, 17, and 18, respectively, or comprise the amino acid sequences of SEQ ID NOs: 37, 38, and 39, respectively. In some embodiments, the nucleic acid comprises a polynucleotide encoding a polypeptide comprising an immunoglobulin heavy chain or a fragment thereof, wherein the fragment comprises a VH, wherein the VH comprises CDR1, CDR2, and CDR3, wherein the CDR1, CDR2, and CDR3 comprise the amino acid sequences set forth in SEQ ID NOs: 19, 20, and 21, respectively, or comprise the amino acid sequences set forth in SEQ ID NOs: 40, 41, and 42, respectively. In some embodiments, the nucleic acid comprises a polynucleotide encoding a polypeptide comprising an immunoglobulin light chain or a fragment thereof, wherein the fragment comprises a VL, wherein the VL comprises CDR1, CDR2, and CDR3, wherein the CDR1, CDR2, and CDR3 comprise the amino acid sequences set forth in SEQ ID NOs: 49, 50, and 51, respectively.

[0050] In some embodiments, when the VH is paired with the VL, it specifically binds to human IL2RA; or when the VL is paired with the VH, it specifically binds to human IL2RA or monkey IL2RA. In some embodiments, the immunoglobulin heavy chain or fragment thereof is a humanized or human immunoglobulin heavy chain or fragment thereof (e.g., a human IgG1 heavy chain or fragment thereof), and the immunoglobulin light chain or fragment thereof is a humanized or human immunoglobulin light chain or fragment thereof. In some embodiments, the nucleic acid encodes a single-chain variable fragment (scFv), a multispecific antibody (e.g., a bispecific antibody), or a chimeric antigen receptor (CAR). In some embodiments, the nucleic acid is a complementary deoxyribonucleic acid (cDNA).

[0051] In one aspect, the present disclosure relates to vectors comprising one or more nucleic acids described herein. In one aspect, the present disclosure relates to vectors comprising two nucleic acids described herein; in some embodiments, the vectors encode a heavy chain variable region (VH) and a light chain variable region (VL) that jointly bind to IL2RA. In one aspect, the present disclosure relates to a pair of vectors; in some embodiments, each vector comprises a nucleic acid described herein; in some embodiments, the pair of vectors jointly encode a VH region and a VL region that can jointly bind to IL2RA.

[0052] In one aspect, the present disclosure relates to a cell comprising a vector or pair of vectors described herein. In some embodiments, the cell is a CHO cell. In one aspect, the present disclosure relates to a cell comprising one or more nucleic acids described herein. In one aspect, the present disclosure relates to a cell comprising two nucleic acids described herein. In some embodiments, the two nucleic acids together encode a VH region and a VL region that can jointly bind to IL2RA.

[0053] In one aspect, the present disclosure relates to a method for producing an antibody or an antigen-binding fragment thereof, the method comprising: (a) culturing a cell as described herein under conditions sufficient for the cell to produce the antibody or antigen-binding fragment thereof; and (b) collecting the antibody or antigen-binding fragment thereof produced by the cell.

[0054] In one aspect, the present disclosure relates to an antibody or antigen-binding fragment thereof that binds to IL2RA, comprising: a heavy chain variable region (VH) comprising an amino acid sequence that is at least 80% identical to a selected VH sequence; and a light chain variable region (VL) comprising an amino acid sequence that is at least 80% identical to a selected VL sequence; in some embodiments, the selected VH sequence and the selected VL sequence are one of the following: (1) the selected VH sequence is SEQ ID NO: 52 and the selected VL sequence is SEQ ID NO: 53; (2) the selected VH sequence is SEQ ID NO: 54 and the selected VL sequence is SEQ ID NO: 55; (3) the selected VH sequence is SEQ ID NO: 56 and the selected VL sequence is SEQ ID NO: 61; (4) the selected VH sequence is SEQ ID NO: 57 and the selected VL sequence is SEQ ID NO: 61; (5) the selected VH sequence is SEQ ID NO: 58 and the selected VL sequence is SEQ ID NO: 61; (6) the selected VH sequence is SEQ ID NO: ID NO:59, the selected VL sequence is SEQ ID NO:61; (7) the selected VH sequence is SEQ ID NO:60, and the selected VL sequence is SEQ ID NO:61.

[0055] In some embodiments, the VH comprises the sequence of SEQ ID NO: 52, and the VL comprises the sequence of SEQ ID NO: 53. In some embodiments, the VH comprises the sequence of SEQ ID NO: 54, and the VL comprises the sequence of SEQ ID NO: 55. In some embodiments, the VH comprises the sequence of SEQ ID NO: 56, and the VL comprises the sequence of SEQ ID NO: 61. In some embodiments, the VH comprises the sequence of SEQ ID NO: 57, and the VL comprises the sequence of SEQ ID NO: 61. In some embodiments, the VH comprises the sequence of SEQ ID NO: 58, and the VL comprises the sequence of SEQ ID NO: 61. In some embodiments, the VH comprises the sequence of SEQ ID NO: 59, and the VL comprises the sequence of SEQ ID NO: 61. In some embodiments, the VH comprises the sequence of SEQ ID NO: 60, and the VL comprises the sequence of SEQ ID NO: 61. In some embodiments, the antibody or antigen-binding fragment thereof specifically binds to human IL2RA or monkey IL2RA. In some embodiments, the antibody or its antigen-binding fragment is a humanized or human antibody or its antigen-binding fragment. In some embodiments, the antibody or its antigen-binding fragment is a single-chain variable fragment (scFv) or a multispecific antibody (e.g., a bispecific antibody).

[0056] In one aspect, the disclosure relates to antibodies or antigen-binding fragments thereof that cross-compete with the antibodies or antigen-binding fragments thereof described herein.

[0057] In one aspect, the present disclosure relates to an antibody or antigen-binding fragment thereof that binds to IL2RA, comprising: a heavy chain variable region (VH) comprising a VH CDR1, VH CDR2, and VH CDR3 identical to the VH CDR1, VH CDR2, and VH CDR3 of a selected VH sequence; and a light chain variable region (VL) comprising a VL CDR1, VL CDR2, and VL CDR3 identical to the VL CDR1, VL CDR2, and VL CDR3 of a selected VL sequence; in some embodiments, the selected VH sequence and the selected VL sequence are one of the following: (1) the selected VH sequence is SEQ ID NO: 52 and the selected VL sequence is SEQ ID NO: 53; (2) the selected VH sequence is SEQ ID NO: 54 and the selected VL sequence is SEQ ID NO: 55; (3) the selected VH sequence is SEQ ID NO: 56 and the selected VL sequence is SEQ ID NO: 61; (4) the selected VH sequence is SEQ ID NO: NO:57, the selected VL sequence is SEQ ID NO:61; (5) the selected VH sequence is SEQ ID NO:58, the selected VL sequence is SEQ ID NO:61; (6) the selected VH sequence is SEQ ID NO:59, the selected VL sequence is SEQ ID NO:61; (7) the selected VH sequence is SEQ ID NO:60, the selected VL sequence is SEQ ID NO:61.

[0058] In one aspect, the present disclosure relates to an antibody-drug conjugate comprising a covalent conjugate of an antibody or antigen-binding fragment thereof as described herein and a therapeutic agent. In some embodiments, the therapeutic agent is a cytotoxic agent or a cytostatic agent.

[0059] In one aspect, the present disclosure relates to a method for treating a subject with cancer, the method comprising administering to the subject a therapeutically effective amount of a composition comprising an antibody or its antigen-binding fragment as described herein or an antibody-drug conjugate. In some embodiments, the subject suffers from solid tumors, brain cancer, lung cancer, melanoma, prostate cancer, esophageal squamous cell carcinoma, leukemia, lymphoma, ovarian cancer, colorectal cancer, head and neck cancer, bladder cancer, kidney cancer and breast cancer. In some embodiments, the method described herein further comprises administering to the subject a therapeutically effective amount of an anti-OX40 antibody, an anti-PD1 antibody, an anti-PDL1 antibody, an anti-PDL2 antibody, an anti-LAG-3 antibody, an anti-TIGIT antibody, an anti-CTLA-4 antibody, an anti-GITR antibody, an anti-TIM-3 antibody, an anti-4-1BB antibody and / or an anti-CD40 antibody.

[0060] In one aspect, the disclosure relates to a method of reducing the growth rate of a tumor, comprising contacting a tumor cell with an effective amount of a composition comprising an antibody or antigen-binding fragment thereof, or an antibody-drug conjugate described herein.

[0061] In one aspect, the present disclosure relates to a method of killing tumor cells, comprising contacting the tumor cells with an effective amount of a composition comprising an antibody or antigen-binding fragment thereof, or an antibody-drug conjugate described herein.

[0062] In one aspect, the present disclosure relates to a pharmaceutical composition comprising an antibody or antigen-binding fragment thereof as described herein and a pharmaceutically acceptable carrier. In one aspect, the present disclosure relates to a pharmaceutical composition comprising an antibody-drug conjugate as described herein and a pharmaceutically acceptable carrier.

[0063] In one aspect, the present disclosure relates to a chimeric antigen receptor (CAR) comprising an antibody or antigen-binding fragment thereof described herein.

[0064] As used herein, the term "cancer" refers to cells with autonomous growth capacity. Examples of such cells include cells that present an abnormal state or condition, characterized by rapid cell proliferation and growth. The term encompasses cancerous growths (e.g., tumors), carcinogenic processes, metastatic tissues, and malignantly transformed cells, tissues, or organs, regardless of their histopathological type or invasion stage. Also included are malignant tumors of various organ systems, such as those of the head and neck, respiratory system, cardiovascular system, kidney, reproductive system, blood system, nervous system, liver, gastrointestinal tract, and endocrine system; and adenocarcinomas, including malignant tumors such as most colon cancers, renal cell carcinomas, prostate cancer, and / or testicular tumors, non-small cell lung cancers, gliomas, and small intestine cancer. "Naturally occurring" cancers include any cancer that is not experimentally induced by implanting cancer cells into a subject, including, for example, spontaneously occurring cancers, cancers caused by exposure of the patient to carcinogens, cancers caused by inserting transgenic oncogenes or knocking out tumor suppressor genes, and cancers caused by infection (e.g., viral infection). The term "carcinoma" is art-recognized and refers to a malignant tumor of epithelial or endocrine tissue. The term also includes carcinosarcoma, which is a malignant tumor composed of both carcinomatous and sarcomatous tissue.

[0065] Adenocarcinoma refers to a cancer that originates from glandular tissue or tumor cells that form a recognizable glandular structure. The term "sarcoma"

[0066] Sarcoma is a well-known term in the art and refers to malignant tumors of mesenchymal origin.

[0067] Hematopoietic neoplastic disorders include diseases involving proliferative / neoplastic cells of hematopoietic origin. Hematopoietic neoplastic diseases can originate from myeloid, lymphoid, erythroid, or their precursor cells. Hematologic cancer is a cancer that begins in hematopoietic tissue (such as the bone marrow) or cells of the immune system. Examples of hematologic cancers include, for example, leukemias, lymphomas, and multiple myeloma.

[0068] As used herein, the term "antibody" refers to any antigen binding molecule that comprises at least one (e.g., one, two, three, four, five, or six) complementary determining regions (CDRs) (e.g., any three CDRs of an immunoglobulin light chain or any three CDRs of an immunoglobulin heavy chain) and is capable of specifically binding to an epitope. Non-limiting examples of antibodies include monoclonal antibodies, polyclonal antibodies, multispecific antibodies (e.g., bispecific antibodies), single-chain antibodies, chimeric antibodies, human antibodies, and humanized antibodies. In some embodiments, an antibody may comprise the Fc region of a human antibody. The term also includes derivatives such as bispecific antibodies, single-chain antibodies, diabodies, linear antibodies, and multispecific antibodies formed from antibody fragments.

[0069] As used herein, the term "antigen-binding fragment" refers to a portion of a full-length antibody that is capable of specifically binding to an antigen. In some embodiments, the antigen-binding fragment comprises at least one variable domain (e.g., a heavy chain variable domain or a light chain variable domain). Non-limiting examples of antibody fragments include, for example, Fab, Fab', F(ab')2, and Fv fragments.

[0070] As used herein, the term "human antibody" refers to an antibody encoded by endogenous nucleic acids derived from humans (e.g., rearranged human immunoglobulin heavy or light chain loci). In some embodiments, human antibodies are collected from the human body or produced in human cell cultures (e.g., human hybridoma cells). In some embodiments, human antibodies are produced in non-human cells (e.g., mouse or hamster cell lines). In some embodiments, human antibodies are produced in bacteria or yeast cells. In some embodiments, human antibodies are produced in transgenic non-human animals (e.g., cattle) comprising unrearranged or rearranged human immunoglobulin loci (e.g., heavy or light chain human immunoglobulin loci).

[0071] As used herein, the term "chimeric antibody" refers to an antibody that comprises sequences present in at least two different species (e.g., two different mammalian species such as human and mouse). A non-limiting example of a chimeric antibody is an antibody that comprises a variable domain sequence (e.g., all or part of a light chain and / or heavy chain variable domain sequence) of a non-human (e.g., mouse) antibody and a human antibody constant domain. Other examples of chimeric antibodies are described herein and are known in the art.

[0072] As used herein, the term "humanized antibody" refers to a non-human antibody that contains minimal sequences derived from a non-human (e.g., mouse) immunoglobulin and contains sequences derived from a human immunoglobulin. In a non-limiting example, a humanized antibody is a human antibody (recipient antibody) in which the hypervariable region (e.g., CDR) residues of the recipient antibody are replaced with hypervariable region (e.g., CDR) residues of a non-human antibody (e.g., a donor antibody, such as a mouse, rat, or rabbit antibody) having the desired specificity, affinity, and capacity. In some embodiments, Fv framework region residues of a human immunoglobulin are replaced with corresponding non-human (e.g., mouse) immunoglobulin residues. In some embodiments, a humanized antibody may contain residues that are not present in the recipient antibody or the donor antibody. These modifications may be made to further optimize antibody performance. In some embodiments, the humanized antibody comprises at least one (usually two) substantially complete variable domains, in which all or substantially all hypervariable loops (CDRs) correspond to the CDRs of a non-human (e.g., mouse) immunoglobulin, and all or substantially all framework regions are those of a human immunoglobulin. The humanized antibody may also comprise at least a portion of an immunoglobulin constant region (Fc), typically the Fc region of a human immunoglobulin. Humanized antibodies can be produced using molecular biology methods known in the art. Non-limiting examples of methods for producing humanized antibodies are described herein.

[0073] As used herein, the term "single-chain antibody" refers to a single peptide chain comprising at least two immunoglobulin variable domains (e.g., variable domains of mammalian immunoglobulin heavy or light chains) and capable of specific binding to an antigen. Non-limiting examples of single-chain antibodies are described herein.

[0074] As used herein, the terms "subject" and "patient" are used interchangeably throughout the specification to refer to animals (human or non-human) that are treated by the methods of the present invention. The present disclosure encompasses veterinary and non-veterinary applications. Human patients may be adults or minors (e.g., humans under the age of 18). In addition to humans, patients include, but are not limited to, mice, rats, hamsters, guinea pigs, rabbits, ferrets, cats, dogs, and primates. For example, non-human primates (e.g., monkeys, chimpanzees, gorillas, etc.), rodents (e.g., rats, mice, gerbils, hamsters, ferrets, rabbits), lagomorphs, suids (e.g., pigs, mini-pigs), equines, canines, felines, bovines, and other domestic, farm, and zoo animals are included.

[0075] As used herein, the phrases "specifically bind" and "specifically binds" when referring to an antibody mean that the antibody interacts with its target molecule (e.g., IL2RA) preferentially over other molecules because the interaction is dependent on the presence of a specific structure (i.e., antigenic determinant or epitope) on the target molecule; in other words, the agent recognizes and binds to molecules containing a specific structure, rather than binding universally to all molecules. An antibody that specifically binds to a target molecule can be referred to as a target-specific antibody. For example, an antibody that specifically binds to an IL2RA molecule can be referred to as an IL2RA-specific antibody or an anti-IL2RA antibody.

[0076] As used herein, the terms "polypeptide," "peptide," and "protein" are used interchangeably to refer to amino acid polymers of any length comprising at least two amino acids.

[0077] As used herein, the terms "polynucleotide," "nucleic acid molecule," and "nucleic acid sequence" are used interchangeably to refer to a nucleotide polymer of any length comprising at least two nucleotides, including but not limited to DNA, RNA, DNA / RNA hybrids, and modified forms thereof.

[0078] 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. Methods and materials for use in the present invention are described herein; other applicable methods and materials known in the art may also be used. The materials, methods, and examples are illustrative only and not restrictive. All publications, patent applications, patents, sequences, database entries, and other references mentioned herein are incorporated herein by reference in their entirety. In the event of a conflict, the present specification (including definitions) shall prevail.

[0079] Other features and advantages of the invention will be apparent from the following detailed description, the accompanying drawings, and the claims. BRIEF DESCRIPTION OF THE DRAWINGS

[0080] Figures 1A-1B Shown in anti-IL2RA antibodies 11C3, 11C12, 11D2, 13F9, 14C2 ( Figure 1A );5D9、7B5( Figure 1B ) in the presence of IL2RA ligands. Daclizumab analog was used as a control.

[0081] Figures 2A-2BThe results showed that the anti-IL2RA antibodies 11C3, 13F9, 11D2, 14C2, 11C12 ( Figure 2A );5D9、7B5( Figure 2B Blockade of mouse IL2 protein binding to IL2 reporter cells in the presence of α-glucose phosphate-buffered saline (BPA). A daclizumab analogue was used as a control.

[0082] Figures 3A-3B Shown in human CD3 + The blocking effect of STAT5 phosphorylation in T cells was observed in the cells and the cells were treated with anti-IL2RA antibodies 11C12, 11D2, 11C3, 13F9, 14C2 ( Figure 3A );7B5、5D9( Figure 3B ) and human IL2 (hIL2) were co-incubated. Daclizumab analogs were used as pSTAT5 blocking antibody controls, and 7G7B6 analog-SI was used as pSTAT5 non-blocking antibody controls.

[0083] Figure 4 Shown are changes in tumor volume over time in B-hIL2RA mice injected with MC38 cancer cells and treated with PBS (Group G1), 3 mg / kg 7B5 (Group G2), 10 mg / kg 7B5 (Group G3), 3 mg / kg 5D9 (Group G4), 10 mg / kg 5D9 (Group G5), 3 mg / kg 7G7B6 analog (Group G6), or 10 mg / kg 7G7B6 analog (Group G7).

[0084] Figure 5 Shown are the changes in tumor volume over time in B-hIL2RA mice injected with MC38 cancer cells and treated with PBS (Group G1), 10 mg / kg 7B5 (Group G2), 10 mg / kg 5D9 (Group G3), 10 mg / kg 7G7B6 analog-SI (Group G4), or 10 mg / kg 7G7B6 analog (Group G5).

[0085] Figure 6 Shown are changes in tumor volume over time in B-hIL2RA mice injected with MC38 cancer cells and treated with PBS (Group G1), 10 mg / kg 7G7B6 analog-SI (Group G2), 10 mg / kg 11C3 (Group G3), 10 mg / kg 13F9 (Group G4), 10 mg / kg 11D2 (Group G5), 10 mg / kg 11C12 (Group G6), or 10 mg / kg 14C2 (Group G7).

