Methods and compositions for treatment of cancer with Anti-human nectin-4 therapeutics

Abstract

The present disclosure provides methods, compositions, uses, and kits for treatment of solid cancer (e.g., urothelial cancer (UC), head and neck squamous cell carcinoma (HNSCC), cervical cancer (CC), endometrial carcinoma (EC), or triple-negative breast cancer (TNBC)) using anti-human Nectin-4 therapeutics.

Description

Attorney Docket No. 14463-733-228METHODS AND COMPOSITIONS FOR TREATMENT OF CANCER WITH ANTIHUMAN NECTIN-4 THERAPEUTICSCROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims priority to International Patent Application No. PCT / CN2024 / 134292, filed November 25, 2024, the disclosure of which is incorporated by reference herein in its entirety.SEQUENCE LISTING

[0002] This application contains an electronic Sequence Listing which has been submitted in XML file format with this application, the entire content of which is incorporated by reference herein in its entirety. The Sequence Listing XML file submitted with this application is entitled “14463-733-228_SEQ_LISTING.xml”, was created on November 21, 2024, and is 67,181 bytes in size.FIELD

[0003] This disclosure relates generally to methods, compositions, uses, and kits for treatment of solid cancer (e.g., urothelial cancer (UC), head and neck squamous cell carcinoma (HNSCC), cervical cancer (CC), endometrial carcinoma (EC), or triple-negative breast cancer (TNBC)) using anti-human Nectin-4 therapeutics.BACKGROUND

[0004] Nectin-4 is a member of an immunoglobulin superfamily which mainly functions in mediating calcium ion-independent cell adhesion junctions. Nectin-4 is a type I transmembrane protein, the structure of which comprises an extracellular segment formed by one IgV domain and two IgC domains, a transmembrane region, and a C-terminal intracellular domain which functionally binds the PDZ domain of Afadin protein.

[0005] In humans, Nectin-4 is normally expressed in keratinocytes of the skin, sweat glands, hair follicles, transitional epithelium of the bladder, salivary gland ducts, esophagus, breast, and stomach. In neoplasia, Nectin-4 promotes tumor cell survival and proliferation by inhibiting anoikis and activating the PI3K-AKT signaling pathway. Studies have shown that Nectin-4 is expressed in gastric cancer, hepatocellular carcinoma, ovarian cancer, head and neck cancer, and esophageal cancer, and is strongly expressed in non-small cell lung cancer, bladder cancer, breast cancer, and pancreatic cancer. High expression of Nectin-4 in tumors is closely related to the poor prognosis of patients.1NAI-5006826801V1

[0006] At present, the antibody drug conjugate (ADC) targeting Nectin-4, enfortumab vedotin, is available for the treatment of urothelial carcinoma. Enfortumab vedotin is fully-human anti- Nectin-4 antibody conjugated to the drug moiety VC-MMAE, with a drug to antibody ratio (DAR) value of 4 (see, international patent publication no. WO2012 / 047724). The main clinical toxic and side effects of enfortumab vedotin are fatigue (54%), with some patients experiencing > grade 3 side effects of anemia (8%), hyponatremia (7%), urinary tract infection (7%), and hyperglycemia (6%). Some patients receiving enfortumab vedotin had serious drug-related side effects, including respiratory failure, urinary tract obstruction, diabetic ketoacidosis, and multiple organ failure.

[0007] Therefore, there is a need in the art for methods and compositions for the safe and effective treatment of Nectin-4-expressing cancers in humans.SUMMARY

[0008] The present disclosure provides, inter alia, methods, compositions, uses, and kits for the treatment of a cancer, e.g., a cancer expressing Nectin-4.

[0009] In one aspect, provided herein is a method for treating a cancer in a patient, the method comprising administering to the patient a therapeutically effective amount of an anti-Nectin-4 antibody conjugate, wherein the antibody conjugate comprises an anti-Nectin-4 antibody, or antigen-binding fragment thereof, and at least one therapeutic agent. In specific embodiments, the anti-Nectin-4 antibody, or antigen-binding fragment thereof, is an anti-human Nectin-4 antibody, or antigen-binding fragment thereof. In specific embodiments, the anti-Nectin-4 antibody, or antigen-binding fragment thereof, of the disclosed methods is a composition of anti-Nectin-4 antibodies, or antigen-binding fragments thereof.

[0010] In some embodiments of the method, the anti-Nectin-4 antibody, or antigen-binding fragment thereof, comprises:(a) the following six complementarity determining regions (CDRs) for heavy and light chains defined according to the Chothia numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 3;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 4; CDR-H3 comprising the sequence as set forth in SEQ ID NO: 5; CDR-L1 comprising the sequence as set forth in SEQ ID NO: 6; CDR-L2 comprising the sequence as set forth in SEQ ID NO: 7; and CDR-L3 comprising the sequence as set forth in SEQ ID NO: 8; or(b) the following six CDRs for heavy and light chains defined according to the Abm numbering system:2NAI-5006826801V1CDR-H1 comprising the sequence as set forth in SEQ ID NO: 9;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 10;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 5;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 6;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 7; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 8; or(c) the following six CDRs for heavy and light chains defined according to the Kabat numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 11;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 12 or 63;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 5;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 6;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 7; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 8; or(d) the following six CDRs for heavy and light chains defined according to the IMGT numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 13;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 14;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 15;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 16;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 17; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 8.

[0011] In some embodiments of the method, the anti-Nectin-4 antibody, or antigen-binding fragment thereof, comprises:(a) the following six CDRs for heavy and light chains defined according to the Chothia numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 21 or 18;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 22;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 23;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 24;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 25; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 26; or(b) the following six CDRs for heavy and light chains defined according to the Abm numbering system:3NAI-5006826801V1CDR-H1 comprising the sequence as set forth in SEQ ID NO: 27 or 36;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 28;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 23;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 24;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 25; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 26; or(c) the following six CDRs for heavy and light chains defined according to the Kabat numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 29 or 44;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 30;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 23;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 24;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 25; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 26; or(d) the following six CDRs for heavy and light chains defined according to the IMGT numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 31 or 54;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 32;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 33;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 34;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 35; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 26.

[0012] In some embodiments of the method, the anti-Nectin-4 antibody, or antigen-binding fragment thereof, comprises:(a) the following six CDRs for heavy and light chains defined according to the Chothia numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 39;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 40;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 41;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 42;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 43; andCDR-L3 comprising SEQ ID NO: 8 or a variant thereof; or(b) the following six CDRs for heavy and light chains defined according to the Abm numbering system:4NAI-5006826801V1CDR-H1 comprising the sequence as set forth in SEQ ID NO: 45;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 46;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 41;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 42;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 43; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 8; or(c) the following six CDRs for heavy and light chains defined according to the Kabat numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 47;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 48 or 64;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 41;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 42;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 43; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 8; or(d) the following six CDRs for heavy and light chains defined according to the IMGT numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 49;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 50;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 51;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 52;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 53; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 8.

[0013] In some embodiments of the method, the anti-Nectin-4 antibody, or antigen-binding fragment thereof, comprises: a VH having the sequence as set forth in SEQ ID NO: 19 or SEQ ID NO: 57 and a VL having the sequence as set forth in SEQ ID NO: 20 or SEQ ID NO: 58; a VH having the sequence as set forth in SEQ ID NO: 1 or SEQ ID NO: 55 and a VL having the sequence as set forth in SEQ ID NO: 2 or SEQ ID NO: 56; or a VH having the sequence as set forth in SEQ ID NO: 37 or SEQ ID NO: 59 and a VL having the sequence as set forth in SEQ ID NO: 38 or SEQ ID NO: 60.

[0014] In some embodiments of the method, the anti-Nectin-4 antibody, or antigen-binding fragment thereof, comprises:(1) a VH having the sequence as set forth in SEQ ID NO: 57 and a VL having the sequence as set forth in SEQ ID NO: 58;5NAI-5006826801V1(2) a VH having the sequence as set forth in SEQ ID NO: 1 and a VL having the sequence as set forth in SEQ ID NO: 2;(3) a VH having the sequence as set forth in SEQ ID NO: 19 and a VL having the sequence as set forth in SEQ ID NO: 20;(4) a VH having the sequence as set forth in SEQ ID NO: 37 and a VL having the sequence as set forth in SEQ ID NO: 38;(5) a VH having the sequence as set forth in SEQ ID NO: 55 and a VL having the sequence as set forth in SEQ ID NO: 56; or(6) a VH having the sequence as set forth in SEQ ID NO: 59 and a VL having the sequence as set forth in SEQ ID NO: 60.