[0086] Figure 7 Kabat CDR sequences of anti-IL2RA antibodies are listed.

[0087] Figure 8 Chothia CDR sequences of anti-IL2RA antibodies are listed.

[0088] Figure 9 The heavy chain variable region and light chain variable region sequences of anti-IL2RA antibodies are listed.

[0089] Figure 10 The amino acid sequences contemplated by the present disclosure are listed. DETAILED DESCRIPTION

[0090] Interleukin-2 (IL-2) is a cytokine that maintains the proliferative potential of T lymphocytes. IL-2 has been identified as the main growth factor for activated T lymphocytes, which can drive clonal expansion and effector cell maturation. IL-2 stimulation can also lead to the growth of natural killer (NK) cells. IL-2 mediates its biological effects through the IL-2 receptor (IL-2R) complex. IL-2R is composed of three different subunits: α chain, β chain and γ chain. IL-2R and activated CD4 - 、CD8 - 、CD4 + 8 + 、CD4 + and CD8 + It is related to the proliferation of T cells. The importance of IL2RA is demonstrated by its high affinity for IL-2. The signaling pathway involving IL-2R includes Jak3-dependent activation of Jak1, which has been shown to be a key proliferation signal in fibroblasts. Elevated expression of sIL2RA and IL2RA protein in tumor cells has been detected in various cancers (such as lung cancer, melanoma, prostate cancer, esophageal squamous cell carcinoma, leukemia, lymphoma, ovarian cancer, colorectal cancer and breast cancer). In addition, elevated expression of IL2RA mRNA and protein has also been observed in various solid tumor types such as ovarian cancer, lung cancer, head and neck cancer and breast cancer. Therefore, anti-IL2RA antibodies have the potential to be used as cancer therapy.

[0091] The present disclosure provides examples of antibodies and antigen-binding fragments thereof that bind to IL2RA (interleukin-2 receptor alpha chain).

[0092] IL2RA

[0093] IL2RA (interleukin-2 receptor alpha chain, or CD25) is a type I transmembrane protein found on the surface of activated T cells, activated B cells, certain thymocytes, myeloid precursor cells, oligodendrocytes, and other cells. While IL2RA has been used as a marker for CD4+FoxP3+ regulatory T cells in mice, it has been found that the majority of resting memory T cells in humans constitutively express IL2RA. IL2RA is expressed in most B cell tumors, some acute nonlymphocytic leukemias, neuroblastoma, mastocytosis, and tumor-infiltrating lymphocytes. It serves as a receptor for the human immunodeficiency virus (HTLV-1) and is therefore expressed on tumor cells in adult T cell lymphomas / leukemias. Its soluble form, sIL-2R, may be elevated in these diseases and is occasionally used to track disease progression.

[0094] The interleukin-2 (IL-2) receptor, composed of α (IL-2RA, CD25), β (IL-2RB, CD122), and a common γ chain (IL-2RG, CD132) subunits, plays a key role in maintaining the immune system. The interleukin-2 (IL2) receptor α (IL2RA) and β (IL2RB) chains, along with the common γ chain (IL2RG), form the high-affinity IL2 receptor. The homodimeric α chain (IL2RA) forms a low-affinity receptor, while the homodimeric β (IL2RB) chain produces an intermediate-affinity receptor. The high-affinity receptor for IL-2 contains all three subunits (α, β, and γ) and is found on the surface of activated T cells, activated B cells, and regulatory T cells (Tregs). The intermediate-affinity receptor, composed solely of the γ and β chains, is expressed on NK cells and resting T and B cells. The low-affinity receptor, composed of the α chain, is expressed on dendritic cells.

[0095] Among IL-2 receptors, IL2RA is the only subunit that specifically binds to IL-2, while CD132 binds to common cytokines of the γc family (IL-4, IL-7, IL-9, IL-15, and IL-21), and the CD122 subunit binds to IL-15. Regulatory T cells (Tregs) constitutively overexpress IL2RA, making them the first responders to IL-2 in immune responses. They amplify IL-2 signaling in a STAT5-dependent manner to promote FOXP3 transcription. Notably, single nucleotide polymorphisms (SNPs) in the IL2RA gene have been associated with various autoimmune diseases, indicating that IL-2 signaling through IL2RA is a key axis in regulating immune tolerance. IL2RA is also crucial for the expansion of effector T cells in response to IL-2 following antigen stimulation.

[0096] The formation of the high-affinity quaternary IL-2 / IL-2R complex mediates signaling through the tyrosine kinases Jak1 and Jak3, which bind to IL-2Rβ and γc, respectively. Phosphorylation of three tyrosine residues in the cytoplasmic tail of IL-2Rβ (Y338 in human; Y341 in mouse) promotes the recruitment of the adaptor protein Shc, which activates the MAPK and PI-3K kinase pathways and primarily activates the Stat5 transcription factor (Y392 and Y510 in human; Y398 and Y505 in mouse), ultimately leading to Stat5-dependent gene regulation. The quaternary IL-2-IL-2R complex is rapidly internalized, with IL-2, IL-2Rβ, and γc rapidly degraded, while IL-2Rα recycles to the cell surface. Therefore, functional activities requiring sustained IL-2R signaling require a continuous supply of IL-2 to bind to IL-2Rα and form new IL-2-IL-2R signaling complexes.

[0097] IL2RA is a protein encoded by the human IL2RA gene. Normally, it is an integral membrane protein, but its soluble form, sIL2RA, has been isolated and demonstrated to be derived from extracellular proteolysis. Alternative splicing variants of IL2RA mRNA have been isolated, but their functional significance remains unclear. Mutations in this gene are associated with interleukin-2 receptor α deficiency.

[0098] For a detailed description of IL2RA and its functions, see, for example, Goudy, Kevin, et al., "Human IL2RA null mutation mediates immunodeficiency with lymphoproliferation and autoimmunity." Clinical Immunology 146.3 (2013): 248-261; Kuhn, Deborah J., and Q. Ping Dou. "The role of interleukin-2 receptor alpha in cancer." Front Biosci 10 (2005): 1462-1474; Malek, Thomas R., and Iris Castro. "Interleukin-2 receptor signaling: at the interface between tolerance and immunity." Immunity 33.2 (2010): 153-165; and Jiang et al., "Role of IL-2 in cancer immunotherapy." Oncoimmunology 5.6 (2016): e1163462; each of which is incorporated herein by reference in its entirety.

[0099] The present disclosure provides anti-IL2RA antibodies and antigen-binding fragments thereof, as well as methods of using such antibodies and antigen-binding fragments to inhibit tumor growth and treat various diseases, including cancer.

[0100] Anti-IL2RA antibodies and antigen-binding fragments thereof

[0101] The present disclosure provides antibodies and antigen-binding fragments thereof that specifically bind to IL2RA (e.g., human IL2RA). The antibodies and antigen-binding fragments described herein are capable of binding to IL2RA. In some embodiments, these antibodies do not block the binding of human IL2RA to IL2RA ligands (e.g., human IL2). In some embodiments, these antibodies do not block the binding of IL2 protein to IL2 reporter cells. In some embodiments, these antibodies do not significantly inhibit human IL2-induced STAT5 phosphorylation. In some embodiments, these antibodies do not block the IL2 / IL2RA signaling pathway to enhance the immune response. In some embodiments, these antibodies can induce complement-dependent cytotoxicity (CDC) or antibody-dependent cell-mediated cytotoxicity (ADCC). In some embodiments, these antibodies bind to cells expressing IL2RA.

[0102] The present disclosure provides, for example, anti-IL2RA antibodies 5D9, 7B5, 11C3, 11C12, 11D2, 13F9, and 14C2, modified antibodies thereof including, for example, chimeric antibodies, humanized antibodies, and human antibodies.

[0103] The CDR sequences of 5D9 and 5D9-derived antibodies (e.g., humanized antibodies) include the heavy chain variable domain CDRs according to the Kabat definition: SEQ ID NOs: 1, 2, and 3, and the light chain variable domain CDRs: SEQ ID NOs: 43, 44, and 45. CDRs may also be defined according to the Chothia system. According to the Chothia system, the heavy chain variable domain CDR sequences are SEQ ID NOs: 22, 23, and 24, and the light chain variable domain CDR sequences are SEQ ID NOs: 43, 44, and 45.

[0104] The CDR sequences of 7B5 and 7B5-derived antibodies include the heavy chain variable domain CDRs according to the Kabat definition: SEQ ID NOs: 4, 5, and 6, and the light chain variable domain CDRs: SEQ ID NOs: 46, 47, and 48. According to the Chothia definition, the heavy chain variable domain CDR sequences are SEQ ID NOs: 25, 26, and 27, and the light chain variable domain CDR sequences are SEQ ID NOs: 46, 47, and 48.

[0105] The CDR sequences of 11C3 and 11C3-derived antibodies include the heavy chain variable domain CDRs according to the Kabat definition: SEQ ID NOs: 7, 8, and 9, and the light chain variable domain CDRs: SEQ ID NOs: 49, 50, and 51. According to the Chothia definition, the heavy chain variable domain CDR sequences are SEQ ID NOs: 28, 29, and 30, and the light chain variable domain CDR sequences are SEQ ID NOs: 49, 50, and 51.

[0106] The CDR sequences of 11C12 and 11C12-derived antibodies include the Kabat-defined heavy chain variable domain CDRs of SEQ ID NOs: 10, 11, and 12, and the Chothia-defined light chain variable domain CDRs of SEQ ID NOs: 31, 32, and 33, respectively, and the Chothia-defined light chain variable domain CDRs of SEQ ID NOs: 49, 50, and 51.

[0107] The CDR sequences of 11D2 and 11D2-derived antibodies include the heavy chain variable domain CDRs according to the Kabat definition: SEQ ID NOs: 13, 14, and 15, and the light chain variable domain CDRs: SEQ ID NOs: 49, 50, and 51. According to the Chothia definition, the heavy chain variable domain CDR sequences are SEQ ID NOs: 34, 35, and 36, and the light chain variable domain CDR sequences are SEQ ID NOs: 49, 50, and 51.

[0108] The CDR sequences of 13F9 and 13F9-derived antibodies include the heavy chain variable domain CDRs according to the Kabat definition: SEQ ID NOs: 16, 17, and 18, and the light chain variable domain CDRs: SEQ ID NOs: 49, 50, and 51. According to the Chothia definition, the heavy chain variable domain CDR sequences are SEQ ID NOs: 37, 38, and 39, and the light chain variable domain CDR sequences are SEQ ID NOs: 49, 50, and 51.

[0109] The CDR sequences of 14C2 and 14C2-derived antibodies include the heavy chain variable domain CDRs according to the Kabat definition: SEQ ID NOs: 19, 20, and 21, and the light chain variable domain CDRs: SEQ ID NOs: 49, 50, and 51. According to the Chothia definition, the heavy chain variable domain CDR sequences are SEQ ID NOs: 40, 41, and 42, and the light chain variable domain CDR sequences are SEQ ID NOs: 49, 50, and 51.

[0110] The amino acid sequence of the heavy chain variable region of the 5D9 antibody is shown in SEQ ID NO: 52. The amino acid sequence of the light chain variable region of the 5D9 antibody is shown in SEQ ID NO: 53.

[0111] The amino acid sequence of the heavy chain variable region of the 7B5 antibody is shown in SEQ ID NO: 54. The amino acid sequence of the light chain variable region of the 7B5 antibody is shown in SEQ ID NO: 55.

[0112] The amino acid sequence of the heavy chain variable region of the 11C3 antibody is shown in SEQ ID NO: 56. The amino acid sequence of the light chain variable region of the 11C3 antibody is shown in SEQ ID NO: 61.

[0113] The amino acid sequence of the heavy chain variable region of the 11C12 antibody is shown in SEQ ID NO: 57. The amino acid sequence of the light chain variable region of the 11C12 antibody is shown in SEQ ID NO: 61.

[0114] The amino acid sequence of the heavy chain variable region of the 11D2 antibody is shown in SEQ ID NO: 58. The amino acid sequence of the light chain variable region of the 11D2 antibody is shown in SEQ ID NO: 61.

[0115] The amino acid sequence of the heavy chain variable region of the 13F9 antibody is shown in SEQ ID NO: 59. The amino acid sequence of the light chain variable region of the 13F9 antibody is shown in SEQ ID NO: 61.

[0116] The amino acid sequence of the heavy chain variable region of the 14C2 antibody is shown in SEQ ID NO: 60. The amino acid sequence of the light chain variable region of the 14C2 antibody is shown in SEQ ID NO: 61.

[0117] The amino acid sequences of the heavy and light chain variable regions of the modified antibodies are also provided. In some embodiments, the heavy chain variable region is at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to SEQ ID NO: 52, 54, 56, 57, 58, 59, or 60. In some embodiments, the light chain variable region is at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to SEQ ID NO: 53, 55, or 61. The heavy chain variable region sequence can be paired with the corresponding light chain variable region sequence to jointly bind to IL2RA.

[0118] In some embodiments, the variable region is fully humanized, e.g., derived from a human heavy chain immunoglobulin locus sequence (e.g., a recombination of human IGHV, IGHD, and IGHJ genes) and / or a human kappa chain immunoglobulin locus sequence (e.g., a recombination of human IGKV and IGKJ genes).

[0119] In addition, in some embodiments, the antibodies or antigen-binding fragments thereof described herein may further comprise one, two or three heavy chain variable region CDRs selected from the following sequence group: SEQ ID NO: 1-3, SEQ ID NO: 4-6, SEQ ID NO: 7-9, SEQ ID NO: 10-12, SEQ ID NO: 13-15, SEQ ID NO: 16-18, SEQ ID NO: 19-21, SEQ ID NO: 22-24, SEQ ID NO: 25-27, SEQ ID NO: 28-30, SEQ ID NO: 31-33, SEQ ID NO: 34-36, SEQ ID NO: 37-39 and SEQ ID NO: 40-42; and / or one, two or three light chain variable region CDRs selected from the following sequence group: SEQ ID NO: 43-45, SEQ ID NO: 46-48 and SEQ ID NO: 49-51.

[0120] In some embodiments, the antibody may have a heavy chain variable region (VH) comprising complementarity determining regions (CDR) 1, 2, and 3, wherein the CDR1 region comprises or consists of an amino acid sequence that is at least 80%, 85%, 90%, or 95% identical to a selected VH CDR1 amino acid sequence; the CDR2 region comprises or consists of an amino acid sequence that is at least 80%, 85%, 90%, or 95% identical to a selected VH CDR2 amino acid sequence; and the CDR3 region comprises or consists of an amino acid sequence that is at least 80%, 85%, 90%, or 95% identical to a selected VH CDR3 amino acid sequence. In some embodiments, the antibody may have a light chain variable region (VL) comprising CDR 1, 2, 3, wherein the CDR1 region comprises or consists of an amino acid sequence that is at least 80%, 85%, 90% or 95% identical to a selected VL CDR1 amino acid sequence; the CDR2 region comprises or consists of an amino acid sequence that is at least 80%, 85%, 90% or 95% identical to a selected VLCDR2 amino acid sequence; and the CDR3 region comprises or consists of an amino acid sequence that is at least 80%, 85%, 90% or 95% identical to a selected VL CDR3 amino acid sequence. Selected VH CDR 1, 2, 3 amino acid sequences and selected VL CDR 1, 2, 3 amino acid sequences are shown in Figure 7 (CDRs according to Kabat definition) and Figure 8 (CDRs under Chothia definition).

[0121] In some embodiments, the antibodies or antigen-binding fragments thereof described herein may comprise a heavy chain variable domain comprising one, two, or three of the CDRs of SEQ ID NO: 1 having 0, 1, or 2 amino acid insertions, deletions, or substitutions; one, two, or three of the CDRs of SEQ ID NO: 2 having 0, 1, or 2 amino acid insertions, deletions, or substitutions; or one, two, or three of the CDRs of SEQ ID NO: 3 having 0, 1, or 2 amino acid insertions, deletions, or substitutions.

[0122] In some embodiments, the antibodies or antigen-binding fragments thereof described herein may comprise a heavy chain variable domain comprising one, two, or three of the CDRs of SEQ ID NO: 4 having 0, 1, or 2 amino acid insertions, deletions, or substitutions; one, two, or three of the CDRs of SEQ ID NO: 5 having 0, 1, or 2 amino acid insertions, deletions, or substitutions; or one, two, or three of the CDRs of SEQ ID NO: 6 having 0, 1, or 2 amino acid insertions, deletions, or substitutions.

[0123] In some embodiments, the antibodies or antigen-binding fragments thereof described herein may comprise a heavy chain variable domain comprising one, two, or three of the CDRs of SEQ ID NO: 7 having 0, 1, or 2 amino acid insertions, deletions, or substitutions; one, two, or three of the CDRs of SEQ ID NO: 8 having 0, 1, or 2 amino acid insertions, deletions, or substitutions; or one, two, or three of the CDRs of SEQ ID NO: 9 having 0, 1, or 2 amino acid insertions, deletions, or substitutions.

[0124] In some embodiments, the antibodies or antigen-binding fragments thereof described herein may comprise a heavy chain variable domain comprising one, two, or three of the CDRs of SEQ ID NO: 10 having 0, 1, or 2 amino acid insertions, deletions, or substitutions; one, two, or three of the CDRs of SEQ ID NO: 11 having 0, 1, or 2 amino acid insertions, deletions, or substitutions; or one, two, or three of the CDRs of SEQ ID NO: 12 having 0, 1, or 2 amino acid insertions, deletions, or substitutions.

[0125] In some embodiments, the antibodies or antigen-binding fragments thereof described herein may comprise a heavy chain variable domain comprising one, two, or three of the CDRs of SEQ ID NO: 13 having 0, 1, or 2 amino acid insertions, deletions, or substitutions; one, two, or three of the CDRs of SEQ ID NO: 14 having 0, 1, or 2 amino acid insertions, deletions, or substitutions; or one, two, or three of the CDRs of SEQ ID NO: 15 having 0, 1, or 2 amino acid insertions, deletions, or substitutions.

[0126] In some embodiments, the antibodies or antigen-binding fragments thereof described herein may comprise a heavy chain variable domain comprising one, two, or three of the CDRs of SEQ ID NO: 16 having 0, 1, or 2 amino acid insertions, deletions, or substitutions; one, two, or three of the CDRs of SEQ ID NO: 17 having 0, 1, or 2 amino acid insertions, deletions, or substitutions; or one, two, or three of the CDRs of SEQ ID NO: 18 having 0, 1, or 2 amino acid insertions, deletions, or substitutions.

[0127] In some embodiments, the antibodies or antigen-binding fragments thereof described herein may comprise a heavy chain variable domain comprising one, two, or three of the CDRs of SEQ ID NO: 19 having 0, 1, or 2 amino acid insertions, deletions, or substitutions; one, two, or three of the CDRs of SEQ ID NO: 20 having 0, 1, or 2 amino acid insertions, deletions, or substitutions; or one, two, or three of the CDRs of SEQ ID NO: 21 having 0, 1, or 2 amino acid insertions, deletions, or substitutions.

[0128] In some embodiments, the antibodies or antigen-binding fragments thereof described herein may comprise a heavy chain variable domain comprising one, two, or three of the CDRs of SEQ ID NO: 22 having 0, 1, or 2 amino acid insertions, deletions, or substitutions; one, two, or three of the CDRs of SEQ ID NO: 23 having 0, 1, or 2 amino acid insertions, deletions, or substitutions; or one, two, or three of the CDRs of SEQ ID NO: 24 having 0, 1, or 2 amino acid insertions, deletions, or substitutions.

[0129] In some embodiments, the antibodies or antigen-binding fragments thereof described herein may comprise a heavy chain variable domain comprising one, two, or three of the CDRs of SEQ ID NO: 25 having 0, 1, or 2 amino acid insertions, deletions, or substitutions; one, two, or three of the CDRs of SEQ ID NO: 26 having 0, 1, or 2 amino acid insertions, deletions, or substitutions; or one, two, or three of the CDRs of SEQ ID NO: 27 having 0, 1, or 2 amino acid insertions, deletions, or substitutions.

[0130] In some embodiments, the antibodies or antigen-binding fragments thereof described herein may comprise a heavy chain variable domain comprising one, two, or three of the CDRs of SEQ ID NO: 28 having 0, 1, or 2 amino acid insertions, deletions, or substitutions; one, two, or three of the CDRs of SEQ ID NO: 29 having 0, 1, or 2 amino acid insertions, deletions, or substitutions; or one, two, or three of the CDRs of SEQ ID NO: 30 having 0, 1, or 2 amino acid insertions, deletions, or substitutions.

[0131] In some embodiments, the antibodies or antigen-binding fragments thereof described herein may comprise a heavy chain variable domain comprising one, two, or three of the CDRs of SEQ ID NO: 31 having 0, 1, or 2 amino acid insertions, deletions, or substitutions; one, two, or three of the CDRs of SEQ ID NO: 32 having 0, 1, or 2 amino acid insertions, deletions, or substitutions; or one, two, or three of the CDRs of SEQ ID NO: 33 having 0, 1, or 2 amino acid insertions, deletions, or substitutions.

[0132] In some embodiments, the antibodies or antigen-binding fragments thereof described herein may comprise a heavy chain variable domain comprising one, two, or three of the CDRs of SEQ ID NO: 34 having 0, 1, or 2 amino acid insertions, deletions, or substitutions; one, two, or three of the CDRs of SEQ ID NO: 35 having 0, 1, or 2 amino acid insertions, deletions, or substitutions; or one, two, or three of the CDRs of SEQ ID NO: 36 having 0, 1, or 2 amino acid insertions, deletions, or substitutions.

[0133] In some embodiments, the antibodies or antigen-binding fragments thereof described herein may comprise a heavy chain variable domain comprising one, two, or three of the CDRs of SEQ ID NO: 37 having 0, 1, or 2 amino acid insertions, deletions, or substitutions; one, two, or three of the CDRs of SEQ ID NO: 38 having 0, 1, or 2 amino acid insertions, deletions, or substitutions; or one, two, or three of the CDRs of SEQ ID NO: 39 having 0, 1, or 2 amino acid insertions, deletions, or substitutions.

[0134] In some embodiments, the antibodies or antigen-binding fragments thereof described herein may comprise a heavy chain variable domain comprising one, two, or three of the CDRs of SEQ ID NO: 40 having 0, 1, or 2 amino acid insertions, deletions, or substitutions; one, two, or three of the CDRs of SEQ ID NO: 41 having 0, 1, or 2 amino acid insertions, deletions, or substitutions; or one, two, or three of the CDRs of SEQ ID NO: 42 having 0, 1, or 2 amino acid insertions, deletions, or substitutions.

[0135] In some embodiments, the antibodies or antigen-binding fragments thereof described herein may comprise a light chain variable domain comprising one, two, or three of the CDRs of SEQ ID NO: 43 having 0, 1, or 2 amino acid insertions, deletions, or substitutions; one, two, or three of the CDRs of SEQ ID NO: 44 having 0, 1, or 2 amino acid insertions, deletions, or substitutions; or one, two, or three of the CDRs of SEQ ID NO: 45 having 0, 1, or 2 amino acid insertions, deletions, or substitutions.

[0136] In some embodiments, the antibodies or antigen-binding fragments thereof described herein may comprise a light chain variable domain comprising one, two, or three of the CDRs of SEQ ID NO: 46 having 0, 1, or 2 amino acid insertions, deletions, or substitutions; one, two, or three of the CDRs of SEQ ID NO: 47 having 0, 1, or 2 amino acid insertions, deletions, or substitutions; or one, two, or three of the CDRs of SEQ ID NO: 48 having 0, 1, or 2 amino acid insertions, deletions, or substitutions.

[0137] In some embodiments, the antibodies or antigen-binding fragments thereof described herein may comprise a light chain variable domain comprising one, two, or three of the CDRs of SEQ ID NO: 49 having 0, 1, or 2 amino acid insertions, deletions, or substitutions; one, two, or three of the CDRs of SEQ ID NO: 50 having 0, 1, or 2 amino acid insertions, deletions, or substitutions; or one, two, or three of the CDRs of SEQ ID NO: 51 having 0, 1, or 2 amino acid insertions, deletions, or substitutions.

[0138] The insertions, deletions, and substitutions may be located within the CDR sequence or at one or both ends of the CDR sequence. In some embodiments, the CDRs are defined based on the Kabat definition scheme. In some embodiments, the CDRs are defined based on the Chothia definition scheme. In some embodiments, the CDRs are defined based on a combination of the Kabat and Chothia definition schemes. In some embodiments, the CDRs are defined based on the IMGT definition. In some embodiments, the CDRs are defined based on the contact definition.

[0139] The present disclosure also provides antibodies or antigen-binding fragments thereof that bind to IL2RA. The antibodies or antigen-binding fragments thereof comprise a heavy chain variable region (VH) comprising or consisting of an amino acid sequence that is at least 80%, 85%, 90%, or 95% identical to a selected VH sequence; and a light chain variable region (VL) comprising or consisting of an amino acid sequence that is at least 80%, 85%, 90%, or 95% identical to a selected VL sequence. In some embodiments, the selected VH sequence is SEQ ID NO: 52 and the selected VL sequence is SEQ ID NO: 53. In some embodiments, the selected VH sequence is SEQ ID NO: 54 and the selected VL sequence is SEQ ID NO: 55. In some embodiments, the selected VH sequence is SEQ ID NO: 56 and the selected VL sequence is SEQ ID NO: 61. In some embodiments, the selected VH sequence is SEQ ID NO: 57 and the selected VL sequence is SEQ ID NO: 61. In some embodiments, the selected VH sequence is SEQ ID NO: 58 and the selected VL sequence is SEQ ID NO: 61. In some embodiments, the selected VH sequence is SEQ ID NO: 59 and the selected VL sequence is SEQ ID NO: 61. In some embodiments, the selected VH sequence is SEQ ID NO: 60 and the selected VL sequence is SEQ ID NO: 61.

[0140] To determine the percent identity of two amino acid sequences or two nucleic acid sequences, the sequences need to be optimally aligned (e.g., gaps can be introduced in the first and / or second amino acid sequence or nucleic acid sequence to optimize alignment, and non-homologous sequences can be ignored). The length of the reference sequence used for alignment is at least 80% of the length of the reference sequence, in some embodiments at least 90%, 95%, or 100%. The amino acid residues or nucleotides at corresponding amino acid positions or nucleotide positions are then compared. When the amino acid residue or nucleotide at a position in the first sequence is the same as the corresponding position in the second sequence, the molecules are identical at that position. The percent identity of the two sequences is a function of the number of identical positions shared by the sequences, taking into account the number of gaps and the length of each gap introduced to optimize the alignment of the two sequences. For example, sequence comparison and percent identity determination can be accomplished using the Blossum 62 scoring matrix, with a gap penalty of 12, a gap extension penalty of 4, and a frameshift gap penalty of 5.

[0141] The present disclosure also provides nucleic acids comprising polynucleotides encoding polypeptides comprising an immunoglobulin heavy chain or an immunoglobulin light chain. Figure 7 or Figure 8 The CDRs shown, or having Figure 9 When the polypeptide is paired with a corresponding polypeptide (e.g., a corresponding heavy chain variable region or a corresponding light chain variable region), the paired polypeptide binds to IL2RA.

[0142] Anti-IL2RA antibodies and antigen-binding fragments thereof may also be variants (including derivatives and conjugates) of antibodies or antibody fragments, multispecific (e.g., bispecific) antibodies, or antibody fragments. Other antibodies provided herein include polyclonal antibodies, monoclonal antibodies, multimeric antibodies, multispecific (e.g., bispecific) antibodies, human antibodies, chimeric antibodies (e.g., human-mouse chimeras), single-chain antibodies, intracellularly produced antibodies (i.e., intrabodies), and antigen-binding fragments thereof. The antibody or antigen-binding fragment thereof may be of any type (e.g., IgG, IgE, IgM, IgD, IgA, and IgY), class (e.g., IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2), or subclass. In some embodiments, the antibody or antigen-binding fragment thereof is an IgG antibody or antigen-binding fragment thereof.

[0143] Antibody fragments are suitable for use in the methods provided herein as long as they retain the desired affinity and specificity of the full-length antibody. Thus, antibody fragments that bind to IL2RA will retain the ability to bind to IL2RA. Fv fragments are antibody fragments that contain a complete antigen recognition and binding site. This region consists of a single heavy-chain variable domain and a light-chain variable domain dimer in close association, which can be covalently bound (e.g., in a scFv). In this configuration, the three CDRs of each variable domain interact to form an antigen-binding site on the surface of the VH-VL dimer. The six CDRs, or a subset thereof, collectively confer antigen-binding specificity to the antibody. However, even a single variable domain (or half an Fv containing only three antigen-specific CDRs) has the ability to recognize and bind antigen, although typically with lower affinity than the complete binding site. Single-chain Fv (scFv) antibody fragments contain the VH and VL domains (or regions) of an antibody, present in a single polypeptide chain. Typically, scFv polypeptides also contain a polypeptide linker between the VH and VL domains, enabling the scFv to form the desired structure for antigen binding.

[0144] The present disclosure also provides antibodies or antigen-binding fragments thereof that cross-compete with any of the antibodies or antigen-binding fragments described herein. Cross-competition assays are known in the art, for example, as described in Moore et al., "Antibody cross-competition analysis of the human immunodeficiency virustype 1gp120 exterior envelope glycoprotein." Journal of Virology 70.3 (1996): 1863-1872, the entirety of which is incorporated herein by reference. In one aspect, the present disclosure also provides antibodies or antigen-binding fragments thereof that bind to the same epitope or region as any of the antibodies or antigen-binding fragments described herein. Epitope sorting assays are known in the art, for example, as described in Estep et al. "High throughput solution-based measurement of antibody-antigen affinity and epitope binning." MAbs. Vol. 5. No. 2. Taylor & Francis, 2013, the entirety of which is incorporated herein by reference.

[0145] Antibodies and their antigen-binding fragments

[0146] The present disclosure provides various antibodies and antigen-binding fragments thereof derived from the anti-IL2RA antibodies described herein. Generally, antibodies (also known as immunoglobulins) are composed of two types of polypeptide chains: light chains and heavy chains. A non-limiting example of an antibody disclosed herein may be a complete four-chain immunoglobulin antibody comprising two heavy chains and two light chains. The antibody heavy chains may be of any isotype, including IgM, IgG, IgE, IgA, or IgD, or subisotypes including IgG1, IgG2, IgG2a, IgG2b, IgG3, IgG4, IgE1, IgE2, and the like. The light chains may be kappa or lambda light chains. An antibody may comprise two identical copies of a light chain and two identical copies of a heavy chain. Each heavy chain comprises a variable domain (or variable region, VH) and multiple constant domains (or constant regions), which are bound to each other by disulfide bonds within their constant domains to form the "backbone" of the antibody. Each light chain comprises a variable domain (or variable region, VL) and a constant domain (or constant region), each bound to a heavy chain by disulfide bonds. The variable domain of each light chain is aligned with the variable domain of the heavy chain to which it is bound.The variable domains of both the light and heavy chains are composed of three hypervariable regions flanked by more conserved framework regions (FR).

[0147] These hypervariable regions, called complementarity-determining regions (CDRs), form a loop-like structure that makes up the antigen-binding surface of antibodies. The four framework regions primarily adopt a β-sheet conformation, with the CDRs forming loops connecting and, in some cases, forming part of the β-sheet structure. The CDRs in each chain are closely juxtaposed by the framework regions and, together with the CDRs from the other chain, contribute to the formation of the antigen-binding region.

[0148] Methods for identifying CDR regions by analyzing the amino acid sequence of an antibody are well known in the art, and there are multiple commonly used CDR definitions: the Kabat definition is based on sequence variability, and the Chothia definition is based on the position of the structural loop regions. These methods and definitions are described, for example, in: Martin, "Protein sequence and structure analysis of antibody variable domains," Antibody engineering, Springer Berlin Heidelberg, 2001. 422-439; Abhinandan, et al. "Analysis and improvements to Kabat and structurally correct numbering of antibody variable domains," Molecular immunology 45.14(2008):3832-3839; Wu, TT and Kabat, EA (1970) J. Exp. Med. 132:211-250; Martin et al., Methods Enzymol. 203:121-53(1991); Morea et al., Biophys Chem. 68(1-3):9-16(Oct. 1997); Morea et al., J Mol Biol. 275(2):269-94(Jan. 1998); Chothia et al. al., Nature 342(6252):877-83 (Dec. 1989); Ponomarenko and Bourne, BMC Structural Biology 7:64 (2007); each of which is incorporated herein by reference in its entirety.

[0149] CDRs are crucial for recognizing antigenic epitopes. As used herein, "epitope" refers to the smallest portion of a target molecule that can be specifically bound by an antibody's antigen-binding domain. An epitope can be as small as approximately three, four, five, six, or seven amino acids, but these amino acids need not lie in a continuous linear sequence within the antigen's primary structure, as the epitope may depend on the antigen's three-dimensional configuration based on its secondary and tertiary structures.

[0150] In some embodiments, the antibody is a complete immunoglobulin molecule (e.g., IgG1, IgG2a, IgG2b, IgG3, IgM, IgD, IgE, IgA). IgG subclasses (IgG1, IgG2, IgG3, IgG4) are highly conserved, with differences in their constant regions, particularly in the hinge region and upper CH2 domain. The sequences and differences of IgG subclasses are known in the art, as described, for example, in Vidarsson, et al, "IgG subclasses and allotypes: from structure to effector functions." Frontiers in immunology 5 (2014); Irani, et al. "Molecular properties of human IgG subclasses and their implications for designing therapeutic monoclonal antibodies against infectious diseases." Molecular immunology 67.2 (2015): 171-182; Shakib, Farouk, ed. The human IgG subclasses: molecular analysis of structure, function and regulation. Elsevier, 2016; each of which is incorporated herein by reference in its entirety.

[0151] The antibody may also be an immunoglobulin molecule derived from any species (e.g., humans, rodents, mice, camelids). Antibodies disclosed herein also include, but are not limited to, polyclonal antibodies, monoclonal antibodies, monospecific antibodies, multispecific antibodies, and chimeric antibodies comprising an immunoglobulin binding domain fused to another polypeptide. "Antigen binding domain" or "antigen binding fragment" refers to an antibody portion that retains the specific binding activity of an intact antibody, i.e., any antibody portion that is capable of specifically binding to an epitope on an intact antibody target molecule. It includes, for example, Fab, Fab', F(ab')2 and variants of these fragments. Therefore, in some embodiments, an antibody or its antigen-binding fragment may be, for example, scFv, Fv, Fd, dAb, bispecific antibodies, bispecific scFv, diabodies, linear antibodies, single-chain antibody molecules, multispecific antibodies formed from antibody fragments, and any polypeptide comprising an antibody binding domain or a binding domain homologous thereto. Non-limiting examples of antigen binding domains include, e.g., the heavy and / or light chain CDRs of an intact antibody, the heavy and / or light chain variable regions of an intact antibody, the full-length heavy or light chain of an intact antibody, or a single CDR within a heavy or light chain of an intact antibody.

[0152] Also provided herein are antibody fragments suitable for use in the methods. A Fab fragment comprises a light chain variable domain and constant domain, and a heavy chain variable domain and first constant domain (CH1). A F(ab')2 antibody fragment comprises a pair of Fab fragments, typically covalently linked near their carboxyl termini by a hinge region cysteine. Other chemical coupling methods for antibody fragments are also known in the art.

[0153] Diabodies are small antibody fragments with two antigen-binding sites, which contain a VH and a VL (VH and VL) linked in the same polypeptide chain. By using a linker that is too short (too short to allow pairing between the two domains on the same chain), these domains are forced to pair with the complementary domains of another chain, thereby forming two antigen-binding sites.

[0154] Linear antibodies consist of a pair of tandem Fd segments (VH-CH1-VH-CH1), which together with complementary light chain polypeptides form a pair of antigen-binding regions. Linear antibodies can be bispecific or monospecific.

[0155] The antibodies and antibody fragments of the present disclosure may be modified in the Fc region to provide desired effector function or serum half-life.

[0156] Antibody multimerization can be achieved through natural antibody aggregation or chemical / recombinant ligation techniques known in the art. For example, partially purified antibody preparations (such as purified IgG1 molecules) will spontaneously form protein aggregates containing antibody homodimers and other higher-order antibody multimers.

[0157] Alternatively, antibody homodimers can be formed by chemical connection techniques known in the art. For example, heterobifunctional cross-linkers (including but not limited to SMCC (succinimidyl-4-(maleimidomethyl)cyclohexane-1-carboxylate) and SATA (N-succinimidyl-S-acetylthioacetate)) can be used to form antibody multimers. An exemplary scheme for the preparation of antibody homodimers is described in Ghetie et al. (Proc. Natl. Acad. Sci. USA 94:7509-7514, 1997). Antibody homodimers can be converted into Fab'2 homodimers by pepsin digestion. Another method for forming antibody homodimers is to use the self-affinity T15 peptide described in Zhao et al. (J. Immunol. 25:396-404, 2002).

[0158] In some embodiments, the multispecific antibody is a bispecific antibody. Bispecific antibodies can optimize the heterodimer ratio recovered from recombinant cell culture by transforming the interface of a pair of antibody molecules. For example, the interface may comprise at least a portion of the CH3 domain of an antibody constant domain. In this method, one or more small amino acid side chains on the interface of the first antibody molecule are replaced with large side chains (e.g., tyrosine or tryptophan); simultaneously, on the interface of the second antibody molecule, by replacing the large amino acid side chains with small side chains (e.g., alanine or threonine), a compensatory "cavity" identical or similar in size to the large side chain is formed. This mechanism can increase heterodimer production and reduce by-products such as homodimers. This method is described in, for example, WO 96 / 27011, which is incorporated herein by reference in its entirety.

[0159] Bispecific antibodies include cross-linked or "heteroconjugate" antibodies. For example, one of the heteroconjugate antibodies can be conjugated to avidin and the other to biotin. Heteroconjugate antibodies can also be prepared using any convenient cross-linking method. Suitable cross-linking agents and cross-linking techniques are well known in the art and are disclosed in U.S. Patent No. 4,676,980 (incorporated herein by reference in its entirety).

[0160] Any of the antibodies or antigen-binding fragments described herein can be conjugated to a stabilizing molecule (e.g., a molecule that extends the half-life of the antibody or antigen-binding fragment thereof in a subject or in solution). Non-limiting examples of stabilizing molecules include polymers (e.g., polyethylene glycol) or proteins (e.g., serum albumin, such as human serum albumin). Conjugation to a stabilizing molecule can extend the half-life of the antibody or antigen-binding fragment in vitro (e.g., in tissue culture or storage as a pharmaceutical composition) or in vivo (e.g., in humans) or enhance its biological activity.