[0015] In some embodiments of the method, the anti-Nectin-4 antibody, or antigen-binding fragment thereof, further comprises a human immunoglobulin heavy chain constant region or a variant thereof, and a light chain constant region or a variant thereof, wherein the variant has a substitution, deletion, or addition of one or more amino acids compared to the wild-type sequence from which it is derived. In some embodiments of the method, the variant has a substitution, deletion, or addition of up to 5 amino acids compared to the wild-type sequence from which it was derived.

[0016] In some embodiments of the method, the anti-Nectin-4 antibody, or antigen-binding fragment thereof, further comprises a heavy chain constant region having the sequence as set forth in SEQ ID NO: 61 and a light chain constant region having the sequence as set forth in SEQ ID NO: 62.

[0017] In some embodiments, the anti-Nectin-4 antibody, or antigen-binding fragment thereof, comprises a heavy chain and light chain, wherein(1) the heavy chain comprises: a VH having the sequence as set forth in SEQ ID NO: 57, and a heavy chain constant region having the sequence as set forth in SEQ ID NO: 61, and the light chain comprises: a VL having the sequence as set forth in SEQ ID NO: 58, and a light chain constant region having the sequence as set forth in SEQ ID NO: 62; or(2) the heavy chain comprises: a VH having the sequence as set forth in SEQ ID NO: 1, and a heavy chain constant region having the sequence as set forth in SEQ ID NO: 61, and the light chain comprises: a VL having the sequence as set forth in SEQ ID NO: 2, and a light chain constant region having the sequence as set forth in SEQ ID NO: 62; or(3) the heavy chain comprises: a VH having the sequence as set forth in SEQ ID NO: 19, and a heavy chain constant region having the sequence as set forth in SEQ ID NO: 61, and the light6NAI-5006826801V1chain comprises: a VL having the sequence as set forth in SEQ ID NO: 20, and a light chain constant region having the sequence as set forth in SEQ ID NO: 62; or(4) the heavy chain comprises: a VH having the sequence as set forth in SEQ ID NO: 37, and a heavy chain constant region having the sequence as set forth in SEQ ID NO: 61, and the light chain comprises: a VL having the sequence as set forth in SEQ ID NO: 38, and a light chain constant region having the sequence as set forth in SEQ ID NO: 62; or(5) the heavy chain comprises: a VH having the sequence as set forth in SEQ ID NO: 55, and a heavy chain constant region having the sequence as set forth in SEQ ID NO: 61, and the light chain comprises: a VL having the sequence as set forth in SEQ ID NO: 56, and a light chain constant region having the sequence as set forth in SEQ ID NO: 62; or(6) the heavy chain comprises: a VH having the sequence as set forth in SEQ ID NO: 59, and a heavy chain constant region having the sequence as set forth in SEQ ID NO: 61, and the light chain comprises: a VL having the sequence as set forth in SEQ ID NO: 60, and a light chain constant region having the sequence as set forth in SEQ ID NO: 62.

[0018] In some embodiments, the anti-Nectin-4 antibody, or antigen-binding fragment thereof, comprises a heavy chain and light chain, wherein (1) the heavy chain comprises: a VH having the sequence as set forth in SEQ ID NO: 57, and a heavy chain constant region having the sequence as set forth in SEQ ID NO: 61, and the light chain comprises: a VL having the sequence as set forth in SEQ ID NO: 58, and a light chain constant region having the sequence as set forth in SEQ ID NO: 62.

[0019] In some embodiments, the anti-Nectin-4 antibody, or antigen-binding fragment thereof, comprises a heavy chain and light chain, wherein (2) the heavy chain comprises: a VH having the sequence as set forth in SEQ ID NO: 1, and a heavy chain constant region having the sequence as set forth in SEQ ID NO: 61, and the light chain comprises: a VL having the sequence as set forth in SEQ ID NO: 2, and a light chain constant region having the sequence as set forth in SEQ ID NO: 62.

[0020] In some embodiments, the anti-Nectin-4 antibody, or antigen-binding fragment thereof, comprises a heavy chain and light chain, wherein (3) the heavy chain comprises: a VH having the sequence as set forth in SEQ ID NO: 19, and a heavy chain constant region having the sequence as set forth in SEQ ID NO: 61, and the light chain comprises: a VL having the sequence as set forth in SEQ ID NO: 20, and a light chain constant region having the sequence as set forth in SEQ ID NO: 62.

[0021] In some embodiments, the anti-Nectin-4 antibody, or antigen-binding fragment thereof, comprises a heavy chain and light chain, wherein (4) the heavy chain comprises: a VH having7NAI-5006826801V1the sequence as set forth in SEQ ID NO: 37, and a heavy chain constant region having the sequence as set forth in SEQ ID NO: 61, and the light chain comprises: a VL having the sequence as set forth in SEQ ID NO: 38, and a light chain constant region having the sequence as set forth in SEQ ID NO: 62.

[0022] In some embodiments, the anti-Nectin-4 antibody, or antigen-binding fragment thereof, comprises a heavy chain and light chain, wherein (5) the heavy chain comprises: a VH having the sequence as set forth in SEQ ID NO: 55, and a heavy chain constant region having the sequence as set forth in SEQ ID NO: 61, and the light chain comprises: a VL having the sequence as set forth in SEQ ID NO: 56, and a light chain constant region having the sequence as set forth in SEQ ID NO: 62.

[0023] In some embodiments, the anti-Nectin-4 antibody, or antigen-binding fragment thereof, comprises a heavy chain and light chain, wherein (6) the heavy chain comprises: a VH having the sequence as set forth in SEQ ID NO: 59, and a heavy chain constant region having the sequence as set forth in SEQ ID NO: 61, and the light chain comprises: a VL having the sequence as set forth in SEQ ID NO: 60, and a light chain constant region having the sequence as set forth in SEQ ID NO: 62.

[0024] In some embodiments of the method, the at least one therapeutic agent of the antibody conjugate is a cytotoxic agent.

[0025] In some embodiments of the method, the antibody conjugate has the structure shown in Formula (I),A-(L-D)y(i); wherein, A represents an antibody or antigen-binding fragment thereof described herein;L comprises an amino acid or a peptide consisting of 2-10 amino acids, or L is selectedfrom Vai, Cit, Phe, Lys, D-Val, Leu, Gly, Ala, Asn,8NAI-5006826801V1of position 1 and position 2 represents the position where L and A are connected, and the other represents the position where L and D are connected;D is selected from the group consisting of cytotoxic agents, therapeutic antibodies, radioisotopes, oligonucleotides and analogs thereof, biologically active peptides, protein toxins and enzymes; preferably, D is a molecule with anti-tumor biological activity;9NAI-5006826801V1y is an integer of 1 to 10.

[0026] In some embodiments of the method, the antibody conjugate is formed by connecting the (L-D) moiety of Formula (1) with a sulfhydryl group of A, for example, the sulfhydryl group of the side chain of a cysteine residue of A, through a thioether bond,Formula (1), wherein the antibody conjugate has the structure of Formula (2):10NAI-5006826801V1Formula (2), wherein y is an integer from 1 to 10, and A represents an antibody or antigen-binding fragment thereof described herein.

[0027] In some embodiments of the method, the molar ratio (DAR value) of therapeutic agent to the antibody, or antigen-binding fragment thereof, in the composition is a decimal or integer between 1 and 10. In some embodiments of the method, the DAR is a decimal or integer between 1 and 8.

[0028] In some embodiments of the method, the DAR value is 7.0, 7.03, 7.1, 7.12, 7.2, 7.3, 7.4, 7.5, 7.6, 7.7, 7.8, 7.9 or 8.0. In some embodiments of the method, the DAR value is 7.71.

[0029] In some embodiments of the method, the age of the patient is greater than or equal to 18 years of age.

[0030] In some embodiments of the method, the cancer is selected from the group consisting of urothelial carcinoma (UC), head and neck squamous cell carcinoma (HNSCC), cervical cancer (CC), endometrial carcinoma (EC), and triple-negative breast cancer (TNBC). In some embodiments of the method, the cancer is selected from the group consisting of cervical cancer (CC) and endometrial carcinoma (EC).

[0031] In some embodiments of the method, the cancer is (i) locally recurrent unresectable and / or (ii) metastatic. In some embodiments of the method, the patient was previously treated for the cancer. In some embodiments of the method, the patient was previously treated with a chemotherapy, an anti-PD-l / anti-PD-Ll therapy, tisotumab vedotin, and / or enfortumab vedotin.

[0032] In some embodiments of the method, the antibody conjugate is administered to the patient via intravenous infusion. In some embodiments of the method, the antibody conjugate is administered to the patient at a dose of about 4 mg / kg or about 5 mg / kg. In some embodiments of the method, the antibody conjugate is administered to the patient every two weeks (Q2W).

[0033] In another aspect, provided herein is a use of an anti-Nectin-4 antibody conjugate in the manufacture of a medicament for treating a cancer in a patient, wherein the antibody conjugate comprises an anti-Nectin-4 antibody, or antigen-binding fragment thereof, and at least one therapeutic agent.11NAI-5006826801V1

[0034] In some embodiments of the use, the anti-Nectin-4 antibody, or antigen-binding fragment thereof, comprises:(a) the following six complementarity determining regions (CDRs) for heavy and light chains defined according to the Chothia numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 3;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 4;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 5;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 6;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 7; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 8; or(b) the following six CDRs for heavy and light chains defined according to the Abm numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 9;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 10;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 5;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 6;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 7; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 8; or(c) the following six CDRs for heavy and light chains defined according to the Kabat numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 11;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 12 or 63;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 5;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 6;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 7; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 8; or(d) the following six CDRs for heavy and light chains defined according to the IMGT numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 13;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 14;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 15;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 16;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 17; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 8.12NAI-5006826801V1

[0035] In some embodiments of the use, the anti-Nectin-4 antibody, or antigen-binding fragment thereof, comprises:(a) the following six CDRs for heavy and light chains defined according to the Chothia numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 21 or 18;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 22;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 23;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 24;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 25; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 26; or(b) the following six CDRs for heavy and light chains defined according to the Abm numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 27 or 36;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 28;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 23;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 24;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 25; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 26; or(c) the following six CDRs for heavy and light chains defined according to the Kabat numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 29 or 44;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 30;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 23;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 24;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 25; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 26; or(d) the following six CDRs for heavy and light chains defined according to the IMGT numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 31 or 54;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 32;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 33;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 34;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 35; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 26.13NAI-5006826801V1

[0036] In some embodiments of the use, the anti-Nectin-4 antibody, or antigen-binding fragment thereof, comprises:(a) the following six CDRs for heavy and light chains defined according to the Chothia numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 39;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 40;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 41;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 42;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 43; andCDR-L3 comprising SEQ ID NO: 8 or a variant thereof; or(b) the following six CDRs for heavy and light chains defined according to the Abm numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 45;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 46;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 41;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 42;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 43; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 8; or(c) the following six CDRs for heavy and light chains defined according to the Kabat numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 47;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 48 or 64;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 41;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 42;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 43; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 8; or(d) the following six CDRs for heavy and light chains defined according to the IMGT numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 49;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 50;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 51;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 52;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 53; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 8.14NAI-5006826801V1

[0037] In some embodiments of the use, the anti-Nectin-4 antibody, or antigen-binding fragment thereof, comprises: a VH having the sequence as set forth in SEQ ID NO: 19 or SEQ ID NO: 57 and a VL having the sequence as set forth in SEQ ID NO: 20 or SEQ ID NO: 58; a VH having the sequence as set forth in SEQ ID NO: 1 or SEQ ID NO: 55 and a VL having the sequence as set forth in SEQ ID NO: 2 or SEQ ID NO: 56; or a VH having the sequence as set forth in SEQ ID NO: 37 or SEQ ID NO: 59 and a VL having the sequence as set forth in SEQ ID NO: 38 or SEQ ID NO: 60.

[0038] In some embodiments of the use, the anti-Nectin-4 antibody, or antigen-binding fragment thereof, comprises:(1) a VH having the sequence as set forth in SEQ ID NO: 57 and a VL having the sequence as set forth in SEQ ID NO: 58;(2) a VH having the sequence as set forth in SEQ ID NO: 1 and a VL having the sequence as set forth in SEQ ID NO: 2;(3) a VH having the sequence as set forth in SEQ ID NO: 19 and a VL having the sequence as set forth in SEQ ID NO: 20;(4) a VH having the sequence as set forth in SEQ ID NO: 37 and a VL having the sequence as set forth in SEQ ID NO: 38;(5) a VH having the sequence as set forth in SEQ ID NO: 55 and a VL having the sequence as set forth in SEQ ID NO: 56; or(6) a VH having the sequence as set forth in SEQ ID NO: 59 and a VL having the sequence as set forth in SEQ ID NO: 60.

[0039] In some embodiments of the use, the anti-Nectin-4 antibody, or antigen-binding fragment thereof, further comprises a human immunoglobulin heavy chain constant region or a variant thereof, and a light chain constant region or a variant thereof, wherein the variant has a substitution, deletion, or addition of one or more amino acids compared to the wild-type sequence from which it is derived. In some embodiments of the use, the variant has a substitution, deletion, or addition of up to 5 amino acids compared to the wild-type sequence from which it was derived. In some embodiments of the use, the anti-Nectin-4 antibody, or antigen-binding fragment thereof, further comprises a heavy chain constant region having the sequence as set forth in SEQ ID NO: 61 and a light chain constant region having the sequence as set forth in SEQ ID NO: 62.

[0040] In some embodiments of the use, the at least one therapeutic agent of the antibody conjugate is a cytotoxic agent.15NAI-5006826801V1

[0041] In some embodiments of the use, the antibody conjugate has the structure shown in Formula (I),A-(L-D)y(i); wherein, A represents the antibody or antigen-binding fragment thereof as described herein;L comprises an amino acid or a peptide consisting of 2-10 amino acids, or L is selected16NAI-5006826801V1of position 1 and position 2 represents the position where L and A are connected, and the other represents the position where L and D are connected;D is selected from the group consisting of cytotoxic agents, therapeutic antibodies, radioisotopes, oligonucleotides and analogs thereof, biologically active peptides, protein toxins and enzymes; preferably, D is a molecule with anti-tumor biological activity;y is an integer of 1 to 10.

[0042] In some embodiments of the use, the antibody conjugate is formed by connecting the (L-D) moiety of Formula (1) with a sulfhydryl group of A, for example, the sulfhydryl group of the side chain of a cysteine residue of A, through a thioether bond,17NAI-5006826801V1Formula (1), wherein the antibody conjugate has the structure of Formula (2):Formula (2), wherein y is an integer from 1 to 10, and A represents the antibody or antigen-binding fragment thereof as described herein.

[0043] In some embodiments of the use, the molar ratio (DAR value) of therapeutic agent to the antibody, or antigen-binding fragment thereof, in the composition is a decimal or integer between 1 and 10. In some embodiments of the use, the DAR is a decimal or integer between 1 and 8.

[0044] In some embodiments of the use, the DAR value is 7.0, 7.03, 7.1, 7.12, 7.2, 7.3, 7.4, 7.5, 7.6, 7.7, 7.8, 7.9 or 8.0. In some embodiments of the use, the DAR value is 7.71.

[0045] In some embodiments of the use, the age of the patient is greater than or equal to 18 years of age.

[0046] In some embodiments of the use, the cancer is selected from the group consisting of urothelial carcinoma (UC), head and neck squamous cell carcinoma (HNSCC), cervical cancer (CC), endometrial carcinoma (EC), and triple-negative breast cancer (TNBC). In some embodiments of the use, the cancer is selected from the group consisting of cervical cancer (CC) and endometrial carcinoma (EC).

[0047] In some embodiments of the use, the cancer is (i) locally recurrent unresectable and / or (ii) metastatic. In some embodiments of the use, the patient was previously treated for the cancer. In some embodiments of the use, the patient was previously treated with a chemotherapy, an anti-PD-l / anti-PD-Ll therapy, tisotumab vedotin, and / or enfortumab vedotin.

[0048] In some embodiments of the use, the antibody conjugate is administered to the patient via intravenous infusion. In some embodiments of the use, the antibody conjugate is administered to the patient at a dose of about 4 mg / kg or about 5 mg / kg. In some embodiments of the use, the antibody conjugate is administered to the patient every two weeks (Q2W). In18NAI-5006826801V1some embodiments of the method or use, the antibody conjugate or medicament is administered at least twice.