[0161] In some embodiments, the antibodies or antigen-binding fragments described herein can be coupled to therapeutic agents. Antibody-drug conjugates comprising the antibodies or antigen-binding fragments thereof can be covalently or non-covalently bound to therapeutic agents. In some embodiments, the therapeutic agent is a cytotoxic agent or a cytostatic agent (e.g., cytochalasin B, gramicidin D, ethidium bromide, emetine, mitomycin, etoposide, teniposide, vincristine, vinblastine, colchicine, doxorubicin, daunorubicin, dihydroxyanthraquinone, maytansine derivatives such as DM-1 and DM-4, ​​diketones, mitoxantrone, mithramycin, actinomycin D, 1-dehydrotestosterone, glucocorticoids, procaine, tetracaine, lidocaine, propranolol, puromycin, epirubicin, cyclophosphamide, and the like).

[0162] In some embodiments, the antigen binding fragment may constitute a component of a chimeric antigen receptor (CAR). In some embodiments, the chimeric antigen receptor is a fusion of a single-chain variable fragment (scFv) and a CD3-zeta transmembrane region and an intracellular domain as described herein. In some embodiments, the chimeric antigen receptor further comprises an intracellular signaling domain from a co-stimulatory protein receptor (e.g., CD28, 41BB, ICOS). In some embodiments, the chimeric antigen receptor comprises multiple signaling domains (e.g., CD3z-CD28-41BB or CD3z-CD28-OX40) to enhance efficacy. Therefore, in one aspect, the present disclosure also provides cells (e.g., T cells) expressing chimeric antigen receptors as described herein.

[0163] In some embodiments, the scFv has one heavy chain variable domain and one light chain variable domain. In some embodiments, the scFv has two heavy chain variable domains and two light chain variable domains.

[0164] In some embodiments, sequences of the antibodies or antigen-binding fragments thereof described herein (e.g., CDR or VH / VL sequences) can be used to generate bispecific antibodies targeting IL2RA and an additional antigen (e.g., OX40, CD3, 4-1BB, CD314, CD47, PD-1, CTLA4, CD40, or PDL1).

[0165] Antibody characteristics

[0166] In some embodiments, the antibodies or antigen-binding fragments thereof described herein do not block the binding of IL2RA to an IL2RA ligand (e.g., IL2). Thus, the antibodies do not inhibit the IL2RA signaling pathway by binding to IL2RA. In some embodiments, the antibodies can upregulate an immune response. In some embodiments, the antibodies can reduce tumor volume in an animal.

[0167] In some embodiments, the antibody (or antigen-binding fragment thereof) has a dissociation rate (koff) of less than 0.1 s -1 , less than 0.01s -1 , less than 0.001s -1 , less than 0.0001s -1 or less than 0.00001s -1 Specifically binds to IL2RA (e.g., human IL2RA or monkey IL2RA). In some embodiments, the off-rate (koff) is greater than 0.01 s -1 , greater than 0.001s -1 , greater than 0.0001s -1 , greater than 0.00001s -1 or greater than 0.000001s -1 .

[0168] In some embodiments, the kinetic binding rate (kon) is greater than 1×10 2 / Ms, greater than 1×10 3 / Ms, greater than 1×10 4 / Ms, greater than 1×10 5 / Ms or greater than 1×10 6 / Ms. In some embodiments, the kinetic binding rate (kon) is less than 1×10 5 / Ms, less than 1×10 6 / Ms or less than 1×10 7 / Ms.

[0169] Affinity can be derived from the kinetic rate constant quotient (KD = koff / kon). In some embodiments, KD is less than 1×10 -6 M, less than 1×10 -7 M, less than 1×10 -8 M, less than 1×10 -9 M or less than 1×10 -10 In some embodiments, the KD is less than 50 nM, 30 nM, 20 nM, 15 nM, 10 nM, 9 nM, 8 nM, 7 nM, 6 nM, 5 nM, 4 nM, 3 nM, 2 nM, or 1 nM. In some embodiments, the KD is greater than 1×10 -7 M, greater than 1×10 -8 M, greater than 1×10 -9 M, greater than 1×10 -10 M, greater than 1×10 -11 M or greater than 1×10 -12 M.

[0170] Common techniques for measuring antibody affinity for an antigen include, for example, ELISA, RIA, and surface plasmon resonance (SPR). In some embodiments, the antibody binds to human IL2RA (SEQ ID NO: 64) and / or monkey IL2RA (eg, SEQ ID NO: 65). In some embodiments, the antibody does not bind to human IL2RA and / or monkey IL2RA.

[0171] In some embodiments, the antibodies or antigen-binding fragments thereof described herein do not inhibit IL2-induced STAT5 phosphorylation, or inhibit it by less than 20%, less than 15%, less than 10%, less than 5%, or less than 1%.

[0172] In some embodiments, the tumor growth inhibition percentage (TGI) of the antibodies or antigen-binding fragments thereof described herein is TV In some embodiments, the antibody has a tumor growth inhibition percentage of less than 60%, 70%, 80%, 90%, 100%, 110%, 120%, 130%, 140%, 150%, 160%, 170%, 180%, 190%, or 200%. TGI% can be measured on day 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 after the start of treatment, or on month 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 after the start of treatment. As used herein, tumor growth inhibition percentage (TGI%) is calculated as follows:

[0173] TGI (%) = [1-(T i -T0) / (V i -V0)]×100

[0174] T i is the average tumor volume of the treatment group on day i. T0 is the average tumor volume of the treatment group on day zero. i is the average tumor volume of the control group on day i. V0 is the average tumor volume of the control group on day zero.

[0175] In some embodiments, the antibodies or antigen-binding fragments thereof described herein are IL2RA antagonists. In some embodiments, the antibodies or antigen-binding fragments thereof described herein are IL2RA agonists.

[0176] In some embodiments, the antibodies or antigen-binding fragments thereof described herein are non-toxic. In some embodiments, there is no significant difference in body weight between the treatment group and the control group (e.g., at a dose of 0.3 mg / kg, 1 mg / kg, 10 mg / kg, or 25 mg / kg).

[0177] In some embodiments, the antibody or antigen-binding fragment can induce complement-dependent cytotoxicity (CDC) and / or antibody-dependent cell-mediated cytotoxicity (ADCC) and kill tumor cells.

[0178] In some embodiments, the antibody or antigen-binding fragment has a functional Fc region. In some embodiments, the effector function of the functional Fc region is antibody-dependent cell-mediated cytotoxicity (ADCC). In some embodiments, the effector function of the functional Fc region is phagocytosis. In some embodiments, the effector function of the functional Fc region is ADCC and phagocytosis.

[0179] In some embodiments, the antibody or antigen-binding fragment can induce complement-dependent cytotoxicity (CDC).

[0180] In some embodiments, the Fc region is human IgG1, human IgG2, human IgG3, or human IgG4. In some embodiments, the antibody is a human IgG1 antibody.

[0181] In some embodiments, the antibody or antigen-binding fragment does not have a functional Fc region. For example, the antibody or antigen-binding fragment is a Fab, Fab', F(ab')2, or Fv fragment. In some embodiments, the Fc region has a LALA mutation (L234A and L235A mutations in EU numbering) or a LALA-PG mutation (EU numbering: L234A, L235A, P329G mutations).

[0182] In some embodiments, the Fc has SI mutations (EU numbering: S239D and I332E).

[0183] Method for preparing anti-IL2RA antibodies

[0184] Isolated fragments of human IL2RA (e.g., the extracellular region) can be used as immunogens to generate antibodies using standard polyclonal and monoclonal antibody production techniques. Polyclonal antibodies can be produced by multiple injections (e.g., subcutaneous or intraperitoneal) of the antigenic peptide or protein into an animal. In some embodiments, the antigenic peptide or protein is co-injected with at least one adjuvant. In some embodiments, the antigenic peptide or protein can be conjugated to an immunogenic agent derived from the species to be immunized. Animals can receive more than one (e.g., two, three, or four) injections of the antigenic peptide or protein.

[0185] The full-length polypeptide or protein, or an antigenic peptide fragment thereof, can be used as an immunogen. The antigenic peptide of the protein comprises at least 8 (e.g., at least 10, 15, 20, or 30) amino acid residues of the IL2RA amino acid sequence and encompasses an epitope of the protein, such that antibodies raised against the peptide can form a specific immune complex with the protein. As previously mentioned, the full-length sequence of human IL2RA is known in the art (SEQ ID NO: 64). In some embodiments, an Fc-tagged human IL2RA protein (the Fc fusion protein comprises all or part of the extracellular domain of human IL2RA, e.g., amino acids 22-213 of SEQ ID NO: 64) is used as an immunogen.

[0186] Antibodies are typically prepared by immunizing a suitable subject (e.g., a human or a transgenic animal expressing at least one human immunoglobulin locus). Suitable immunogenic preparations can include, for example, recombinantly expressed or chemically synthesized polypeptides (e.g., human IL2RA fragments). The preparations can further include an adjuvant, such as Freund's complete or incomplete adjuvant, or similar immunostimulants.

[0187] Polyclonal antibodies can be prepared by the methods described above: immunizing a suitable subject with an IL2RA polypeptide or antigenic peptide thereof (e.g., a portion of IL2RA, such as the extracellular domain) as an immunogen. Antibody titers in the immunized subject can be monitored over time using standard techniques, such as enzyme-linked immunosorbent assay (ELISA) using immobilized IL2RA polypeptide or peptide. If necessary, antibody molecules can be isolated from a mammal (e.g., blood) and purified to obtain an IgG fraction using known techniques (e.g., protein A or protein G chromatography). At an appropriate time after immunization (e.g., when the specific antibody titer is highest), antibody-producing cells can be obtained from the subject and monoclonal antibodies can be prepared using standard techniques, such as the hybridoma technology pioneered by Kohler et al. (Nature 256:495-497, 1975), human B cell hybridoma technology (Kozbor et al., Immunol. Today 4:72, 1983), EBV-hybridoma technology (Cole et al., Monoclonal Antibodies and Cancer Therapy, Alan R. Liss, Inc., pp.77-96, 1985), or trioma technology. Hybridoma production technology is well known in the art (see Current Protocols in Immunology, 1994, Coligan et al. (Eds.), John Wiley & Sons, Inc., New York, NY). Hybridoma cells producing monoclonal antibodies are identified by screening hybridoma culture supernatants for antibodies that bind to the polypeptide or epitope of interest (eg, using a standard ELISA assay).

[0188] Variants of the antibodies or antigen-binding fragments described herein can be prepared by introducing appropriate nucleotide changes into DNA encoding a humanized, chimeric, or human antibody, or by peptide synthesis. Such variants include, for example, deletions, insertions, or substitutions of residues in the amino acid sequence that constitutes the antigen-binding site of the antibody or antigen-binding domain. Within such a population of variants, some antibodies or antigen-binding fragments will have enhanced affinity for the target protein (e.g., IL2RA). Antibodies or antigen-binding fragments thereof with enhanced binding affinity for the target can be obtained by any combination of deletions, insertions, and / or substitutions. Amino acid changes introduced into the antibody or antigen-binding fragment can also alter or introduce new post-translational modifications, such as changing (e.g., increasing or decreasing) the number of glycosylation sites, changing the type of glycosylation site (e.g., changing the amino acid sequence so that cellular enzymes can bind different carbohydrates), or introducing new glycosylation sites.

[0189] The antibodies disclosed herein can be derived from any animal species, including mammals. Non-limiting examples of natural antibodies include antibodies derived from humans, primates (such as monkeys and apes), cattle, pigs, horses, sheep, camelids (such as camels and alpacas), chickens, goats, and rodents (such as rats, mice, hamsters, and rabbits), including transgenic rodents that have been genetically engineered to produce human antibodies.

[0190] Humanized and human antibodies include antibodies whose variable and constant regions are derived from human germline immunoglobulin sequences (or have the same amino acid sequence). Human antibodies may contain amino acid residues encoded by non-human germline immunoglobulin sequences (such as mutations introduced by random / site-directed mutagenesis in vitro or somatic mutation in vivo), for example, in the CDR regions.

[0191] Humanized antibodies typically have human framework regions (FRs) grafted with non-human CDRs. Thus, humanized antibodies contain one or more amino acid sequences derived from non-human sources, and these non-human amino acid residues are often referred to as "import" residues, typically taken from "import" variable domains. Humanization can be achieved by, for example, replacing the corresponding sequences of human antibodies with rodent CDRs or CDR sequences. These methods are described in: Jones et al. "Replacing the complementarity-determining regions in a human antibody with those from a mouse." Nature 321.6069 (1986): 522; Riechmann et al. "Reshaping human antibodies for therapy." Nature 332.6162 (1988): 323; Dall'Acqua et al. "Antibody humanization by framework shuffling." Methods 36.1 (2005): 43-60; Each document is incorporated herein by reference in its entirety. Therefore, "humanized" antibodies refer to chimeric antibodies in which the intact human V domains are substantially not replaced by corresponding sequences from non-human species. In practice, humanized antibodies are usually mouse antibodies in which some CDR residues and FR residues are replaced by residues at analogous sites in human antibodies.

[0192] The selection of human VH and VL domains for the preparation of humanized antibodies is crucial for reducing immunogenicity. According to the "best match" method, the mouse antibody V domain sequence is compared with a library of known human domain sequences, and the human sequence closest to the mouse sequence is selected as the FR of the humanized antibody (Sims et al. "A humanized CD18 antibody can block function without cell destruction." The Journal of Immunology 151.4 (1993): 2296-2308; Chothia, et al., "Canonical structures for the hypervariable regions of immunoglobulins." Journal of Molecular Biology 196.4 (1987): 901-917).

[0193] It is crucial to ensure that humanized antibodies are modified while maintaining high specificity, high affinity and other excellent biological properties for antigens. To achieve this goal, humanized antibodies can be prepared by analyzing the parental sequence and a variety of conceptual humanized products, and using three-dimensional models of the parental and humanized sequences. Three-dimensional immunoglobulin models are widely used and well known to those skilled in the art. Existing computer programs can simulate and display the possible three-dimensional conformational structures of selected candidate immunoglobulin sequences. By examining these display results, the potential role of the residues in the function of the candidate immunoglobulin can be analyzed, that is, the residues that affect the ability of the candidate immunoglobulin to bind to the antigen can be analyzed. In this way, FR residues can be selected and combined from the receptor sequence and the imported sequence to achieve the desired antibody characteristics (such as enhancing affinity for the target antigen).

[0194] Generally, amino acid sequence variants of humanized, chimeric or human anti-IL2RA antibodies will comprise an amino acid sequence that is at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98% or 99% identical to the original antibody light or heavy chain sequence.

[0195] In some embodiments, antibodies are generated using mice (e.g., RenMab mice) carrying humanized heavy chain immunoglobulin loci and humanized kappa chain immunoglobulin loci. The heavy chain immunoglobulin locus is a region on a chromosome comprising antibody heavy chain genes, and the locus may include, for example, human IGHV (variable region) genes, human IGHD (diversity) genes, human IGHJ (joining region) genes, and mouse heavy chain constant domain genes. The kappa chain immunoglobulin locus is a region on a chromosome comprising coding antibody light chain (kappa chain) genes, and the locus may include, for example, human IGKV (variable region) genes, human IGKJ (joining region) genes, and mouse light chain constant domain genes. For a detailed description of RenMab mice, see PCT / CN2020 / 075698 (incorporated herein by reference in its entirety). The antibody generated by the mouse has fully human VH, fully human VL, and mouse constant region. In some embodiments, human VH and human VL are connected to human IgG constant region (e.g., IgG1, IgG2, IgG3, and IgG4).

[0196] In some embodiments, mice carrying humanized heavy chain immunoglobulin loci and humanized kappa chain immunoglobulin loci are used (e.g., RenLite TMThe heavy chain immunoglobulin locus is a region on the chromosome that contains antibody heavy chain genes. This locus can include, for example, human IGHV (variable region) genes, human IGHD (diversity) genes, human IGHJ (joining region) genes, and mouse heavy chain constant domain genes. The kappa chain immunoglobulin locus is a region on the chromosome that contains genes encoding common light chains. This locus can include, for example, human IGKV (variable region) genes, human IGKJ (joining region) genes, and mouse light chain constant domain genes. About RenLite TM For a detailed description of mice, see PCT / CN2021 / 097652 (incorporated herein by reference in its entirety).

[0197] In some embodiments, to facilitate production efficiency by avoiding Fab-arm exchange, the Fc region of the antibodies was further engineered to replace the serine at position 228(EU numbering) of IgG4 with proline(S228P). Adetailed description regarding S228 mutation is described, e.g., in Silva etal. andphysiological matrix preparation."Journal of Biological Chemistry 290.9(2015):5462-5469, which is incorporated by reference in its entirety.

[0198] Identity or homology to a native sequence generally refers to the percentage of amino acid residues in a candidate sequence that are identical to the sequence of a humanized, chimeric, or human anti-IL2RA antibody or fragment thereof, after aligning the sequences and introducing gaps, if necessary, to achieve the maximum percent sequence identity, and conservative substitutions are not included in sequence identity calculations.

[0199] Additional modifications may be made to the anti-IL2RA antibodies or antigen-binding fragments. For example, cysteine ​​residues may be introduced into the Fc region to form interchain disulfide bonds in this region. The resulting homodimeric antibodies may have an increased half-life in vitro and / or in vivo. Homodimeric antibodies with increased half-life in vitro and / or in vivo may also be prepared using heterobifunctional cross-linkers, such as those described by Wolff et al. ("Monoclonal antibody homodimers: enhanced antitumor activity in nude mice." Cancer Research 53.11 (1993): 2560-2565). Alternatively, antibodies with dual Fc regions may be designed.

[0200] In some embodiments, the anti-IL2RA antibody or antigen-binding fragment thereof may be covalently modified. Such covalent modifications can be achieved by chemical or enzymatic synthesis, or enzymatic / chemical cleavage. Other types of covalent modifications are introduced into the molecule by reacting targeted amino acid residues of the antibody or fragment thereof with an organic derivatizing agent that selectively reacts with side chains or N-terminal / C-terminal residues.

[0201] In some embodiments, antibody variants are provided that have sugar chain structures that do not contain fucose (directly or indirectly) attached to the Fc region. For example, the fucose content in the antibody composition may be 1% to 80%, 1% to 65%, 5% to 65%, or 20% to 40%. Fucose content is determined by MALDI-TOF mass spectrometry, calculating the average fucose content within the sugar chain at Asn297 as a percentage of the total amount of all sugar structures (such as complex, hybrid, and high-mannose) attached to Asn297, as described, for example, in WO 2008 / 077546. Asn297 refers to the asparagine residue located at approximately position 297 in the Fc region (EU numbering of Fc region residues; position 314 in Kabat numbering); however, due to minor sequence differences in antibodies, Asn297 may also be located within a range of approximately ±3 amino acids upstream or downstream of position 297 (i.e., positions 294 to 300). Such defucosylated variants may have enhanced ADCC function. In some embodiments, to reduce sugar chain heterogeneity, the Fc region of the antibody can be further engineered by replacing asparagine at position 297 with alanine (N297A).