[0049] In another aspect, provided herein is a kit comprising an antibody conjugate for use as described herein, and instructions to administer the antibody conjugate to a patient having a cancer.

[0050] In some embodiments of the kit, the age of the patient is greater than or equal to 18 years of age.

[0051] In some embodiments of the kit, the cancer is selected from the group consisting of urothelial carcinoma (UC), head and neck squamous cell carcinoma (HNSCC), cervical cancer (CC), endometrial carcinoma (EC), and triple-negative breast cancer (TNBC). In some embodiments of the kit, the cancer is selected from the group consisting of cervical cancer (CC) and endometrial carcinoma (EC). In some embodiments of the kit, the cancer is (i) locally recurrent unresectable and / or (ii) metastatic.

[0052] In some embodiments of the method, use, or kit, the antibody conjugate or medicament is used in combination with one or more therapeutic agents selected from the group consisting of antineoplastic agents, chemotherapeutic agents, radioisotopes, immune checkpoint inhibitors.

[0053] In some embodiments of the method, use, or kit, the antibody conjugate or medicament is used in combination with one or more therapeutic regimens selected from the group consisting of surgery, radiotherapy, chemotherapy, targeted therapy, immunotherapy, hormone therapy, angiogenesis inhibition, and palliative care.

[0054] The summary of the technology described above is non-limiting and other features and advantages of the technology will be apparent from the following detailed description, and from the claims.BRIEF DESCRIPTION OF THE DRAWINGS

[0055] FIG. 1A is a line graph plotting the results of anti-human Nectin-4 ADCs 31HZ-TL001 and 74HZ-TL001 killing A549-Nectin-4 cells. FIG. IB is a line graph plotting the results of anti-human Nectin-4 ADC 31HZ-TL001 and 74HZ-TL001 killing A549 cells. FIG. 1C is a line graph plotting the results of anti-human Nectin-4 ADCs 31HZ-TL001 and 56HZ-TL001 killing T24-Nectin-4 cells. FIG. ID is a line graph plotting the results of anti-human Nectin-4 ADC 31HZ-TL001 and 56HZ-TL001 killing T24 cells.

[0056] FIG. 2A is a line graph plotting the results of anti-human Nectin-4 ADC 31HZ-TL001 and 74HZ-TL001 killing T47D cells. FIG. 2B is a line graph plotting the results of anti-human Nectin-4 ADC 31HZ-TL001 and 74HZ-TL001 killing MDA-MB-468 cells. FIG. 2C is a line19NAI-5006826801V1graph plotting the results of anti-human Nectin-4 ADC 31HZ-TL001 and 56HZ-TL001 killing NCI-N87 cells.

[0057] FIG. 3 is a line graph plotting the change in tumor volume over time of each group of NOD / SCID mice in a subcutaneous NCI-H322M cell xenograft tumor model.

[0058] FIG. 4 is a line graph plotting the change in body weight over time of each group of NOD / SCID mice in a subcutaneous NCI-H322M cell xenograft tumor model.

[0059] FIG. 5 is a line graph plotting the change in tumor volume over time of each group of NOD / SCID mice in a subcutaneous HuPrime® gastric cancer BL9200 PDX model.

[0060] FIG. 6 is a line graph plotting the change in body weight over time of each group of NOD / SCID mice in a subcutaneous HuPrime® gastric cancer BL9200 PDX model.

[0061] FIG. 7 is a schematic diagram showing the 56HZ-TL001 study design. EV, enfortumab vedotin; HNSCC, head and neck squamous cell carcinoma; IV, intravenous; q2w, every 2 weeks; TNBC, triple negative breast cancer.DETAILED DESCRIPTION

[0062] The present disclosure provides, inter alia, methods, compositions, uses, and kits for the treatment of a cancer, e.g., a cancer expressing Nectin-4. In particular, the present disclosure provides methods, compositions, uses, and kits for the treatment of a cancer comprising administering to a patient a therapeutically effective amount of an anti-Nectin-4 antibody conjugate, wherein the antibody conjugate comprises an anti-Nectin-4 antibody, or antigenbinding fragment thereof, and optionally, at least one therapeutic agent.Definitions

[0063] Listed below are definitions of various terms used herein. These definitions apply to the terms as they are used throughout this specification and claims, unless otherwise limited in specific instances, either individually or as part of a larger group.

[0064] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art. Generally, the nomenclature used herein and the laboratory procedures in cell culture, molecular genetics, organic chemistry, and peptide chemistry are those well-known and commonly employed in the art.

[0065] As used herein, the articles “a” and “an” refer to one or to more than one (i.e., to at least one) of the grammatical object of the article. By way of example, “an element” means one element or more than one element. Furthermore, use of the term “including” as well as other forms, such as “include,” “includes,” and “included,” is not limiting.20NAI-5006826801V1

[0066] Reference to “or” indicates either or both possibilities unless the context clearly dictates one of the indicated possibilities. In some cases, “and / or” was employed to highlight either or both possibilities.

[0067] The term “about,” when modifying the quantity (e.g., mg) of a substance or composition, or the value of a parameter characterizing a step in a method, or the like, refers to variation in the numerical quantity that can occur, for example, through typical measuring, handling and sampling procedures involved in the preparation, characterization and / or use of the substance or composition; through inadvertent error in these procedures; through differences in the manufacture, source, or purity of the ingredients employed to make or use the compositions or carry out the procedures; and the like. In certain embodiments, “about” can mean a variation of ± 0.1%, ± 0.5%, ± 1%, ± 2%, ± 3%, ± 4%, ± 5%, ± 6%, ± 7%, ± 8%, ± 9% or ± 10%.

[0068] All ranges disclosed herein are inclusive of the recited endpoint and independently combinable (for example, the range of “from 50 mg to 500 mg” is inclusive of the endpoints, 50 mg and 500 mg, and all the intermediate values). The endpoints of the ranges and any values disclosed herein are not limited to the precise range or value; they are sufficiently imprecise to include values approximating these ranges and / or values.

[0069] As used herein, the term “comprising” may include the embodiments “consisting of’ and “consisting essentially of.” The terms “comprise(s),” “include(s),” “having,” “has,” “may,” “contain(s),” and variants thereof, as used herein, are intended to be open-ended transitional phrases, terms, or words that require the presence of the named ingredients / steps and permit the presence of other ingredients / steps. However, such description should be construed as also describing compositions or processes as “consisting of’ and “consisting essentially of’ the enumerated components.

[0070] “ Consists essentially of,” and variations such as “consist essentially of’ or “consisting essentially of,” as used herein, indicate the inclusion of any recited elements or group of elements, and the optional inclusion of other elements, of similar or different nature than the recited elements, that do not materially change the basic or novel properties of the specified dosage regimen, method, or composition. As a non-limiting example, an anti-Nectin-4 antibody that consists essentially of a recited amino acid sequence may also include one or more amino acids, including substitutions of one or more amino acid residues, which do not materially affect the properties of the binding compound.

[0071] “Administration” and “treatment,” as it applies to an animal, human, experimental subject, cell, tissue, organ, or biological fluid, refers to contact of an exogenous pharmaceutical, therapeutic, diagnostic agent, or composition to the animal, human, subject, cell, tissue, organ,21NAI-5006826801V1or biological fluid. “Treat” or “treating” a cancer (e.g., a cancer expressing Nectin-4), as used herein, means to administer an anti-Nectin-4 antibody, antigen-binding fragment, or antibody conjugate, alone or in combination with one or more additional therapeutic agents, to a subject having a cancer, or diagnosed with cancer. “Treatment” achieves at least one positive therapeutic effect, such as for example, reduced number of cancer cells, reduced tumor size, reduced rate of cancer cell infiltration into peripheral organs, or reduced rate of tumor metastasis or tumor growth. “Treatment” may include one or more of the following: inducing / increasing an antitumor immune response, decreasing the number of one or more tumor markers, halting or delaying the growth of a tumor or blood cancer or progression of disease such as cancer, stabilization of disease, inhibiting the growth or survival of tumor cells, eliminating or reducing the size of one or more cancerous lesions or tumors, decreasing the level of one or more tumor markers, ameliorating or abrogating the clinical manifestations of disease, reducing the severity or duration of the clinical symptoms of disease such as cancer, prolonging the survival of a patient relative to the expected survival in a similar untreated patient, and inducing complete or partial remission of a cancerous condition or other disease.

[0072] Positive therapeutic effects in cancer can be measured in a number of ways (see, e.g., W. A. Weber, J. Nucl. Med. 5O: 1S-1OS (2009)). For example, with respect to tumor growth inhibition, according to NCI standards, a T / C ^42% is the minimum level of anti-tumor activity. A T / C < 10% is considered a high anti -tumor activity level, with T / C (%) = Median tumor volume of the treated / Median tumor volume of the control x 100. In some embodiments, the treatment achieved by a therapeutically effective amount is any of progression free survival (PFS), disease free survival (DFS) or overall survival (OS). PFS, also referred to as “Time to Tumor Progression” indicates the length of time during and after treatment that the cancer does not grow, and includes the amount of time patients have experienced a complete response or a partial response, as well as the amount of time patients have experienced stable disease. DFS refers to the length of time during and after treatment that the patient remains free of disease. OS refers to a prolongation in life expectancy as compared to naive or untreated individuals or patients. While an embodiment of the treatment methods, compositions and uses of the present invention may not be effective in achieving a positive therapeutic effect in every patient, it should do so in a statistically significant number of subjects as determined by any statistical test known in the art such as the Student’s t-test, the chi2-test, the U-test according to Mann and Whitney, the Kruskal-Wallis test (H-test), Jonckheere-Terpstra-test and the Wilcoxon-test.22NAI-5006826801V1

[0073] “RECIST 1.1 Response Criteria” as used herein means the definitions set forth in Eisenhauer, E.A. et al., Eur. J. Cancer 45:228-247 (2009) for target lesions or nontarget lesions, as appropriate based on the context in which response is being measured.

[0074] The term “patient” (alternatively referred to as “subject” or “individual” interchangeably herein) refers to a mammal (e.g., human, non-human primate (e.g., cynomolgus monkey), rat, mouse, dog, cat, rabbit) capable of being treated with the methods and compositions of the invention, most preferably a human. In some embodiments, the patient is an adult patient. In other embodiments, the patient is a pediatric patient.

[0075] A “biomarker” is an objectively measured indicator, compound, or molecule that reflects the presence, or likely progression, or successful treatment of a particular condition. Biomarkers have long been used in drug development, and the discovery and validation of new efficacy biomarkers is expected to improve predictive disease models, reduce the time and cost associated with drug development, and increase the success rate of translating experimental drugs into clinical therapeutics. In addition, biomarkers are valuable in early detection of disease development, changes in disease status, and effectiveness of behavioral modifications and therapeutics in disease control.

[0076] As used herein, the term “antibody” refers to any form of antibody that exhibits the desired biological activity. Thus, it is used in the broadest sense and specifically covers, but is not limited to, monoclonal antibodies (including full length monoclonal antibodies comprising two light chains and two heavy chains), polyclonal antibodies, multispecific antibodies (e.g., bispecific antibodies), humanized antibodies, fully human antibodies, and chimeric antibodies.

[0077] In general, the basic antibody structural unit comprises a tetramer. Each tetramer includes two identical pairs of polypeptide chains, each pair having one “light” (e.g., about 25 kDa) and one “heavy” chain (e.g., about 50-70 kDa). The amino-terminal portion of each chain includes a variable region of about 100 to 110 or more amino acids primarily responsible for antigen recognition. The carboxy-terminal portion of the heavy chain may define a constant region primarily responsible for effector function. Typically, human light chains are classified as kappa and lambda light chains. Furthermore, human heavy chains are typically classified as mu, delta, gamma, alpha, or epsilon, and define the antibody's isotype as IgM, IgD, IgG, IgA, and IgE, respectively. Within light and heavy chains, the variable and constant regions are joined by a “J” region of about 12 or more amino acids, with the heavy chain also including a “D” region of about 10 more amino acids. See generally, Fundamental Immunology Ch. 7 (Paul, W., ed., 2nd ed. Raven Press, N.Y. (1989). Antibodies and antigen-binding fragments of any of these classes or subclasses of antibodies may be utilized and administered in the present disclosure.23NAI-5006826801V1

[0078] “Variable regions” or “V region” as used herein means the segment of IgG chains which is variable in sequence between different antibodies. The V region typically extends to Kabat residue 109 in the light chain and 113 in the heavy chain. The variable regions of each light / heavy chain pair form the antibody binding site. The “heavy chain variable region” or “heavy chain variable domain” is also referred to herein as “VH,” and the “light chain variable region” or “light chain variable domain” is also referred to herein as “VL .” Thus, in general, an intact antibody has two binding sites. Except in bifunctional or bispecific antibodies, the two binding sites are, in general, the same.

[0079] Typically, the variable domains of both the heavy and light chains comprise three hypervariable regions, also called complementarity determining regions (referred to here as “CDRs”), which are located within relatively conserved framework regions (FR). The CDRs are usually aligned by the framework regions, enabling binding to a specific epitope. In general, from N-terminal to C- terminal, both light and heavy chains variable domains comprise FR1, CDR1, FR2, CDR2, FR3, CDR3, and FR4. The assignment of amino acids to each domain is, generally, in accordance with the definitions of Sequences of Proteins of Immunological Interest, Kabat, et al.; National Institutes of Health, Bethesda, Md.; 5th ed.; NIH Publ. No. 91- 3242 (1991); Kabat (1978) Adv. Prot. Chem. 32: 1-75; Kabat, et al., (1977) J. Biol. Chem.252:6609-6616; Chothia, et al., (1987) J Mol. Biol. 196:901-917 or Chothia, et al., (1989) Nature 342:878-883.

[0080] “Kabat,” as used herein, means an immunoglobulin alignment and numbering system pioneered by Elvin A. Kabat ((1991) Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, Md.). Other alternate alignment and number systems (e.g., Chothia, Abm, IGMT, Aho, etc.) may be used in combination with, or in place of, the Kabat numbering system for the immunoglobulins described herein.

[0081] As used herein, the term “framework” or “FR” residues refers to those variable domain residues other than the hypervariable region residues defined herein as CDR residues.

[0082] As used herein, the term “hypervariable region” refers to the amino acid residues of an antibody or antigen-binding fragment thereof that are responsible for antigen-binding. The hypervariable region comprises amino acid residues from a CDR (i.e., CDRL1, CDRL2 and CDRL3 in the light chain variable domain and CDRH1, CDRH2 and CDRH3 in the heavy chain variable domain). See Kabat et al. (1991) Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, Md. (defining the CDR regions of an antibody by sequence); see also Chothia and Lesk (1987) J. Mol. Biol. 196: 901- 917 (defining the CDR regions of an antibody by structure).24NAI-5006826801V1

[0083] “Monoclonal antibody,” “mAb,” or “Mab,” as used herein, refers to a population of substantially homogeneous antibodies, i.e., the antibody molecules comprising the population are identical in amino acid sequence except for possible naturally occurring mutations that may be present in minor amounts. In contrast, polyclonal antibody preparations typically include a multitude of different antibodies having different amino acid sequences in their variable domains, particularly their CDRs, which are often specific for different epitopes. The modifier “monoclonal” indicates the character of the antibody as being obtained from a substantially homogeneous population of antibodies, and is not to be construed as requiring production of the antibody by any particular method. For example, the monoclonal antibodies to be used in accordance with the present invention may be made by the hybridoma method first described by Kohler et al. (1975) Nature 256: 495, or may be made by recombinant DNA methods (see, e.g., U.S. Pat. No. 4,816,567). The “monoclonal antibodies” may also be isolated from phage antibody libraries using the techniques described in Clackson et al. (1991) Nature 352: 624-628 and Marks et al. (1991) J. Mol. Biol. 222: 581-597, for example. See also Presta (2005) J. Allergy Clin. Immunol. 116:731.

[0084] As used herein, unless otherwise indicated, “antibody fragment,” “antigen binding fragment thereof,” or “antigen-binding fragment" refer to antigen-binding fragments of antibodies, i.e., antibody fragments that retain the ability to bind specifically to the antigen (e.g., Nectin-4) bound by the full-length antibody, e.g., fragments that retain one or more CDR (e.g., CDR1, CDR2, and CDR3) regions. Examples of antigen-binding fragments include, but are not limited to, Fab, Fab’, F(ab’)2, and Fv fragments; diabodies; linear antibodies; single-chain antibody molecules, e.g., scFv; and multispecific antibodies formed from antibody fragments.