[0202] In some embodiments, to avoid Fab arm exchange to improve production efficiency, the Fc region of the antibody is further transformed, and the 228th serine (EU numbering) of IgG4 is replaced with proline (S228P). For a detailed description of the S228 mutation, see, for example, Silva et al. ("The S228P mutation preventsin vivo and in vitro IgG4 Fab-arm exchange as demonstrated using a combination of novel quantitative immunoassays and physiological matrix preparation." Journal of Biological Chemistry 290.9(2015):5462-5469), which is incorporated herein by reference in its entirety.

[0203] recombinant vector

[0204] The present disclosure also provides a recombinant vector (e.g., an expression vector) comprising an isolated polynucleotide described herein (e.g., a polynucleotide encoding a polypeptide described herein), a host cell into which the recombinant vector is introduced (i.e., the host cell contains the polynucleotide and / or a vector comprising the polynucleotide), and a method for producing a recombinant antibody polypeptide or fragment thereof by recombinant technology.

[0205] As used herein, "vector" refers to any construct capable of delivering one or more target polynucleotides to a host cell when introduced into the host cell. "Expression vector" refers to a vector capable of delivering and expressing one or more target polynucleotides as a coded polypeptide in the introduced host cell. Therefore, in the expression vector, the target polynucleotide is positioned in the vector for expression by being operably connected to a regulatory element (such as a promoter, an enhancer and / or a poly A tail), or is located at, near or on both sides of the target polynucleotide integration site in the host cell genome so that the target polynucleotide can be translated in the host cell into which the expression vector is introduced.

[0206] The vector can be introduced into the host cell by methods known in the art, such as electroporation, chemical transfection (such as DEAE-dextran), transformation, transfection, infection and / or transduction (such as using recombinant viruses). Therefore, non-limiting examples of vectors include viral vectors (which can be used to produce recombinant viruses), naked DNA or RNA, plasmids, cosmids, phage vectors, and DNA or RNA expression vectors combined with cationic condensing agents.

[0207] In some embodiments, a polynucleotide described herein (e.g., a polynucleotide encoding a polypeptide described herein) is introduced using a viral expression system (e.g., vaccinia virus or other poxvirus, retrovirus, or adenovirus), which may involve the use of a non-pathogenic (defective) replication-competent virus or a replication-defective virus, the latter of which typically undergoes viral propagation only in complementing virus packaging cells. Suitable systems are described, for example, in Fisher-Hoch et al., 1989, Proc. Natl. Acad. Sci. USA 86:317-321; Flexner et al., 1989, Ann. NY Acad Sci. 569:86-103; Flexner et al., 1990, Vaccine, 8:17-21; U.S. Pat. Nos. 4,603,112, 4,769,330, and 5,017,487; WO 89 / 01973; U.S. Pat. No. 4,777,127; GB 2,200,651; EP 0,345,242; WO 91 / 02805; Berkner-Biotechniques, 6:616-627, 1988; Rosenfeld et al. et al., 1991, Science, 252:431-434; Kolls et al., 1994, Proc. Natl. Acad. Sci. USA, 91:215-219; Kass-Eisler et al., 1993, Proc. Natl. Acad. Sci. USA, 90:11498-11502; Guzman et al., 1993, Circulation, 88:2838-2848; Guzman et al., 1993, Cir. Res., 73:1202-1207. Techniques for integrating DNA into such expression systems are well known to those of ordinary skill in the art. DNA can also be in "naked" form, as described, for example, by Ulmer et al., 1993, Science, 259:1745-1749 and Cohen, 1993, Science, 259:1691-1692. The efficiency of naked DNA uptake can be increased by coating the DNA on degradable microbeads that are efficiently transported into cells.

[0208] For expression purposes, the DNA insert comprising the polynucleotides described herein encoding the antibody or polypeptide can be operably linked to an appropriate promoter (e.g., a heterologous promoter), such as the bacteriophage lambda PL promoter, the E. coli lac, trp, and tac promoters, the SV40 early and late promoters, the retroviral LTR promoters, and the like. Other suitable promoters are known to those skilled in the art. In some embodiments, the promoter is a cytomegalovirus (CMV) promoter. The expression construct may also comprise a transcription initiation site, a termination site, and a ribosome binding site for translation in the transcribed region. The coding region of the mature transcript expressed by the construct may comprise a start codon for initiating translation and a stop codon (UAA, UGA, or UAG) at the end of the polypeptide to be translated.

[0209] As previously mentioned, the expression vector may contain at least one selection marker. Such markers include dihydrofolate reductase or neomycin resistance genes for eukaryotic cell culture, and tetracycline or ampicillin resistance genes for bacterial culture such as Escherichia coli. Representative examples of suitable host cells include, but are not limited to, bacterial cells (such as Escherichia coli, Streptomyces, and Salmonella typhimurium cells); fungal cells (such as yeast cells); insect cells (such as Drosophila S2 cells and Spodoptera frugiperda Sf9 cells); animal cells (such as CHO, COS, Bowes melanoma, and HK 293 cells); and plant cells. Suitable culture media and culture conditions for the host cells described herein are known in the art.

[0210] Non-limiting bacterial vectors include pQE70, pQE60, and pQE-9 (available from Qiagen); pBS vectors, Phagescript vectors, Bluescript vectors, pNH8A, pNH16a, pNH18A, pNH46A (available from Stratagene); and ptrc99a, pKK223-3, pKK233-3, pDR540, and pRIT5 (available from Pharmacia). Non-limiting eukaryotic vectors include pWLNEO, pSV2CAT, pOG44, pXT1, and pSG (available from Stratagene); and pSVK3, pBPV, pMSG, and pSVL (available from Pharmacia. Other suitable vectors are readily known to those skilled in the art.

[0211] Non-limiting promoters suitable for use in bacteria include the E. coli lacI and lacZ promoters, the T3 and T7 promoters, the gpt promoter, the λPR and PL promoters, and the trp promoter. Suitable eukaryotic promoters include the CMV immediate early promoter, the HSV thymidine kinase promoter, the SV40 early and late promoters, retroviral LTR promoters (such as the Rous sarcoma virus RSV promoter), and metallothionein promoters (such as the mouse metallothionein-I promoter).

[0212] In Saccharomyces cerevisiae, a variety of vectors containing constitutive or inducible promoters (such as the alpha factor, alcohol oxidase, and PGH promoters) are available.

[0213] The construct can be introduced into the host cell by calcium phosphate transfection, DEAE-dextran mediated transfection, cationic liposome mediated transfection, electroporation, transduction, infection or other methods. Such methods are described in detail in various standard laboratory manuals, such as Davis et al., Basic Methods In Molecular Biology (1986), which is incorporated herein by reference in its entirety.

[0214] Transcription of the disclosed antibody-encoding DNA by higher eukaryotes can be increased by inserting an enhancer sequence into the vector. Enhancers are cis-acting DNA elements, typically about 10-300 bp in length, that increase the transcriptional activity of a promoter in a specific host cell type. Examples of enhancers include the SV40 enhancer located 100-270 bp from the late side of the SV40 replication origin, the cytomegalovirus early promoter enhancer, the polyoma virus late enhancer on the replication origin, and adenovirus enhancers.

[0215] In order to secrete the translated protein into the endoplasmic reticulum, periplasmic space or extracellular environment, an appropriate secretion signal can be incorporated into the expressed polypeptide. The signal can be derived from the endogenous sequence of the polypeptide or a heterologous signal.

[0216] Polypeptides (e.g., antibodies) can be expressed in modified forms, such as fusion proteins (e.g., GST fusion proteins) or histidine-tagged forms, and can include not only secretion signals but also additional heterologous functional regions. For example, an additional amino acid region (particularly charged amino acids) can be added to the N-terminus of the polypeptide to enhance stability within the host cell, the purification process, or durability during subsequent processing and storage. In addition, peptide segments can be added to facilitate purification, and such regions can be removed before the final preparation of the polypeptide. Techniques for achieving secretion / excretion, enhancing stability, and facilitating purification by adding peptide segments are conventional methods well known in the art.

[0217] Treatment

[0218] The antibodies or antigen-binding fragments thereof disclosed herein can be used for a variety of therapeutic purposes.

[0219] In one aspect, the present disclosure provides methods for treating cancer in a subject, methods for reducing the rate of tumor growth over time in a subject, methods for reducing the risk of developing metastasis, or methods for reducing the risk of developing additional metastases in a subject. In some embodiments, the treatment can halt, slow, delay, or inhibit cancer progression. In some embodiments, the treatment can reduce the number, severity, and / or duration of cancer-related symptoms in a subject.

[0220] In one aspect, the disclosure features a method comprising administering a therapeutically effective amount of an antibody or antigen-binding fragment thereof to a subject in need thereof (e.g., a subject suffering from or identified / diagnosed as having cancer), such as breast cancer (e.g., triple-negative breast cancer), carcinoid tumors, cervical cancer, endometrial cancer, nervous system cancer, head and neck cancer, liver cancer, lung cancer, small cell lung cancer, lymphoma, melanoma, ovarian cancer, pancreatic cancer, prostate cancer, kidney cancer, colorectal cancer, gastric cancer, testicular cancer, thyroid cancer, bladder cancer, urethral cancer, skin cancer, or hematologic malignancies. In some embodiments, the cancer is unresectable melanoma or metastatic melanoma, non-small cell lung cancer (NSCLC), small cell lung cancer (SCLC), bladder cancer, gastroesophageal junction cancer, or metastatic hormone-refractory prostate cancer. In some embodiments, the subject suffers from a solid tumor. In some embodiments, the cancer is squamous cell carcinoma of the head and neck (SCCHN), renal cell carcinoma (RCC), triple-negative breast cancer (TNBC), or colorectal cancer. In some embodiments, the subject has Hodgkin lymphoma. In some embodiments, the subject has triple-negative breast cancer (TNBC), gastric cancer, urothelial carcinoma, Merkel cell carcinoma, or head and neck cancer. In some embodiments, the cancer is melanoma, pancreatic cancer, mesothelioma, glioma, hematologic malignancy (particularly non-Hodgkin lymphoma), lymphoma, chronic lymphocytic leukemia, or advanced solid tumors.

[0221] In some embodiments, the anti-IL2RA antibody is designed to treat melanoma (e.g., advanced melanoma), non-small cell lung cancer (NSCLC), small cell lung cancer (SCLC), B-cell non-Hodgkin lymphoma, bladder cancer, and / or prostate cancer (e.g., metastatic hormone-refractory prostate cancer). In some embodiments, the anti-IL2RA antibody is designed to treat hepatocellular carcinoma, ovarian cancer, colon cancer, or cervical cancer. In some embodiments, the anti-IL2RA antibody is designed to treat advanced breast cancer, advanced ovarian cancer, and / or advanced refractory solid tumors. In some embodiments, the anti-IL2RA antibody is designed to treat metastatic solid tumors, NSCLC, melanoma, non-Hodgkin lymphoma, colorectal cancer, and multiple myeloma. In some embodiments, the anti-IL2RA antibody is designed to treat melanoma, pancreatic cancer, mesothelioma, hematological malignancies (e.g., non-Hodgkin lymphoma, lymphoma, chronic lymphocytic leukemia), or solid tumors (e.g., advanced solid tumors). In some embodiments, the anti-IL2RA antibody is designed to treat cancer (e.g., nasopharyngeal, bladder, cervical, renal, or ovarian cancer).

[0222] In some embodiments, the IL2RA antibodies are designed to treat acute lymphoblastic leukemia (ALL), B-cell chronic lymphocytic leukemia (B-CLL), hairy cell leukemia (HCL), solid tumors, colorectal cancer, ovarian cancer, prostate cancer, melanoma, lung cancer, breast cancer, gastric cancer, esophageal squamous cell carcinoma (ESCC), leukemia, lymphoma, multiple myeloma, sarcoma, and / or head and neck cancer.

[0223] In some embodiments, the compositions and methods disclosed herein can be used to treat patients at high risk for cancer.Cancer patients can be identified by various methods known in the art.

[0224] In some aspects, the disclosure relates to a method of treating an autoimmune disease or inflammation, comprising administering to a subject an effective amount of a composition comprising an antibody or antigen-binding fragment thereof described herein, or an antibody-drug conjugate described herein.

[0225] In one aspect, the present disclosure provides methods for treating, preventing, or reducing the risk of diseases associated with abnormal or undesirable immune responses, such as autoimmune diseases. Such autoimmune diseases include, but are not limited to, alopecia areata, lupus, ankylosing spondylitis, Meniere's disease, antiphospholipid syndrome, mixed connective tissue disease, autoimmune Addison's disease, multiple sclerosis, autoimmune hemolytic anemia, myasthenia gravis, autoimmune hepatitis, pemphigus vulgaris, Behçet's disease, pernicious anemia, bullous pemphigoid, polyarteritis nodosa, cardiomyopathy, polychondritis, celiac disease-dermatitis, polyglandular syndrome, chronic fatigue syndrome (CFIDS), polymyalgia rheumatica, chronic inflammatory demyelinating polyneuropathy, polymyositis and dermatomyositis, essential agammaglobulinemia, eosinophilic granulomatosis with polyangiitis (Churg -Strauss syndrome), primary biliary cholangitis, cicatricial pemphigoid, psoriasis, CREST syndrome, Raynaud's phenomenon, cold agglutinin disease, Reiter's syndrome, Crohn's disease, rheumatic fever, discoid lupus erythematosus, rheumatoid arthritis, cryoglobulinemia, sarcoidosis, fibromyalgia, scleroderma, Graves' disease, Sjögren's syndrome, Guillain-Barré syndrome, stiff-person syndrome, Hashimoto's thyroiditis, Takayasu's arteritis, idiopathic pulmonary fibrosis, temporal arteritis / giant cell arteritis, idiopathic thrombocytopenic purpura (ITP), ulcerative colitis, IgA nephropathy, uveitis, diabetes (e.g., type 1), vasculitis, lichen planus, and vitiligo. The anti-IL2RA antibodies or antigen-binding fragments thereof can also be administered to a subject to treat, prevent, or reduce the risk of abnormal or undesirable immune responses associated with cell, tissue, or organ transplantation (e.g., kidney, liver, and heart transplantation), such as graft-versus-host disease (GVHD), or to prevent allogeneic transplant rejection. In some embodiments, the subject has Crohn's disease, ulcerative colitis, or type 1 diabetes. In some embodiments, the antibody or antigen-binding fragment can be used to treat inflammation. In some embodiments, the anti-IL2RA antibody is designed to treat other diseases or conditions, such as microbial infections and allergic diseases.

[0226] In some embodiments, the antibodies or antigen-binding fragments described herein can specifically target regulatory T cells (Tregs) without blocking the interaction between IL2 and IL2RA. Therefore, they can be used to eliminate Treg cells and thereby relieve immunosuppression. At the same time, preserving the IL2 / IL2RA interaction can effectively stimulate T cells by activating the IL2 / IL2RA signaling pathway.

[0227] In some aspects, the present disclosure relates to a method of suppressing an immune response in a subject, comprising administering to the subject an effective amount of a composition comprising an antibody or antigen-binding fragment thereof described herein or an antibody-drug conjugate described herein. In some embodiments, the subject suffers from an autoimmune disease.

[0228] As used herein, an "effective amount" refers to an amount or dosage sufficient to produce a beneficial or desired result, including stopping, slowing, hindering, or inhibiting the progression of a disease (e.g., cancer). The effective amount will vary depending on, for example, the age and weight of the subject to whom the antibody, antigen-binding fragment, polynucleotide encoding the antibody, vector comprising the polynucleotide, and / or composition thereof is to be administered, the severity of the symptoms, and the route of administration, and thus the mode of administration can be determined on an individual basis.

[0229] An effective amount can be administered once or multiple times. For example, an effective amount of an antibody or antigen binding fragment is an amount sufficient to improve, stop, stabilize, reverse, inhibit, slow down and / or delay the progression of a patient's autoimmune disease or cancer, or an amount sufficient to improve, stop, stabilize, reverse, slow down and / or delay the proliferation of cells (e.g., biopsy cells, any cancer cell or cell line described herein (e.g., cancer cell line)) in vitro. As understood in the art, the effective amount of an antibody or antigen binding fragment may vary, depending on factors such as the patient's medical history and the type (and / or dosage) of the antibody used, among other factors.

[0230] The effective amount and regimen of administering the antibodies, polynucleotides encoding the antibodies, and / or compositions disclosed herein can be determined empirically, and such determinations are within the skill of those skilled in the art. It will be understood by those skilled in the art that the dosage that must be administered will vary depending on factors such as the mammal to which the antibodies, polynucleotides encoding the antibodies, and / or compositions disclosed herein will be administered, the route of administration, the specific type of antibody, polynucleotide encoding the antibodies, antigen-binding fragments, and / or compositions disclosed herein used, and other drugs administered to the mammal.

[0231] A typical daily dose of an effective amount of the antibody is 0.01 mg / kg to 100 mg / kg (milligrams per kilogram of patient body weight). In some embodiments, the dosage can be less than 100 mg / kg, 50 mg / kg, 40 mg / kg, 30 mg / kg, 25 mg / kg, 20 mg / kg, 10 mg / kg, 9 mg / kg, 8 mg / kg, 7 mg / kg, 6 mg / kg, 5 mg / kg, 4 mg / kg, 3 mg / kg, 2 mg / kg, 1 mg / kg, 0.5 mg / kg, 0.3 mg / kg, or 0.1 mg / kg. In some embodiments, the dose may be greater than 50 mg / kg, 40 mg / kg, 30 mg / kg, 20 mg / kg, 10 mg / kg, 9 mg / kg, 8 mg / kg, 7 mg / kg, 6 mg / kg, 5 mg / kg, 4 mg / kg, 3 mg / kg, 2 mg / kg, 1 mg / kg, 0.5 mg / kg, 0.3 mg / kg, 0.1 mg / kg, 0.05 mg / kg, or 0.01 mg / kg. In some embodiments, the dose is about 50 mg / kg, 40 mg / kg, 30 mg / kg, 25 mg / kg, 20 mg / kg, 10 mg / kg, 9 mg / kg, 8 mg / kg, 7 mg / kg, 6 mg / kg, 5 mg / kg, 4 mg / kg, 3 mg / kg, 2 mg / kg, 1 mg / kg, 0.9 mg / kg, 0.8 mg / kg, 0.7 mg / kg, 0.6 mg / kg, 0.5 mg / kg, 0.4 mg / kg, 0.3 mg / kg, 0.2 mg / kg, or 0.1 mg / kg.

[0232] In any of the methods described herein, the at least one antibody, antigen-binding fragment thereof, or pharmaceutical composition (e.g., any antibody, antigen-binding fragment, or pharmaceutical composition described herein) and optionally at least one additional therapeutic agent can be administered to the subject at least once a week (e.g., once a week, twice a week, three times a week, four times a week, once a day, twice a day, or three times a day). In some embodiments, at least two different antibodies and / or antigen-binding fragments are administered in the same composition (e.g., a liquid composition). In some embodiments, at least one antibody or antigen-binding fragment and at least one additional therapeutic agent are administered in the same composition (e.g., a liquid composition). In some embodiments, the at least one antibody or antigen-binding fragment and the at least one additional therapeutic agent are administered in two different compositions (e.g., a liquid composition comprising at least one antibody or antigen-binding fragment and a solid oral composition comprising at least one additional therapeutic agent). In some embodiments, the at least one additional therapeutic agent is administered as a pill, tablet, or capsule. In some embodiments, the at least one additional therapeutic agent is administered in a sustained-release oral formulation.

[0233] In some embodiments, the one or more additional therapeutic agents may be administered to the subject before or after administration of the at least one antibody, antigen-binding antibody fragment, or pharmaceutical composition (e.g., any described herein). In some embodiments, the one or more additional therapeutic agents and the at least one antibody, antigen-binding antibody fragment, or pharmaceutical composition (e.g., any described herein) are administered to the subject such that the one or more additional therapeutic agents and the at least one antibody or antigen-binding fragment (e.g., any described herein) have an overlapping period of biological activity in the subject.

[0234] In some embodiments, the subject may be administered the at least one antibody, antigen-binding antibody fragment, or pharmaceutical composition (e.g., any of the antibodies, antigen-binding antibody fragments, or pharmaceutical compositions described herein) for an extended period of time (e.g., at least 1 week, 2 weeks, 3 weeks, 1 month, 2 months, 3 months, 4 months, 5 months, 6 months, 7 months, 8 months, 9 months, 10 months, 11 months, 12 months, 1 year, 2 years, 3 years, 4 years, or 5 years). A skilled medical professional can determine the length of the treatment period using any of the methods described herein for diagnosing or tracking the effectiveness of treatment (e.g., observing at least one symptom of cancer). As described herein, a skilled medical professional can also change the type and amount (e.g., increase or decrease) of the antibody or antigen-binding antibody fragment (and / or one or more additional therapeutic agents) administered to the subject, and can adjust (e.g., increase or decrease) the dose or frequency of administration of the at least one antibody or antigen-binding antibody fragment (and / or one or more additional therapeutic agents) administered to the subject based on an assessment of the effectiveness of the treatment (e.g., using any of the methods described herein and known in the art).

[0235] In some embodiments, one or more additional therapeutic agents may be administered to the subject. The additional therapeutic agent may include one or more inhibitors selected from the group consisting of: B-Raf inhibitors, EGFR inhibitors, MEK inhibitors, ERK inhibitors, K-Ras inhibitors, c-Met inhibitors, anaplastic lymphoma kinase (ALK) inhibitors, phosphatidylinositol 3-kinase (PI3K) inhibitors, Akt inhibitors, mTOR inhibitors, dual PI3K / mTOR inhibitors, Bruton's tyrosine kinase (BTK) inhibitors, and isocitrate dehydrogenase 1 (IDH1) and / or isocitrate dehydrogenase 2 (IDH2) inhibitors. In some embodiments, the additional therapeutic agent is an indoleamine 2,3-dioxygenase-1 (IDO1) inhibitor (e.g., ipacastat).

[0236] In some embodiments, the additional therapeutic agent may comprise one or more inhibitors selected from the group consisting of: a HER3 inhibitor, a LSD1 inhibitor, an MDM2 inhibitor, a BCL2 inhibitor, a CHK1 inhibitor, an activated hedgehog signaling pathway inhibitor, and an agent that selectively degrades the estrogen receptor.

[0237] In some embodiments, the additional therapeutic agent may comprise one or more therapeutic agents selected from the group consisting of trabectedin, nab-paclitaxel, trabentanib, pazopanib, cediranib, palbociclib, everolimus, fluoropyrimidine, IFL, regorafenib, Reolysin, Alimta, Zykadia, Sutent, temsirolimus, axitinib, everolimus, sorafenib, Votrient, pazopanib, IMA-901, AGS-003, cabozantinib, vinflunine, Hsp90 inhibitors, Ad-GM-CSF, temozolomide, IL-2, IFNa, vinblastine, thalidomide, dacarbazine, cyclophosphamide, lenalidomide, azacitidine, lenalidomide, bortezomib, amrubicin, carfilzomib, pralatrexate, and enzastaurin.

[0238] In some embodiments, the additional therapeutic agent may comprise one or more therapeutic agents selected from the group consisting of an adjuvant, a TLR agonist, IL-1, HMGB1, an IL-10 antagonist, an IL-4 antagonist, an IL-6 antagonist (e.g., an IL-6 receptor), an IL-13 antagonist, an IL-17 antagonist, an HVEM antagonist, an ICOS agonist, a CX3CL1-targeted therapy, a CXCL9-targeted therapy, a CXCL10-targeted therapy, a CCL5-targeted therapy, an LFA-1 agonist, an ICAM1 agonist, and a Selectin agonist.

[0239] In some embodiments, the subject is administered carboplatin, nab-paclitaxel, paclitaxel, cisplatin, pemetrexed, gemcitabine, FOLFOX, or FOLFIRI.

[0240] In some embodiments, the additional therapeutic agent is an anti-OX40 antibody, an anti-PD-1 antibody, an anti-PD-L1 antibody, an anti-PD-L2 antibody, an anti-LAG-3 antibody, an anti-TIGIT antibody, an anti-BTLA antibody, an anti-CTLA-4 antibody, an anti-GITR antibody, an anti-TIM-3 antibody, an anti-4-1BB antibody, or an anti-CD40 antibody.

[0241] Pharmaceutical compositions and routes of administration

[0242] Also provided herein are pharmaceutical compositions comprising at least one (e.g., one, two, three, or four) antibodies or antigen-binding fragments described herein. Any two or more (e.g., two, three, or four) antibodies or antigen-binding fragments described herein may be present in any combination in a pharmaceutical composition. The pharmaceutical composition may be formulated in any manner known in the art.

[0243] Pharmaceutical composition is formulated to be compatible with its expected route of administration (e.g., intravenous, intra-arterial, intramuscular, intradermal, subcutaneous or intraperitoneal). The composition can include a sterile diluent (e.g., sterile water or saline), fixed oil, polyethylene glycol, glycerol, propylene glycol or other synthetic solvents, antibacterial or antifungal agents, such as benzyl alcohol or methyl paraben, chlorobutanol, phenol, ascorbic acid, thimerosal, etc., antioxidants, such as ascorbic acid or sodium bisulfite, chelating agents, such as ethylenediaminetetraacetic acid, buffers, such as acetates, citrates or phosphates, and isotonic agents, such as sugars (e.g., glucose), polyols (e.g., mannitol or sorbitol) or salts (e.g., sodium chloride), or any combination thereof. Liposomal suspensions can also be used as pharmaceutically acceptable carriers. The preparation of the composition can be prepared and packaged in ampoules, disposable syringes or multiple-dose bottles. Where necessary (e.g., in injectable formulations), suitable fluidity can be maintained, for example, by the use of a coating such as lecithin or a surfactant. Absorption of the antibody or antigen-binding fragment thereof can be prolonged by including agents that delay absorption (e.g., aluminum monostearate and gelatin). Alternatively, controlled release can be achieved by implants and microencapsulated delivery systems, which may include biodegradable, biocompatible polymers (e.g., ethylene vinyl acetate, polyanhydrides, polyglycolic acid, collagen, polyorthoesters, and polylactic acid).

[0244] Compositions containing any one or more of the antibodies or antigen-binding fragments described herein can be formulated in dosage unit form (i.e., physically discrete units containing a predetermined quantity of active compound to allow for ease of administration and uniformity of dosage) for parenteral (e.g., intravenous, intraarterial, intramuscular, intradermal, subcutaneous, or intraperitoneal) administration.

[0245] Pharmaceutical compositions for parenteral administration are preferably sterile, substantially isotonic, and produced under good manufacturing practice (GMP) conditions. Pharmaceutical compositions can be provided in unit dosage form (i.e., a single dose). Pharmaceutical compositions can be formulated using one or more physiologically acceptable carriers, diluents, excipients, or adjuvants. The formulation depends on the chosen route of administration. For injection, the antibody can be formulated in an aqueous solution, preferably in a physiologically compatible buffer, to reduce discomfort at the injection site. The solution may contain a formulation agent, such as a suspending agent, a stabilizer, and / or a dispersant. Alternatively, the antibody can be provided in a lyophilized form for redissolution with a suitable carrier (e.g., sterile, pyrogen-free water) before use.

[0246] The toxicity and therapeutic efficacy of a composition can be determined by standard pharmaceutical procedures in cell cultures or experimental animals (e.g., monkeys). For example, the LD50 (the dose lethal to 50% of the population) and the ED50 (the dose therapeutically effective to 50% of the population) can be determined: the therapeutic index is the ratio of LD50:ED50. Agents exhibiting high therapeutic indices are preferred. When an agent exhibits adverse side effects, care should be taken to minimize potential harm (i.e., reduce adverse side effects). Toxicity and therapeutic efficacy can be determined by other standard pharmaceutical procedures.

[0247] The data obtained from cell culture assays and animal studies can be used to formulate the appropriate dose of any given agent for a subject (e.g., a human). The therapeutically effective amount of one or more (e.g., one, two, three, or four) antibodies or their antigen-binding fragments (e.g., any antibody or antibody fragment described herein) will be the amount of the disease (e.g., killing cancer cells) in a subject (e.g., a human subject determined to have cancer), or the amount of the disease in a subject determined to have a risk of illness (e.g., previously had cancer but now cured), reducing the severity, frequency, and / or duration of one or more symptoms of the disease in a subject (e.g., a human). The effectiveness and dosage of any antibody or antigen-binding fragment described herein can be determined by a healthcare professional or veterinary professional using methods known in the art and by observing one or more disease symptoms in a subject (e.g., a human). Certain factors may affect the dosage and timing required for the effective treatment of a subject (e.g., the severity of the disease or illness, previous treatments, the overall health and / or age of the subject, and the presence of other diseases).

[0248] Exemplary dosages include milligram or microgram amounts of any antibody or antigen-binding fragment described herein per kilogram of subject body weight (e.g., about 1 μg / kg to about 500 mg / kg; about 100 μg / kg to about 500 mg / kg; about 100 μg / kg to about 50 mg / kg; about 10 μg / kg to about 5 mg / kg; about 10 μg / kg to about 5 mg / kg; about 10 μg / kg to about 0.5 mg / kg; about 1 μg / kg to about 50 μg / kg; about 0.3 mg / kg to about 25 mg / kg; about 1 mg / kg to about 10 mg / kg; or about 1 mg / kg to about 5 mg / kg). Although these dosages cover a wide range, those of ordinary skill in the art will understand that the efficacy of therapeutic agents (including antibodies and antigen-binding fragments thereof) varies, and effective amounts can be determined by methods known in the art. Typically, a relatively low dose is administered first, and then the attending healthcare professional or veterinary professional (in the case of therapeutic applications) or researcher (when still in the development stage) can gradually increase the dose until an appropriate response is obtained. Furthermore, it will be understood that the specific dosage level for any particular subject will depend on a variety of factors, including the activity of the specific compound used, the age, weight, general health, sex and diet of the subject, the time of administration, the route of administration, the rate of excretion, and the half-life of the antibody or antibody fragment in vivo.

[0249] The pharmaceutical composition can be contained in a container, pack, or dispenser together with instructions for administration. The present disclosure also provides methods for producing the antibodies or antigen-binding fragments thereof having various uses described herein.

[0250] Example

[0251] The present invention is further described in the following examples, which do not limit the scope of the invention described in the claims.

[0252] Example 1. Generation of anti-IL2RA antibodies

[0253] To generate antibodies against human IL2RA, RenMice (i.e., engineered mice containing DNA encoding the variable regions of human immunoglobulin heavy and kappa light chains, such as RenMab) were immunized with Fc-tagged human IL2RA protein (ACROBiosystems Inc., Cat#: ILA-H5251) or plasmid encoding IL2RA protein. TM Mouse, RenLite TM Mice). Antibody immune responses were monitored by antigen-specific immunoassays.

[0254] A total of three immunizations were performed, with two weeks between immunizations. One week after the final immunization, retroorbital blood was collected, and serum antibody titers were determined by fluorescence-activated cell sorting (FACS). Two weeks later, mice with high titers were selected for pulse immunization. Pulse immunizations were performed using either intraperitoneal or tail vein injections of human IL2RA protein or CHO-S cells expressing human IL2RA protein.

[0255] When the desired immune response is achieved, antigen-specific immune cells are isolated from the immunized mice to further obtain anti-IL2RA antibodies or to obtain the light chain and heavy chain variable region sequences of anti-IL2RA antibodies. For example, using single cell technology (e.g., using Plasma cells secreting antigen-specific monoclonal antibodies were screened and identified using an optical fluidics system (Berkeley Lights Inc.). Antibody variable region sequences were obtained using reverse transcription and PCR sequencing. The obtained variable region sequences were cloned into a vector encoding a human IgG constant region sequence for antibody expression. Binding affinity of the expressed antibodies to IL2RA was verified using FACS.

[0256] Exemplary antibodies obtained include: 5D9, 7B5, 11C3, 11C12, 11D2, 13F9 and 14C2. The heavy and light chain variable regions of 5D9, 7B5, 11C3, 11C12, 11D2, 13F9 and 14C2 are shown in FIG. Figure 9 shown. Figure 7 The CDR sequences according to the Kabat definition are shown. Figure 8 The CDR sequences according to the Chothia definition are shown.

[0257] Various IgG1, IgG2, and IgG4 antibodies were prepared. Regarding antibody names, when the antibody VH / VL is linked to a different isotype, the isotype is added to the name. For example, if the VH and VL of 5D9 are linked to the IgG1 constant region, the antibody is named 5D9-IgG1 (or 5D9). Examples of other isotypes are as follows: 5D9-IgG2 and 5D9-IgG4. The constant region of an antibody may contain some mutations. For example, when the SI mutation (EU numbering: S239D and I332E mutations) is introduced into the Fc region of 5D9-IgG1, the resulting antibody is named 5D9-IgG1-SI (or 5D9-SI).

[0258] Example 2. Binding affinity of anti-IL2RA antibodies

[0259] Using Biacore with pre-immobilized Protein A sensor chips TMThe affinity of anti-IL2RA antibodies to His-tagged human IL2RA protein (hIL2RA-his, ACROBiosystems Inc., Cat#: ILA-H52H9) and His-tagged recombinant monkey (cynomolgus macaque, or Macaca fascicularis) IL2RA protein (cynoIL2RA-his, Sino Biological, Inc., Cat#: 90265-C08H) was measured using a Biacore 8K biosensor (Biacore, INC, Piscataway, N.J.) by surface plasmon resonance (SPR).

[0260] Purified anti-IL2RA antibody was diluted to 2 μg / mL and then injected into the Biacore at a flow rate of 10 μL / min. TM The 8K biosensor was titrated for approximately 50 seconds to reach the desired protein density (e.g., approximately 150 response units (RU)). His-tagged IL2RA protein at concentrations of 200, 100, 50, 25, 12.5, 6.25, 3.125, or 0 nM was then injected at a flow rate of 30 μL / min for 180 seconds. Dissociation was monitored for 400 seconds. After the last injection of each titration, the chip was regenerated with glycine (pH 2.0, 30 μL / min for 30 seconds).

[0261] Using Biacore TM 8K Evaluation Software 3.0 was used to globally fit the data to a 1:1 Langmuir binding model (Karlsson, R. Roos, H. Fagerstam, L. Petersson, B., 1994. Methods Enzymology 6.99-110), simultaneously obtaining kinetic association rates (k on ) and dissociation rates (k off ). Affinity was derived from the quotient of the kinetic rate constants (K = k off / k on ).

[0262] As will be understood by one of ordinary skill in the art, the same method was used for each test antibody with appropriate adjustment of parameters (e.g., antibody concentration). The results for the tested antibodies are summarized in the table below. ISO is an antibody of the same IgG1 subtype but targeting an unrelated target.

[0263] Table 1. Affinity test results

[0264]

[0265] (“—” indicates no combination)

[0266] RG6292 is a human IgG1 monoclonal antibody targeting IL2RA, which is in early clinical development by Roche for the treatment of patients with advanced and / or metastatic solid tumors. Its VH and VL sequences are shown in SEQ ID NO:62 and SEQ ID NO:63, respectively.

[0267] The results showed that the anti-IL2RA antibodies 5D9, 7B5, 11C3, 11C12, 11D2, 13F9, and 14C2 had good binding affinity to human IL2RA and monkey IL2RA.

[0268] Example 3. Binding activity of anti-IL2RA antibodies

[0269] The binding activity of anti-IL2RA antibodies (5D9, 7B5, 11C3, 11C12, 11D2, 13F9, and 14C2) to CHO-S-hIL2RA cells (CHO-S cells expressing human IL2RA (hIL2RA, SEQ ID NO: 64)) or CHO-S-fasIL2RA cells (CHO-S cells expressing monkey ( Macaca fascicularis ) IL2RA (fasIL2RA, SEQ ID NO: 65)) was verified by flow cytometry.

[0270] CHO-S-hIL2RA cells or CHO-S-fasIL2RA cells were cultured at 2×10 5 Cells were plated at a density of 100 cells / well in a 96-well plate. Serial dilutions of purified anti-IL2RA antibody (maximum concentration 9 μg / mL, 2-fold dilution, 11 steps) were added to each well and incubated at 4°C for 30 minutes. After washing once with PBS, the cells were incubated with secondary antibody Alexa Fluor. The cells were incubated with 647 anti-human IgG Fcγ (Jackson ImmunoResearch Laboratories, Inc., Cat#: 109-606-170) at 4°C for 15 minutes before flow cytometric analysis.

[0271] Cells were harvested and the mean fluorescence intensity (MFI) was measured. A fitted curve was generated using log(antibody concentration (μg / mL)) as the X-axis and mean MFI as the Y-axis. EC50 values ​​were determined, and the test results are shown in the table below.

[0272] Table 2. Antibody binding to cells

[0273]

[0274] The results showed that all seven antibodies, 5D9, 7B5, 11C3, 11C12, 11D2, 13F9, and 14C2, had good binding activity to both human and monkey IL2RA.

[0275] Example 4. Blocking experiment

[0276] Blockade of IL2RA ligand binding to IL2RA by flow cytometry

[0277] Using human CHO-S-hIL2RA cells, the blocking effect of anti-IL2RA antibodies on the binding of human IL2RA ligand (biotinylated human IL-2 protein with His and Avitag tags, ACROBiosystems Inc., Cat#: IL2-H82E4) to human IL2RA was tested by flow cytometry.

[0278] CHO-S-hIL2RA cells were seeded in 96-well plates (cell density was 2×10 5

[0279] cells / well) and incubated at 37°C overnight.

[0280] μg / mL (2-fold dilution, 11 steps in total) and incubate with cells at 4°C for 15 minutes, then add human IL2RA ligand to each well. The working concentration of human IL 2RA ligand is 2.5

[0281] μg / mL. The 96-well plate was incubated at 4°C for 20 minutes. After washing once with PBS, the cells were incubated with the secondary antibody APC-labeled streptavidin (APC Streptavidin, BioLegend, Cat#:

[0282] 405207) were incubated at 4°C for 15 min and then analyzed by flow cytometry.

[0283] The mean fluorescence intensity (MFI) was determined. A fitting curve was obtained using Log(antibody concentration (μg / mL)) or antibody concentration (μg / mL) as the X-axis and Log(mean MFI) or MFI as the Y-axis. The results are shown in Figure 2. Figures 1A-1B shown.

[0284] Daclizumab is a humanized monoclonal antibody that binds to the IL-2 receptor (α subunit, CD25) on activated T cells and inhibits IL-2-mediated lymphocyte activation, thereby preventing transplant rejection and the activation of inflammatory cytokine responses commonly seen in autoimmune and inflammatory diseases. Its VH and VL sequences are shown in SEQ ID NO:68 and SEQ ID NO:69, respectively.