[0085] A “Fab fragment” is comprised of one light chain and the CHI and variable regions of one heavy chain. The heavy chain of a Fab molecule cannot form a disulfide bond with another heavy chain molecule. A Fab fragment can be the product of papain cleavage of an antibody. In other examples, a Fab fragment may be produced using recombinant antibody production.

[0086] An “Fc” region contains two heavy chain fragments comprising the CH2 and CH3 domains of an antibody. The two heavy chain fragments are held together by two or more disulfide bonds and by hydrophobic interactions of the CH3 domains.

[0087] A “Fab’ fragment” contains one light chain and a portion or fragment of one heavy chain that contains the VH domain and the CHI domain and also the region between the CHI and CH2 domains, such that an interchain disulfide bond can be formed between the two heavy chains of two Fab’ fragments to form a F(ab’)2 fragment.25NAI-5006826801V1

[0088] A “F(ab’)2 fragment” contains two light chains and two heavy chains containing a portion of the constant region between the CHI and CH2 domains, such that an interchain disulfide bond is formed between the two heavy chains. A F(ab’)2 fragment thus is composed of two Fab’ fragments that are held together by a disulfide bond between the two heavy chains. A F(ab’)2 fragment can be the product of pepsin cleavage of an antibody.

[0089] The “Fv region” comprises the variable regions from both the heavy and light chains but lacks the constant regions.

[0090] The term “single-chain Fv” or “scFv” antibody refers to an antibody fragment comprising the VH and VL domains of an antibody, in which these domains are present in a single polypeptide chain. Generally, the Fv polypeptide further comprises a polypeptide linker between the VH and VL domains which enables the scFv to form the desired structure for antigen-binding. For a review of scFv, see Pluckthun (1994) The Pharmacology of Monoclonal Antibodies, vol. 113, Rosenburg and Moore eds. Springer-Verlag, New York, pp. 269-315. See also International Patent Application Publication No. WO 88 / 01649 and U.S. Pat. Nos. 4,946, 778 and 5,260,203. The scFv may have an orientation in the N-terminal to C-terminal direction of VH-linker- VL or VL-linker-VH.

[0091] A “bivalent antibody” comprises two antigen-binding sites. In some instances, the two binding sites have the same antigen specificities. However, bivalent antibodies may be bispecific (see below).

[0092] As used herein, the term “diabodies” refers to small antibody fragments with two antigen-binding sites, which fragments comprise a VH connected to a VL in the same polypeptide chain (VH-VL or VL-VH). By using a linker that is too short to allow pairing between the two domains on the same chain, the domains are forced to pair with the complementary domains of another chain and create two antigen-binding sites. Diabodies are described more fully in, e.g., EP 404,097; WO 93 / 11161; and Holliger et al. (1993) Proc. Natl. Acad. Sci. USA 90: 6444-6448. For a review of engineered antibody variants generally see Holliger and Hudson (2005) Nat. Biotechnol. 23: 1126-1136.

[0093] An antibody that “specifically binds to” a specified target protein (e.g., Nectin-4) is an antibody that exhibits preferential binding to that target as compared to other proteins, but this specificity does not require absolute binding specificity. An antibody is considered “specific” for its intended target if its binding is determinative of the presence of the target protein in a sample, e.g., without producing undesired results such as false positives. Antibodies, or binding fragments thereof, useful in the present invention will bind to the target protein with an affinity that is at least two-fold greater, preferably at least ten times greater, more preferably at least 20-26NAI-5006826801V1times greater, and most preferably at least 100-times greater than the affinity with non-target proteins. As used herein, an antibody is said to bind specifically to a polypeptide comprising a given amino acid sequence, e.g., the amino acid sequence of a mature human Nectin-4 molecule, if it binds to polypeptides comprising that sequence but does not bind to proteins lacking that sequence.

[0094] As used herein, a “chimeric antibody” is an antibody having the variable domain from a first antibody and the constant domain from a second antibody, where the first and second antibodies are from different species. See, e.g., U.S. Pat. No. 4,816,567; and Morrison et al., (1984) Proc. Natl. Acad. Sci. USA 81 : 6851-6855. Typically, the variable domains are obtained from an antibody from an experimental animal (the “parental antibody”), such as a rodent, and the constant domain sequences are obtained from human antibodies, so that the resulting chimeric antibody will be less likely to elicit an adverse immune response in a human subject than the parental (e.g., mouse) antibody.

[0095] “Humanized antibody” refers to forms of antibodies that contain sequences from nonhuman (e.g., murine) antibodies as well as human antibodies. Such antibodies contain minimal sequence derived from non-human immunoglobulin. In general, the humanized antibody will comprise substantially all of at least one, and typically two, variable domains, in which all or substantially all of the hypervariable loops correspond to those of a non-human immunoglobulin and all or substantially all of the FR regions are those of a human immunoglobulin sequence. The humanized antibody optionally also will comprise at least a portion of an immunoglobulin constant region (Fc), typically that of a human immunoglobulin. The prefix “hum”, “hu” or “h” is added to antibody clone designations when necessary to distinguish humanized antibodies from parental rodent antibodies. The humanized forms of rodent antibodies will generally comprise the same CDR sequences of the parental rodent antibodies, although certain amino acid substitutions may be included to increase affinity, increase stability of the humanized antibody, or for other reasons.

[0096] As used herein, “germline sequence” refers to a sequence of unrearranged immunoglobulin DNA sequences. Any suitable source of unrearranged immunoglobulin sequences may be used. Human germline sequences may be obtained, for example, from JOINSOLVER germline databases on the website for the National Institute of Arthritis and Musculoskeletal and Skin Diseases of the United States National Institutes of Health. Mouse germline sequences may be obtained, for example, as described in Giudicelli et al. (2005) Nucleic Acids Res. 33:D256-D261.27NAI-5006826801V1

[0097] “ Tissue section” refers to a single part or piece of a tissue sample, e.g., a thin slice of tissue cut from a sample of a normal tissue or of a tumor.

[0098] “ Tumor” as it applies to a subject diagnosed with, or suspected of having, a cancer refers to a malignant or potentially malignant neoplasm or tissue mass of any size and includes primary tumors and secondary neoplasms. A solid tumor is an abnormal growth or mass of tissue that usually does not contain cysts or liquid areas. Different types of solid tumors are named for the type of cells that form them. Examples of solid tumors are sarcomas, carcinomas, and lymphomas. Leukemias (cancers of the blood) generally do not form solid tumors (National Cancer Institute, Dictionary of Cancer Terms).

[0099] “Conservatively modified variants” or “conservative substitution” refers to substitutions of amino acids in a protein with other amino acids having similar characteristics (e.g., charge, side-chain size, hydrophobicity / hydrophilicity, backbone conformation and rigidity, and the like), such that the changes can frequently be made without altering the biological activity of the protein. Those of skill in this art recognize that, in general, single amino acid substitutions in non-essential regions of a polypeptide do not substantially alter biological activity (see, e.g., Watson et al. (1987) Molecular Biology of the Gene, The Benjamin / Cummings Pub. Co., p. 224 (4th Ed.)). In addition, substitutions of structurally or functionally similar amino acids are less likely to disrupt biological activity. Exemplary conservative substitutions are set forth in the table below.28NAI-5006826801V1

[0100] “Function-conservative variants,” as used herein, refers to antibodies or fragments thereof in which one or more amino acid residues have been changed without altering a desired property, such as an antigen affinity and / or specificity. Such variants include, but are not limited to, replacement of an amino acid with one having similar properties, such as the conservative amino acid substitutions set forth in the table above.

[0101] Typically, an antibody or antigen-binding fragment thereof as used herein is modified in some way retains at least 10% of its binding activity (when compared to the parental antibody) when that activity is expressed on a molar basis. Preferably, an antibody or antigenbinding fragment thereof of the invention retains at least 20%, 50%, 70%, 80%, 90%, 95% or 100% or more of the Nectin-4 binding affinity as the parental antibody. It is also intended that an antibody or antigen-binding fragment thereof of the invention can include conservative or non-conservative amino acid substitutions (e.g., conservative variants or function-conserved variants of the antibody) that do not substantially alter its biologic activity.

[0102] The term “effector function” as used herein refers to one or more of antibody dependent dell mediated cytotoxic activity (ADCC), complement-dependent cytotoxic activity (CDC) mediated responses, Fc-mediated phagocytosis or antibody dependent cellular phagocytosis (ADCP) and antibody recycling via the FcRn receptor.

[0103] As used herein, the terms “N-glycan” and “glycoform” are used interchangeably and refer to an N-linked oligosaccharide, for example, one that is attached by an asparagine-N- acetylglucosamine linkage to an asparagine residue of a polypeptide. N-linked glycoproteins contain an N-acetylglucosamine residue linked to the amide nitrogen of an asparagine residue in the protein. The predominant sugars found on glycoproteins are glucose, galactose, mannose, fucose, N-acetylgalactosamine (GalNAc), N-acetylglucosamine (GlcNAc) and sialic acid (e.g., N-acetyl-neuraminic acid (NANA)). The processing of the sugar groups occurs co- translationally in the lumen of the ER and continues post-translationally in the Golgi apparatus for N-linked glycoproteins.

[0104] N-glycans have a common pentasaccharide core of MansGlcNAc2 (“Man” refers to mannose; “Glc” refers to glucose; and "NAc" refers to N-acetyl; GlcNAc refers to N- acetylglucosamine). Usually, N-glycan structures are presented with the non-reducing end to the left and the reducing end to the right. The reducing end of the N-glycan is the end that is attached to the Asn residue comprising the glycosylation site on the protein. N-glycans differ with respect to the number of branches (antennae) comprising peripheral sugars (e.g., GlcNAc, galactose, fucose and sialic acid) that are added to the MamGlcN Ac2 (“Mans”) core structure which is also referred to as the “trimannose core,” the “pentasaccharide core,” or the29NAI-5006826801V1“paucimannose core.” N-glycans are classified according to their branched constituents (e.g., high mannose, complex or hybrid). A “high mannose” type N-glycan has five or more mannose residues. A “complex” type N-glycan typically has at least one GlcNAc attached to the 1,3 mannose arm and at least one GlcNAc attached to the 1,6 mannose arm of a “trimannose” core. Complex N-glycans may also have galactose (“Gal”) or N- acetylgalactosamine (“GalNAc”) residues that are optionally modified with sialic acid or derivatives (e.g., “NANA” or “NeuAc,” where “Neu” refers to neuraminic acid and “Ac” refers to acetyl). Complex N-glycans may also have intrachain substitutions comprising “bisecting” GlcNAc and core fucose (“Fuc”). Complex N-glycans may also have multiple antennae on the “trimannose core,” often referred to as “multiple antennary glycans.” A “hybrid” N-glycan has at least one GlcNAc on the terminal of the 1,3 mannose arm of the trimannose core and zero or more mannoses on the 1,6 mannose arm of the trimannose core. The various N-glycans are also referred to as “glycoforms.”

[0105] “ Isolated” antibodies or antigen-binding fragments thereof are at least partially free of other biological molecules from the cells or cell cultures in which they are produced. Such biological molecules include nucleic acids, proteins, lipids, carbohydrates, or other material such as cellular debris and growth medium. An isolated antibody or antigen-binding fragment thereof may further be at least partially free of expression system components such as biological molecules from a host cell or of the growth medium thereof. Generally, the term “isolated” is not intended to refer to a complete absence of such biological molecules or to an absence of water, buffers, or salts or to components of a pharmaceutical formulation that includes the antibodies or fragments.

[0106] “ Isolated nucleic acid molecule” or “isolated polynucleotide” means a DNA or RNA of genomic, mRNA, cDNA, or synthetic origin or some combination thereof which is not associated with all or a portion of a polynucleotide in which the isolated polynucleotide is found in nature, or is linked to a polynucleotide to which it is not linked in nature. For purposes of this disclosure, it should be understood that “a nucleic acid molecule comprising” a particular nucleotide sequence does not encompass intact chromosomes. Isolated nucleic acid molecules comprising specified nucleic acid sequences may include, in addition to the specified sequences, coding sequences for up to ten or even up to twenty or more other proteins or portions or fragments thereof, or may include operably linked regulatory sequences that control expression of the coding region of the recited nucleic acid sequences, and / or may include vector sequences.

[0107] The phrase “control sequences” refers to DNA sequences necessary for the expression

[0108] of an operably linked coding sequence in a particular host organism. The control sequences that are suitable for prokaryotes, for example, include a promoter, optionally an30NAI-5006826801V1operator sequence, and a ribosome binding site. Eukaryotic cells are known to use promoters, polyadenylation signals, and enhancers.

[0109] “Percentage of sequence identity,” “percent sequence identity,” “percent identity,” and “percent identical” are used herein to refer to comparisons between polynucleotide sequences or polypeptide sequences, and are determined by comparing two optimally aligned sequences over a comparison window, in which the portion of the polynucleotide or polypeptide sequence in the comparison window may comprise additions or deletions (i.e., gaps) as compared to the reference sequence for optimal alignment of the two sequences. The percentage is calculated by determining the number of positions at which either the identical nucleic acid base or amino acid residue occurs in both sequences or a nucleic acid base or amino acid residue is aligned with a gap to yield the number of matched positions, dividing the number of matched positions by the total number of positions in the window of comparison and multiplying the result by 100 to yield the percentage of sequence identity. Determination of optimal alignment and percent sequence identity can be performed using the BLAST and BLAST 2.0 algorithms (see e.g., Altschul et al., 1990, J. Mol. Biol. 215: 403-410; and Altschul et al., 1977, Nucleic Acids Res. 3389-3402). Software for performing BLAST analyses is publicly available through the National Center for Biotechnology Information website.

[0110] Briefly, the BLAST analyses involve first identifying high scoring sequence pairs (HSPs) by identifying short words of length W in the query sequence, which either match or satisfy some positive-valued threshold score T when aligned with a word of the same length in a database sequence. T is referred to as the neighborhood word score threshold (Altschul et al., supra). These initial neighborhood word hits act as seeds for initiating searches to find longer HSPs containing them. The word hits are then extended in both directions along each sequence for as far as the cumulative alignment score can be increased. Cumulative scores are calculated using, for nucleotide sequences, the parameters M (reward score for a pair of matching residues; always >0) and N (penalty score for mismatching residues; always <0). For amino acid sequences, a scoring matrix is used to calculate the cumulative score. Extension of the word hits in each direction are halted when: the cumulative alignment score falls off by the quantity X from its maximum achieved value; the cumulative score goes to zero or below, due to the accumulation of one or more negative-scoring residue alignments; or the end of either sequence is reached. The BLAST algorithm parameters W, T, and X determine the sensitivity and speed of the alignment. The BLASTN program (for nucleotide sequences) uses as defaults a word length (W) of 11, an expectation (E) of 10, M = 5, N = -4, and a comparison of both strands. For amino acid sequences, the BLASTP program uses as defaults a word length (W) of 3, an31NAI-5006826801V1expectation (E) of 10, and the BLOSUM62 scoring matrix (see Henikoff and Henikoff, 1989, PROC. NATL. ACAD. SCI. USA 89: 10915).[oni] Numerous other algorithms are available that function similarly to BLAST in providing percent identity for two sequences. Optimal alignment of sequences for comparison can be conducted, e.g., by the local homology algorithm of Smith and Waterman, 1981, Adv. Appl. Math. 2:482, by the homology alignment algorithm of Needleman and Wunsch, 1970, J. Mol. Biol. 48:443, by the search for similarity method of Pearson and Lipman, 1988, Proc. Natl. Acad. Sci. USA 85:2444, by computerized implementations of these algorithms (GAP, BESTFIT, FASTA, and TFASTA in the GCG Wisconsin Software Package), or by visual inspection (see generally, Current Protocols in Molecular Biology, F. M. Ausubel et al., eds., Current Protocols, a joint venture between Greene Publishing Associates, Inc. and John Wiley & Sons, Inc., (1995 Supplement)). Additionally, determination of sequence alignment and percent sequence identity can employ the BESTFIT or GAP programs in the GCG Wisconsin Software package (Accelrys, Madison WI), using default parameters provided.

[0112] A nucleic acid or polynucleotide is “operably linked” when it is placed into a functional relationship with another nucleic acid sequence. For example, DNA for a presequence or secretory leader is operably linked to DNA for a polypeptide if it is expressed as a preprotein that participates in the secretion of the polypeptide; a promoter or enhancer is operably linked to a coding sequence if it affects the transcription of the sequence; or a ribosome binding site is operably linked to a coding sequence if it is positioned so as to facilitate translation. Generally, but not always, “operably linked” means that the DNA sequences being linked are contiguous, and, in the case of a secretory leader, contiguous and in reading phase. However, enhancers do not have to be contiguous. Linking is accomplished by ligation at convenient restriction sites. If such sites do not exist, the synthetic oligonucleotide adaptors or linkers are used in accordance with conventional practice.