[0285] There was no corresponding decrease in MFI when the concentration of antibodies (5D9, 7B5, 11C3, 11C12, 11D2, 13F9, and 14C2) was increased, indicating that the anti-IL2RA antibodies did not block the binding of human IL2RA to its ligand, whereas the daclizumab analogs did.

[0286] IL2 reporter cell blocking experiment

[0287] This experiment was performed to test whether anti-IL2RA antibodies 5D9, 7B5, 11C3, 11C12, 11D2, 13F9, and 14C2 could block the binding of mouse IL2 protein (mIL2) to IL2 reporter cells.

[0288] 20 μL of 100 ng / mL diluted mouse IL2 protein (mIL2, ACRO, Cat#: IL2-M52H3) was added to a 96-well plate. 20 μL of diluted antibody solution was then added to each well. The working concentration of the antibody was 100 μg / mL, 33.3 μg / mL, or 11.1 μg / mL. Subsequently, 160 μL of IL2 reporter cells (HEK293-Blue-IL2 cells, Invivogen, Cat#: hkb-il2) were seeded into a 96-well plate (cell density was 5 × 10 4 The 96-well plate was incubated at 37°C under 5% CO2 for 20-24 hours. After incubation, the plate was removed and 180 μL of QUANTI-Blue was added to each well. TM Solution (QUANTI Blue reagent: QUANTI Blue buffer: sterile water = 1:1:98) and 20 μL of cell supernatant. The plate was incubated at 37°C for 1 hour and then placed in a microplate reader to measure the OD value at 630 nm. The results are shown in Figure 2. Figures 2A-2B shown.

[0289] The results showed that the anti-IL2RA antibodies 5D9, 7B5, 11C3, 11C12, 11D2, 13F9, and 14C2 did not block the binding of mouse IL2 protein to IL2 reporter cells, while the daclizumab analog blocked the binding of mouse IL2 protein to IL2 reporter cells.

[0290] STAT5 phosphorylation blocking activity assay

[0291] human CD3 +T cells were incubated at 37°C for 2 hours and then seeded into 96-well plates (100 μL / well). The antibody was diluted and added to the wells and incubated with the cells at 37°C, 5% CO2 for 30 minutes (the working concentration of the diluted antibody was 10 μg / mL). Human IL2 protein (hIL2, ACROBiosystems Inc., Cat#: IL2-H4113) was then added to each well, and the 96-well plates were incubated at 37°C, 5% CO2 for 15 minutes. Subsequently, the cells were collected and single-cell suspensions were prepared from the samples. The cells were labeled with PE anti-STAT5 phosphorylation (Tyr694) antibody (BioLegend, Cat#: 936904) and then analyzed by flow cytometry.

[0292] 7G7B6 is a mouse IgG2 monoclonal antibody targeting human IL2RA that is in preclinical development at Leukemia Therapy. Its VH and VL sequences are shown in SEQ ID NO:66 and SEQ ID NO:67, respectively.

[0293] The results are as follows Figures 3A-3B Similar to the 7G7B6 analog-SI, the anti-IL2RA antibodies 5D9, 7B5, 11C3, 11C12, 11D2, 13F9, and 14C2 had little effect on human IL2-induced STAT5 phosphorylation, indicating that they are non-pSTAT5 blocking antibodies. The daclizumab analog significantly inhibited STAT5 phosphorylation, thus acting as a pSTAT5 blocking antibody.

[0294] Example 5. Antitumor activity in MC38 xenograft model

[0295] Antitumor activity of anti-IL2RA antibodies 7B5 and 5D9

[0296] The effect of anti-IL2RA antibodies on tumor growth in vivo was tested in a colon cancer model. Approximately 5×10 5 MC38 cells (mouse colon cancer cells). When the mouse tumor volume reaches about 100mm 3 At the time of the study, mice were randomly divided into different groups based on tumor size. Mice were then injected intraperitoneally (ip) with phosphate-buffered saline (PBS) or anti-IL2RA antibody. Details are shown in the table below.

[0297] Table 3. Grouping

[0298] Group Number of mice Antibody dose Route of administration frequency Total number of doses G1 6 PBS - ip BIW 6 G2 6 7B5 3 mg / kg ip BIW 6 G3 6 7B5 10 mg / kg ip BIW 6 G4 6 5D9 3 mg / kg ip BIW 6 G5 6 5D9 10 mg / kg ip BIW 6 G6 6 7G7B6 analogs 3 mg / kg ip BIW 6 G7 6 7G7B6 analogs 10 mg / kg ip BIW 6

[0299] The long and short axis lengths of the tumor were measured, and the tumor volume was calculated using the formula 0.5×(long axis)×(short axis) 2 calculate.

[0300] The tumor growth inhibition percentage (TGI%) was calculated using the following formula: TGI (%) = [1-(T i -T0) / (V i -V0)]×100. T i is the mean tumor volume of the treatment group on day i. T0 is the mean tumor volume of the treatment group on day 0. V i is the mean tumor volume of the control group on day i. V0 is the mean tumor volume of the control group on day 0.

[0301] The values ​​are expressed as mean ± standard error (SEM). Statistical analysis was performed using a T-test. P < 0.05 was considered a significant difference.

[0302] Mice in all groups showed increased weight. On the day of grouping (day 0), the average body weight of each group ranged from 21.2g to 21.5g. At the end of the experiment (day 21), the average body weight of each group ranged from 24.6g to 25.6g, with the average body weight change in each group ranging from 115.4% to 119.8%. These results demonstrate that these anti-IL2RA antibodies were well tolerated and showed no significant toxicity in mice.

[0303] The following table summarizes the results of this experiment, including the tumor volume on the day of grouping (day 0), 11 days after grouping (day 11), and 21 days after grouping (day 21); tumor growth inhibition value (TGI) and the statistical difference (P value) in tumor volume and body weight between the treatment group and the control group.

[0304] Table 4. Changes in tumor size

[0305]

[0306] The tumor volumes of mice in different groups after treatment with antibodies or PBS were as follows Figure 4 At the same dose level, anti-IL2RA antibodies (G2-G5) showed better tumor inhibition than the positive control 7G7B6 analogs (G6-G7) in a dose-dependent manner.

[0307] In another experiment, approximately 5 × 10 5 MC38 cells were used to determine the anti-tumor activity of 7B5 and 5D9. When the tumor volume reached approximately 80-90 mm 3 At the same time, mice were randomly divided into a control group and different treatment groups according to tumor size. Grouping and dosing details are shown in the table below.

[0308] Table 5. Grouping

[0309] Group Number of mice Antibody dose Route of administration frequency Total number of doses G1 6 PBS - ip BIW 6 G2 6 7B5 10 mg / kg ip BIW 6 G3 6 5D9 10 mg / kg ip BIW 6 G4 6 7G7B6 analogue-SI 10 mg / kg ip BIW 6 G5 6 7G7B6 analogs 10 mg / kg ip BIW 6

[0310] The body weights of mice in all groups increased, indicating that 7B5 and 5D9 were well tolerated and had no significant toxicity to the mice. The following table summarizes the results of this experiment, including tumor volumes on the day of grouping (Day 0), 10 days after grouping (Day 10), and at the end of the experiment (Day 21); TGI (%), and the statistical differences (P values) in tumor volume and body weight between the treatment and control groups.

[0311] Table 6. Changes in tumor size

[0312]

[0313] The tumor volumes of mice in different groups after treatment with antibodies or PBS were as follows Figure 5 As shown, at the 10 mg / kg dose level, the anti-IL2RA antibodies 7B5 and 5D9 showed better tumor inhibition than the 7G7B6 analog-SI and 7G7B6 analog.

[0314] Antitumor activity of anti-IL2RA antibodies 11C3, 11C12, 11D2, 13F9, and 14C2

[0315] B-hIL2RA mice were subcutaneously injected with approximately 5×10 5 MC38 cells (mouse colon cancer cells) were used to determine the anti-tumor activity of 11C3, 11C12, 11D2, 13F9, and 14C2. 3 At the time of the study, mice were randomly divided into different groups based on tumor size. Mice were then injected intraperitoneally (ip) with either PBS or anti-IL2RA antibody. Details are shown in the table below.

[0316] Table 7. Grouping

[0317]

[0318] The mice in different groups showed increased weight. On the day of grouping (day 0), the average weight of each group ranged from 20.7g to 21.7g. At the end of the experiment (day 22), the average weight of each group ranged from 23.8g to 24.9g, with the average weight change in each group ranging from 113.5% to 117.8%. These results demonstrate that these anti-IL2RA antibodies were well tolerated and showed no significant toxicity in mice.

[0319] The following table summarizes the results of this experiment, including the tumor volume on the day of grouping (day 0), 12 days after grouping (day 12), and 22 days after grouping (day 22); tumor growth inhibition value (TGI) and the statistical difference (P value) in tumor volume and body weight between the treatment group and the control group.

[0320] Table 8. Changes in tumor size

[0321]

[0322] The tumor volumes of mice in different groups after treatment with antibodies or PBS were as follows Figure 6 At a dose level of 10 mg / kg, anti-IL2RA antibodies (G3-G7) showed better tumor inhibition compared to PBS in the control group (G1) and the positive control 7G7B6 analog-SI (G2).

[0323] Example 6. Antitumor activity in GL261 xenograft model

[0324] Each B-hIL2RA mouse was subcutaneously injected with approximately 1×10 6 When the mouse tumor volume reached about 80-90mm 3 At the time of the study, mice were randomly divided into different groups based on tumor size. Mice were then injected intraperitoneally (ip) with either PBS or anti-IL2RA antibody. Details are shown in the table below.

[0325] Table 9. Grouping

[0326]

[0327] The mice in different groups showed an increase in weight. On the day of grouping (Day 0), the average weight of each group ranged from 20.5g to 20.9g. At the end of the experiment (Day 14), the average weight of each group ranged from 23.3g to 25.0g. The average weight change in each group ranged from 113.5% to 119.4%. The results showed that the tested antibodies were well tolerated and had no significant toxicity to the mice.

[0328] The following table summarizes the results of this experiment, including the tumor volume on the day of grouping (day 0), 7 days after grouping (day 7), and 14 days after grouping (day 14); tumor growth inhibition value (TGI) and the statistical difference (P value) in tumor volume and body weight between the treatment group and the control group.

[0329] Table 10. Changes in tumor size

[0330]

[0331] Compared with the control group (G1) treated with PBS, the treatment groups (G2-G4) showed better tumor inhibition. In addition, 7B5 showed better tumor inhibition than the positive control 7G7B6 analog, and the tumor inhibition effect of 5D9 was basically equivalent to that of the positive control 7G7B6 analog.

[0332] Other implementation plans

[0333] It should be understood that although the invention has been described in conjunction with the detailed description thereof, the foregoing description is intended to illustrate and not limit the scope of the invention, which is defined by the scope of the appended claims. Other aspects, advantages and modifications are within the scope of the following claims.

Claims

1. An antibody or antigen-binding fragment thereof that binds to interleukin-2 receptor alpha chain (IL2RA), comprising: a heavy chain variable region (VH) comprising complementarity determining regions (CDRs) 1, 2, and 3, wherein the VH CDR1 region comprises an amino acid sequence that is at least 80% identical to a selected VH CDR1 amino acid sequence, the VH CDR2 region comprises an amino acid sequence that is at least 80% identical to a selected VH CDR2 amino acid sequence, and the VH CDR3 region comprises an amino acid sequence that is at least 80% identical to a selected VH CDR3 amino acid sequence; and a light chain variable region (VL) comprising CDRs 1, 2, and 3, wherein the VL CDR1 region comprises an amino acid sequence that is at least 80% identical to a selected VL CDR1 amino acid sequence, the VL CDR2 region comprises an amino acid sequence that is at least 80% identical to a selected VL CDR2 amino acid sequence, and the VL CDR3 region comprises an amino acid sequence that is at least 80% identical to a selected VL CDR3 amino acid sequence, wherein the selected VH CDR 1, 2 and 3 amino acid sequences and the selected VL CDR 1, 2 and 3 amino acid sequences are one of the following: (1) The amino acid sequences of the selected VH CDRs 1, 2, and 3 are shown in SEQ ID NOs: 1, 2, and 3, respectively, and the selected VL The amino acid sequences of CDRs 1, 2, and 3 are shown in SEQ ID NOs: 43, 44, and 45, respectively; (2) The amino acid sequences of the selected VH CDRs 1, 2, and 3 are shown in SEQ ID NOs: 4, 5, and 6, respectively, and the selected VL The amino acid sequences of CDRs 1, 2, and 3 are shown in SEQ ID NOs: 46, 47, and 48, respectively; (3) The amino acid sequences of the selected VH CDRs 1, 2, and 3 are shown in SEQ ID NOs: 7, 8, and 9, respectively, and the selected VL The amino acid sequences of CDRs 1, 2, and 3 are shown in SEQ ID NOs: 49, 50, and 51, respectively; (4) the selected VH CDR 1, 2, and 3 amino acid sequences are shown in SEQ ID NOs: 10, 11, and 12, respectively, and the selected VLCDR 1, 2, and 3 amino acid sequences are shown in SEQ ID NOs: 49, 50, and 51, respectively; (5) the selected VH CDR 1, 2, and 3 amino acid sequences are shown in SEQ ID NOs: 13, 14, and 15, respectively, and the selected VLCDR 1, 2, and 3 amino acid sequences are shown in SEQ ID NOs: 49, 50, and 51, respectively; (6) the selected VH CDR 1, 2, and 3 amino acid sequences are shown in SEQ ID NOs: 16, 17, and 18, respectively, and the selected VLCDR 1, 2, and 3 amino acid sequences are shown in SEQ ID NOs: 49, 50, and 51, respectively; (7) the selected VH CDR 1, 2, and 3 amino acid sequences are shown in SEQ ID NOs: 19, 20, and 21, respectively, and the selected VLCDR 1, 2, and 3 amino acid sequences are shown in SEQ ID NOs: 49, 50, and 51, respectively; (8) the selected VH CDR 1, 2, and 3 amino acid sequences are shown in SEQ ID NOs: 22, 23, and 24, respectively, and the selected VLCDR 1, 2, and 3 amino acid sequences are shown in SEQ ID NOs: 43, 44, and 45, respectively; (9) the selected VH CDR 1, 2, and 3 amino acid sequences are shown in SEQ ID NOs: 25, 26, and 27, respectively, and the selected VLCDR 1, 2, and 3 amino acid sequences are shown in SEQ ID NOs: 46, 47, and 48, respectively; (10) the selected VH CDR 1, 2, and 3 amino acid sequences are shown in SEQ ID NOs: 28, 29, and 30, respectively, and the selected VLCDR 1, 2, and 3 amino acid sequences are shown in SEQ ID NOs: 49, 50, and 51, respectively; (11) the selected VH CDR 1, 2, and 3 amino acid sequences are shown in SEQ ID NOs: 31, 32, and 33, respectively, and the selected VLCDR 1, 2, and 3 amino acid sequences are shown in SEQ ID NOs: 49, 50, and 51, respectively; (12) the selected VH CDR 1, 2, and 3 amino acid sequences are shown in SEQ ID NOs: 34, 35, and 36, respectively, and the selected VLCDR 1, 2, and 3 amino acid sequences are shown in SEQ ID NOs: 49, 50, and 51, respectively; (13) the selected VH CDR 1, 2, and 3 amino acid sequences are shown in SEQ ID NOs: 37, 38, and 39, respectively, and the selected VLCDR 1, 2, and 3 amino acid sequences are shown in SEQ ID NOs: 49, 50, and 51, respectively; and (14) The selected VH CDR 1, 2, and 3 amino acid sequences are shown in SEQ ID NOs: 40, 41, and 42, respectively, and the selected VLCDR 1, 2, and 3 amino acid sequences are shown in SEQ ID NOs: 49, 50, and 51, respectively.

2. The antibody or antigen-binding fragment thereof of claim 1, wherein the VH comprises CDRs 1, 2, and 3 of the amino acid sequences shown in SEQ ID NOs: 1, 2, and 3, respectively, and the VL comprises CDRs 1, 2, and 3 of the amino acid sequences shown in SEQ ID NOs: 43, 44, and 45, respectively, according to the Kabat definition.

3. The antibody or antigen-binding fragment thereof of claim 1, wherein the VH comprises CDRs 1, 2, and 3 of the amino acid sequences shown in SEQ ID NOs: 4, 5, and 6, respectively, and the VL comprises CDRs 1, 2, and 3 of the amino acid sequences shown in SEQ ID NOs: 46, 47, and 48, respectively, according to the Kabat definition.

4. The antibody or antigen-binding fragment thereof of claim 1, wherein the VH comprises CDRs 1, 2, and 3 of the amino acid sequences shown in SEQ ID NOs: 7, 8, and 9, respectively, and the VL comprises CDRs 1, 2, and 3 of the amino acid sequences shown in SEQ ID NOs: 49, 50, and 51, respectively, according to the Kabat definition.

5. The antibody or antigen-binding fragment thereof of claim 1, wherein the VH comprises CDRs 1, 2, and 3 of the amino acid sequences shown in SEQ ID NOs: 10, 11, and 12, respectively, and the VL comprises CDRs 1, 2, and 3 of the amino acid sequences shown in SEQ ID NOs: 49, 50, and 51, respectively, according to the Kabat definition.

6. The antibody or antigen-binding fragment thereof of claim 1 , wherein the VH comprises CDRs 1, 2, and 3 of the amino acid sequences shown in SEQ ID NOs: 13, 14, and 15, respectively, and the VL comprises CDRs 1, 2, and 3 of the amino acid sequences shown in SEQ ID NOs: 49, 50, and 51, respectively, according to the Kabat definition.

7. The antibody or antigen-binding fragment thereof of claim 1, wherein the VH comprises CDRs 1, 2, and 3 of the amino acid sequences shown in SEQ ID NOs: 16, 17, and 18, respectively, and the VL comprises CDRs 1, 2, and 3 of the amino acid sequences shown in SEQ ID NOs: 49, 50, and 51, respectively, according to the Kabat definition.

8. The antibody or antigen-binding fragment thereof of claim 1, wherein the VH comprises CDRs 1, 2, and 3 of the amino acid sequences shown in SEQ ID NOs: 19, 20, and 21, respectively, and the VL comprises CDRs 1, 2, and 3 of the amino acid sequences shown in SEQ ID NOs: 49, 50, and 51, respectively, according to the Kabat definition.

9. The antibody or antigen-binding fragment thereof of any one of claims 1-8, wherein the antibody or antigen-binding fragment thereof specifically binds to human IL2RA or monkey IL2RA.

10. The antibody or antigen-binding fragment thereof of any one of claims 1-9, wherein the antibody or antigen-binding fragment thereof is a human or humanized antibody or antigen-binding fragment thereof (eg, a human IgG1 antibody).

11. The antibody or antigen-binding fragment thereof of any one of claims 1-10, wherein the antibody or antigen-binding fragment thereof is a single-chain variable fragment (scFv) or a multispecific antibody (eg, a bispecific antibody).

12. A nucleic acid comprising a polynucleotide encoding the following polypeptide: (1) an immunoglobulin heavy chain or fragment thereof comprising a heavy chain variable region (VH) comprising complementarity determining regions (CDRs) 1, 2, and 3 of the amino acid sequences set forth in SEQ ID NOs: 1, 2, and 3, respectively, and wherein the VH, when paired with a light chain variable region (VL) comprising the amino acid sequence set forth in SEQ ID NO: 53, binds to IL2RA; (2) an immunoglobulin light chain or fragment thereof comprising a light chain variable region (VL) comprising CDRs 1, 2, and 3 of the amino acid sequences set forth in SEQ ID NOs: 43, 44, and 45, respectively, and wherein the VL, when paired with a VH comprising the amino acid sequence set forth in SEQ ID NO: 52, binds to IL2RA; (3) an immunoglobulin heavy chain or fragment thereof comprising a heavy chain variable region (VH) comprising CDRs 1, 2, and 3 of the amino acid sequences set forth in SEQ ID NOs: 4, 5, and 6, respectively, and wherein the VH, when paired with a light chain variable region (VL) comprising the amino acid sequence set forth in SEQ ID NO: 55, binds to IL2RA; (4) an immunoglobulin light chain or fragment thereof comprising a light chain variable region (VL) comprising CDRs 1, 2, and 3 of the amino acid sequences set forth in SEQ ID NOs: 46, 47, and 48, respectively, and wherein the VL, when paired with a VH comprising the amino acid sequence set forth in SEQ ID NO: 54, binds to IL2RA; (5) an immunoglobulin heavy chain or fragment thereof comprising a heavy chain variable region (VH) comprising CDRs 1, 2, and 3 of the amino acid sequences set forth in SEQ ID NOs: 7, 8, and 9, respectively, and wherein the VH, when paired with a light chain variable region (VL) comprising the amino acid sequence set forth in SEQ ID NO: 61, binds to IL2RA; (6) an immunoglobulin light chain or fragment thereof comprising a light chain variable region (VL) comprising CDRs 1, 2, and 3 of the amino acid sequences set forth in SEQ ID NOs: 49, 50, and 51, respectively, and wherein the VL, when paired with a VH comprising the amino acid sequence set forth in SEQ ID NO: 56, binds to IL2RA; (7) an immunoglobulin heavy chain or fragment thereof comprising a heavy chain variable region (VH) comprising CDRs 1, 2, and 3 of the amino acid sequences set forth in SEQ ID NOs: 10, 11, and 12, respectively, and wherein the VH, when paired with a light chain variable region (VL) comprising the amino acid sequence set forth in SEQ ID NO: 61, binds to IL2RA; (8) an immunoglobulin light chain or fragment thereof comprising a light chain variable region (VL) comprising CDRs 1, 2, and 3 of the amino acid sequences set forth in SEQ ID NOs: 49, 50, and 51, respectively, and wherein the VL, when paired with a VH comprising the amino acid sequence set forth in SEQ ID NO: 57, binds to IL2RA; (9) an immunoglobulin heavy chain or fragment thereof comprising a heavy chain variable region (VH) comprising CDRs 1, 2, and 3 of the amino acid sequences set forth in SEQ ID NOs: 13, 14, and 15, respectively, and wherein the VH, when paired with a light chain variable region (VL) comprising the amino acid sequence set forth in SEQ ID NO: 61, binds to IL2RA; (10) an immunoglobulin light chain or fragment thereof comprising a light chain variable region (VL) comprising CDRs 1, 2, and 3 of the amino acid sequences set forth in SEQ ID NOs: 49, 50, and 51, respectively, and wherein the VL, when paired with a VH comprising the amino acid sequence set forth in SEQ ID NO: 58, binds to IL2RA; (11) an immunoglobulin heavy chain or fragment thereof comprising a heavy chain variable region (VH) comprising CDRs 1, 2, and 3 of the amino acid sequences set forth in SEQ ID NOs: 16, 17, and 18, respectively, and wherein the VH, when paired with a light chain variable region (VL) comprising the amino acid sequence set forth in SEQ ID NO: 61, binds to IL2RA; (12) an immunoglobulin light chain or fragment thereof comprising a light chain variable region (VL) comprising CDRs 1, 2, and 3 of the amino acid sequences set forth in SEQ ID NOs: 49, 50, and 51, respectively, and wherein the VL, when paired with a VH comprising the amino acid sequence set forth in SEQ ID NO: 59, binds to IL2RA; (13) an immunoglobulin heavy chain or fragment thereof comprising a heavy chain variable region (VH) comprising CDRs 1, 2, and 3 of the amino acid sequences set forth in SEQ ID NOs: 19, 20, and 21, respectively, and wherein the VH, when paired with a light chain variable region (VL) comprising the amino acid sequence set forth in SEQ ID NO: 61, binds to IL2RA; (14) an immunoglobulin light chain or fragment thereof comprising a light chain variable region (VL) comprising CDRs 1, 2, and 3 of the amino acid sequences set forth in SEQ ID NOs: 49, 50, and 51, respectively, and wherein the VL, when paired with a VH comprising the amino acid sequence set forth in SEQ ID NO: 60, binds to IL2RA; (15) an immunoglobulin heavy chain or fragment thereof comprising a heavy chain variable region (VH) comprising CDRs 1, 2, and 3 of the amino acid sequences set forth in SEQ ID NOs: 22, 23, and 24, respectively, and wherein the VH, when paired with a light chain variable region (VL) comprising the amino acid sequence set forth in SEQ ID NO: 53, binds to IL2RA; (16) an immunoglobulin heavy chain or fragment thereof comprising a heavy chain variable region (VH) comprising CDRs 1, 2, and 3 of the amino acid sequences set forth in SEQ ID NOs: 25, 26, and 27, respectively, and wherein the VH, when paired with a light chain variable region (VL) comprising the amino acid sequence set forth in SEQ ID NO: 55, binds to IL2RA; (17) an immunoglobulin heavy chain or fragment thereof comprising a heavy chain variable region (VH) comprising CDRs 1, 2, and 3 of the amino acid sequences shown in SEQ ID NOs: 28, 29, and 30, respectively, and wherein the VH, when paired with a light chain variable region (VL) comprising the amino acid sequence shown in SEQ ID NO: 61, binds to IL2RA; (18) an immunoglobulin heavy chain or fragment thereof comprising a heavy chain variable region (VH) comprising CDRs 1, 2, and 3 of the amino acid sequences shown in SEQ ID NOs: 31, 32, and 33, respectively, and wherein the VH, when paired with a light chain variable region (VL) comprising the amino acid sequence shown in SEQ ID NO: 61, binds to IL2RA; (19) an immunoglobulin heavy chain or fragment thereof comprising a heavy chain variable region (VH) comprising CDRs 1, 2, and 3 of the amino acid sequences shown in SEQ ID NOs: 34, 35, and 36, respectively, and wherein the VH, when paired with a light chain variable region (VL) comprising the amino acid sequence shown in SEQ ID NO: 61, binds to IL2RA; (20) an immunoglobulin heavy chain or fragment thereof comprising a heavy chain variable region (VH) comprising CDRs 1, 2, and 3 of the amino acid sequences set forth in SEQ ID NOs: 37, 38, and 39, respectively, and wherein the VH, when paired with a light chain variable region (VL) comprising the amino acid sequence set forth in SEQ ID NO: 61, binds to IL2RA; or (21) An immunoglobulin heavy chain or fragment thereof comprising a heavy chain variable region (VH) comprising CDRs 1, 2, and 3 of the amino acid sequences shown in SEQ ID NOs: 40, 41, and 42, respectively, and wherein the VH, when paired with a light chain variable region (VL) comprising the amino acid sequence shown in SEQ ID NO: 61, binds to IL2RA.

13. The nucleic acid of claim 12, wherein the nucleic acid comprises a polynucleotide encoding an immunoglobulin heavy chain or fragment thereof comprising a heavy chain variable region (VH) comprising complementarity determining regions (CDRs) 1, 2, and 3 of the amino acid sequences shown in SEQ ID NOs: 1, 2, and 3, respectively, or SEQ ID NOs: 22, 23, and 24, respectively.

14. The nucleic acid of claim 12, wherein the nucleic acid comprises a polynucleotide encoding an immunoglobulin light chain or fragment thereof comprising a light chain variable region (VL) comprising CDRs 1, 2, and 3 of the amino acid sequences shown in SEQ ID NOs: 43, 44, and 45, respectively.

15. The nucleic acid of claim 12, wherein the nucleic acid comprises a polynucleotide encoding an immunoglobulin heavy chain or fragment thereof comprising a heavy chain variable region (VH) comprising CDRs 1, 2, and 3 of the amino acid sequences shown in SEQ ID NOs: 4, 5, and 6, respectively, or SEQ ID NOs: 25, 26, and 27, respectively.

16. The nucleic acid of claim 12, wherein the nucleic acid comprises a polynucleotide encoding an immunoglobulin light chain or fragment thereof comprising a light chain variable region (VL) comprising CDRs 1, 2, and 3 of the amino acid sequences shown in SEQ ID NOs: 46, 47, and 48, respectively.

17. The nucleic acid of claim 12, wherein the nucleic acid comprises a polynucleotide encoding an immunoglobulin heavy chain or fragment thereof comprising a heavy chain variable region (VH) comprising CDRs 1, 2, and 3 of the amino acid sequences shown in SEQ ID NOs: 7, 8, and 9, respectively, or SEQ ID NOs: 28, 29, and 30, respectively.

18. The nucleic acid of claim 12, wherein the nucleic acid comprises a polynucleotide encoding an immunoglobulin heavy chain or fragment thereof comprising a heavy chain variable region (VH) comprising CDRs 1, 2, and 3 of the amino acid sequences shown in SEQ ID NOs: 10, 11, and 12, respectively, or SEQ ID NOs: 31, 32, and 33, respectively.

19. The nucleic acid of claim 12, wherein the nucleic acid comprises a polynucleotide encoding an immunoglobulin heavy chain or fragment thereof comprising a heavy chain variable region (VH) comprising CDRs 1, 2, and 3 of the amino acid sequences shown in SEQ ID NOs: 13, 14, and 15, respectively, or SEQ ID NOs: 34, 35, and 36, respectively.

20. The nucleic acid of claim 12, wherein the nucleic acid comprises a polynucleotide encoding an immunoglobulin heavy chain or fragment thereof comprising a heavy chain variable region (VH) comprising CDRs 1, 2, and 3 of the amino acid sequences shown in SEQ ID NOs: 16, 17, and 18, respectively, or SEQ ID NOs: 37, 38, and 39, respectively.

21. The nucleic acid of claim 12, wherein the nucleic acid comprises a polynucleotide encoding an immunoglobulin heavy chain or fragment thereof comprising a heavy chain variable region (VH) comprising CDRs 1, 2, and 3 of the amino acid sequences shown in SEQ ID NOs: 19, 20, and 21, respectively, or SEQ ID NOs: 40, 41, and 42, respectively.

22. The nucleic acid of claim 12, wherein the nucleic acid comprises a polynucleotide encoding an immunoglobulin light chain or fragment thereof comprising a light chain variable region (VL) comprising CDRs 1, 2, and 3 of the amino acid sequences shown in SEQ ID NOs: 49, 50, and 51, respectively.

23. The nucleic acid of any one of claims 12-22, wherein the VH specifically binds human IL2RA when paired with the VL, or the VL specifically binds human IL2RA or monkey IL2RA when paired with the VH.

24. The nucleic acid of any one of claims 12-23, wherein the immunoglobulin heavy chain or fragment thereof is a human or humanized immunoglobulin heavy chain or fragment thereof (e.g., a human IgG1 heavy chain or fragment thereof), and the immunoglobulin light chain or fragment thereof is a human or humanized immunoglobulin light chain or fragment thereof.

25. The nucleic acid of any one of claims 12-24, wherein the nucleic acid encodes a single-chain variable fragment (scFv), a multispecific antibody (e.g., a bispecific antibody), or a chimeric antigen receptor (CAR).

26. The nucleic acid of any one of claims 12-25, wherein the nucleic acid is a cDNA.

27. A vector comprising one or more nucleic acids of any one of claims 12-26.

28. A vector comprising two nucleic acids of any one of claims 12-26, wherein the VH region and VL region encoded by the vector jointly bind to IL2RA.

29. A pair of vectors, wherein each vector comprises a nucleic acid according to any one of claims 12-26, wherein the VH region and VL region encoded by the pair of vectors jointly bind to IL2RA.

30. A cell comprising the vector of claim 27 or 28, or a pair of vectors of claim 29.

31. The cell of claim 30, wherein the cell is a CHO cell.

32. A cell comprising one or more nucleic acids of any one of claims 12-26.

33. A cell comprising the two nucleic acids of any one of claims 12-26.

34. The cell of claim 33, wherein the VH and VL regions encoded by the two nucleic acids jointly bind IL2RA.

35. A method for producing an antibody or an antigen-binding fragment thereof, the method comprising (a) culturing the cell of any one of claims 30-34 under conditions sufficient for the cell to produce the antibody or antigen-binding fragment thereof; and (b) collecting the antibodies or antigen-binding fragments thereof produced by the cells.

36. An antibody or antigen-binding fragment thereof that binds to IL2RA, comprising a heavy chain variable region (VH) comprising an amino acid sequence that is at least 80% identical to a selected VH sequence; and a light chain variable region (VL) comprising an amino acid sequence that is at least 80% identical to a selected VL sequence, wherein the selected VH sequence and the selected VL sequence are one of the following: (1) the selected VH sequence is SEQ ID NO: 52, and the selected VL sequence is SEQ ID NO: 53; (2) the selected VH sequence is SEQ ID NO: 54, and the selected VL sequence is SEQ ID NO: 55; (3) the selected VH sequence is SEQ ID NO: 56, and the selected VL sequence is SEQ ID NO: 61; (4) the selected VH sequence is SEQ ID NO: 57, and the selected VL sequence is SEQ ID NO: 61; (5) the selected VH sequence is SEQ ID NO: 58, and the selected VL sequence is SEQ ID NO: 61; (6) the selected VH sequence is SEQ ID NO: 59, and the selected VL sequence is SEQ ID NO: 61; and (7) The selected VH sequence is SEQ ID NO:60, and the selected VL sequence is SEQ ID NO:

61.

37. The antibody or antigen-binding fragment thereof of claim 36, wherein the VH comprises the sequence shown in SEQ ID NO: 52, and the VL comprises the sequence shown in SEQ ID NO:

53.

38. The antibody or antigen-binding fragment thereof of claim 36, wherein the VH comprises the sequence set forth in SEQ ID NO: 54, and the VL comprises the sequence set forth in SEQ ID NO:

55.

39. The antibody or antigen-binding fragment thereof of claim 36, wherein the VH comprises the sequence set forth in SEQ ID NO: 56, and the VL comprises the sequence set forth in SEQ ID NO:

61.

40. The antibody or antigen-binding fragment thereof of claim 36, wherein the VH comprises the sequence set forth in SEQ ID NO: 57, and the VL comprises the sequence set forth in SEQ ID NO:

61.

41. The antibody or antigen-binding fragment thereof of claim 36, wherein the VH comprises the sequence set forth in SEQ ID NO: 58, and the VL comprises the sequence set forth in SEQ ID NO:

61.

42. The antibody or antigen-binding fragment thereof of claim 36, wherein the VH comprises the sequence set forth in SEQ ID NO: 59, and the VL comprises the sequence set forth in SEQ ID NO:

61.

43. The antibody or antigen-binding fragment thereof of claim 36, wherein the VH comprises the sequence set forth in SEQ ID NO: 60, and the VL comprises the sequence set forth in SEQ ID NO:

61.

44. The antibody or antigen-binding fragment thereof of any one of claims 36-43, wherein the antibody or antigen-binding fragment thereof specifically binds human IL2RA or monkey IL2RA.

45. The antibody or antigen-binding fragment thereof of any one of claims 36-44, wherein the antibody or antigen-binding fragment thereof is a human or humanized antibody or antigen-binding fragment thereof.

46. ​​The antibody or antigen-binding fragment thereof of any one of claims 36-45, wherein the antibody or antigen-binding fragment thereof is a single-chain variable fragment (scFv) or a multispecific antibody (e.g., a bispecific antibody).

47. An antibody or antigen-binding fragment thereof that cross-competes with the antibody or antigen-binding fragment thereof of any one of claims 1-11 and 36-46.

48. An antibody or antigen-binding fragment thereof that binds to IL2RA, comprising a heavy chain variable region (VH) comprising VH CDR1, VH CDR2, and VH CDR3 identical to the VH CDR1, VH CDR2, and VH CDR3 of the selected VH sequence; and a light chain variable region (VL) comprising a VL CDR1, a VL CDR2, and a VL CDR3 that are identical to the VL CDR1, VL CDR2, and VL CDR3 of a selected VL sequence, wherein the selected VH sequence and the selected VL sequence are one of the following: (1) the selected VH sequence is SEQ ID NO: 52, and the selected VL sequence is SEQ ID NO: 53; (2) the selected VH sequence is SEQ ID NO: 54, and the selected VL sequence is SEQ ID NO: 55; (3) the selected VH sequence is SEQ ID NO: 56, and the selected VL sequence is SEQ ID NO: 61; (4) the selected VH sequence is SEQ ID NO: 57, and the selected VL sequence is SEQ ID NO: 61; (5) the selected VH sequence is SEQ ID NO: 58, and the selected VL sequence is SEQ ID NO: 61; (6) the selected VH sequence is SEQ ID NO: 59, and the selected VL sequence is SEQ ID NO: 61; and (7) The selected VH sequence is SEQ ID NO:60, and the selected VL sequence is SEQ ID NO:

61.

49. An antibody-drug conjugate comprising the antibody or antigen-binding fragment thereof of any one of claims 1-11 and 36-48, covalently bound to a therapeutic agent.

50. The antibody-drug conjugate of claim 49, wherein the therapeutic agent is a cytotoxic agent or a cytostatic agent.

51. A method of treating a subject having cancer, the method comprising administering to the subject a therapeutically effective amount of a composition comprising the antibody or antigen-binding fragment thereof of any one of claims 1-11 and 36-48, or the antibody-drug conjugate of claim 49 or 50.

52. The method of claim 51, wherein the subject has a solid tumor, brain cancer, lung cancer, melanoma, prostate cancer, esophageal squamous cell carcinoma, leukemia, lymphoma, ovarian cancer, colorectal cancer, head and neck cancer, bladder cancer, kidney cancer, and breast cancer.

53. The method of claim 51 or 52, further comprising administering to the subject a therapeutically effective amount of an anti-OX40 antibody, an anti-PD1 antibody, an anti-PDL1 antibody, an anti-PDL2 antibody, an anti-LAG-3 antibody, an anti-TIGIT antibody, an anti-CTLA-4 antibody, an anti-GITR antibody, an anti-TIM-3 antibody, an anti-4-1BB antibody, and / or an anti-CD40 antibody.

54. A method for reducing tumor growth rate, the method comprising The tumor cells are contacted with an effective amount of a composition comprising the antibody or antigen-binding fragment thereof of any one of claims 1-11 and 36-48, or the antibody-drug conjugate of claim 49 or 50.

55. A method for killing tumor cells, the method comprising The tumor cells are contacted with an effective amount of a composition comprising the antibody or antigen-binding fragment thereof of any one of claims 1-11 and 36-48, or the antibody-drug conjugate of claim 49 or 50.

56. A pharmaceutical composition comprising the antibody or antigen-binding fragment thereof according to any one of claims 1-11 and 36-48, and a pharmaceutically acceptable carrier.

57. A pharmaceutical composition comprising the antibody-drug conjugate of claim 49 or 50, and a pharmaceutically acceptable carrier.

58. A chimeric antigen receptor (CAR) comprising the antibody or antigen-binding fragment thereof of any one of claims 1-11 and 36-48.

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