[0113] As used herein, the expressions “cell,” “cell line,” and “cell culture” are used interchangeably and all such designations include progeny. Thus, the words “transformants” and “transformed cells” include the primary subject cell and cultures derived therefrom without regard for the number of transfers. It is also understood that not all progeny will have precisely identical DNA content, due to deliberate or inadvertent mutations. Mutant progeny that have the same function or biological activity as screened for in the originally transformed cell are included. Where distinct designations are intended, it will be clear from the context.

[0114] The term “solid tumor cancer” is defined as a cancer disease wherein the malignant tumor is present at a distinct location and does not contain a cyst and / or liquid area. Solid32NAI-5006826801V1tumors may be benign (not cancer), or malignant (cancer). Bladder cancer, breast cancer (e.g., TNBC), cervical cancer, colon and rectal cancer (colorectal cancer), endometrial cancer, head and neck cancer, kidney (renal cell) cancer, lung cancer, pancreatic cancer, prostate cancer, thyroid cancer, and urothelial cancer are non-limiting examples of different types of solid tumor cancers. Leukemia is an example of a cancer that is not a solid tumor cancer. See U.S. Patent No. 10,451,626.

[0115] The term “triple-negative breast cancer” as used herein refers to a breast cancer that tests negative for estrogen receptors, progesterone receptors, and HER2.

[0116] The term “monotherapy” refers to the use of a single agent or one type of treatment (e.g., radiation therapy) to treat a certain disease or conditions, e.g., a cancer.

[0117] The term “in association with” indicates that the components administered to a subject, e.g., an anti-Nectin-4 antibody, antigen-binding fragment thereof, or antibody conjugate, along with one or more additional therapeutic agents, can be formulated into a single composition for simultaneous delivery or formulated separately into two or more compositions (e.g., a kit). Each component can be administered to a subject at a different time than when the other component is administered; for example, each administration may be given non-simultaneously (e.g., separately or sequentially) at several intervals over a given period of time. Moreover, the separate components may be administered to a subject by the same or by a different route. The term “in association with” encompasses co-administration and co-formulation.

[0118] “ Co-administration" as used herein for therapeutic agents (e.g., an anti-Nectin-4 antibody, antigen-binding fragment thereof, or antibody conjugate, and one or more additional therapeutic agents) means that the agents are administered so as to have overlapping therapeutic activities, and not necessarily that the agents are administered simultaneously to the subject. The agents may or may not be in physical combination prior to administration. In an embodiment, the agents are administered to a subject simultaneously or at about the same time. For example, an anti-Nectin-4 antibody, antigen-binding fragment thereof, or antibody conjugate, and another therapeutic agent, may be contained in separate vials in liquid solution, and may be mixed into the same intravenous infusion bag or injection device and administered simultaneously to the patient. In other embodiments, the agents are administered sequentially. For example, e.g., an anti-Nectin-4 antibody, antigen-binding fragment thereof, or antibody conjugate may be administered prior to or following one or more additional therapeutic agents, e.g., another therapeutic antibody, antigen binding fragment thereof, antibody conjugate, and / or one or more chemotherapeutic agents.33NAI-5006826801V1

[0119] “ Co-formulated” or “co-formulation” as used herein refers to at least two agents (e.g., different antibodies, antigen binding fragments thereof, antibody conjugates, and / or therapeutic agents) thereof which are formulated together and stored as a combined product in a single vial or vessel (e.g., an injection device) rather than being formulated and stored individually and then mixed before administration or separately administered. In one embodiment, the co-formulation contains two different antibodies, antigen binding fragments thereof, and / or antibody conjugates described herein, for example, an anti-Nectin-4 antibody conjugate described herein (e.g., enfortumab vedotin) and a therapeutic described herein (e.g., a chemotherapeutic agent).

[0120] “Chemotherapeutic agent” is a chemical compound useful in the treatment of cancer. Classes of chemotherapeutic agents include, but are not limited to: platinum-containing chemotherapeutic agents (e.g., cisplatin, carboplatin, and oxaliplatin), alkylating agents, antimetabolites, kinase inhibitors, spindle poison plant alkaloids, cytotoxic / cytostatic agents, topoisomerase inhibitors, photosensitizers, anti-estrogens and selective estrogen receptor modulators (SERMs), anti-progesterones, estrogen receptor down-regulators (ERDs), estrogen receptor antagonists, luteinizing hormone-releasing hormone agonists, anti-androgens, aromatase inhibitors, EGFR inhibitors, VEGF inhibitors, anti-sense oligonucleotides that that inhibit expression of genes implicated in abnormal cell proliferation or tumor growth.

[0121] “Cytotoxic / cytostatic agents” refer to compounds which cause cell death or inhibit cell proliferation primarily by interfering directly with the cell’s functioning or inhibit or interfere with cell myosis, including alkylating agents, tumor necrosis factors, intercalators, hypoxia activatable compounds, microtubule inhibitors / microtubule-stabilizing agents, inhibitors of mitotic kinesins, histone deacetylase inhibitors, inhibitors of kinases involved in mitotic progression, inhibitors of kinases involved in growth factor and cytokine signal transduction pathways, antimetabolites, biological response modifiers, hormonal / anti-hormonal therapeutic agents, hematopoietic growth factors, monoclonal antibody targeted therapeutic agents, topoisomerase inhibitors, proteosome inhibitors, ubiquitin ligase inhibitors, and aurora kinase inhibitors.

[0122] Drug-to- Antibody Ratio (DAR) as used herein: (i) when referring to a composition of the antibody conjugates, is the average number of therapeutic agents D (or units of L-D) that are conjugated to the antibody or functional fragment thereof (A) in the antibody conjugate, and (ii) when referring to a single antibody conjugate, is the discrete integer of therapeutic agents D (or units of L-D) that are conjugated to the antibody or functional fragment thereof (A) in the antibody conjugate. Methods to determine DAR are well known to the skilled person and include methods using Reverse Phase Chromatography, or HPLC-MS.34NAI-5006826801V1

[0123] As used herein, “therapeutically effective amount” refers to an amount of a therapeutic agent (e.g., an anti-Nectin-4 antibody, antigen-binding fragment thereof, or antibody conjugate), which is effective as a monotherapy or in combination with one or more active ingredients upon single or multiple dose administration (e.g., an infusion treatment over a period of time, such as 30 minutes, or a bolus and / or maintenance doses) to a subject, in inhibiting, delaying or reducing the growth of a cancer (e.g., a cancer characterized by presence of a solid tumor expressing Nectin-4). “Therapeutically effective” is also intended to refer to an amount of the therapeutic agent that is suitable for therapeutic uses. In various embodiments, a therapeutically effective amount of an anti-Nectin-4 antibody, antigen-binding fragment thereof, or antibody conjugate, is the amount of the anti-Nectin-4 antibody, antigen-binding fragment thereof, or antibody conjugate that is needed to treat a solid tumor cancer in combination with one or more additional therapeutic agents, for example, one or more chemotherapeutic agents, or a combination thereof.

[0124] A therapeutically effective dose of any of the antibodies, antigen binding fragments thereof, or Antibody conjugates described herein, administered alone or in combination with one or more additional therapeutic agents to a cell, tissue, or subject, is effective to cause a measurable improvement in one or more symptoms of disease (e.g., cancer) or the progression of the disease. A therapeutically effective dose may be an amount of the antibody, fragment thereof, or antibody conjugate sufficient to result in at least partial amelioration of symptoms, e.g., tumor shrinkage or elimination, lack of tumor growth, or increased survival time. When applied to an individual active ingredient administered alone, an effective dose refers to that ingredient alone. When applied to a combination, a therapeutically effective dose may be combined with amounts of the active ingredients that result in the therapeutic effect, whether administered in combination, serially or simultaneously. An effective amount of a therapeutic may result in an improvement of a biomarker, measure, or parameter by at least 10%; usually by at least 20%; preferably at least about 30%; more preferably at least 40%, and most preferably by at least 50%. An effective amount can also result in an improvement in a subjective measure in cases where subjective measures are used to assess disease severity.

[0125] “Diagnostic anti-Nectin-4 monoclonal antibody” means a monoclonal antibody (mAb) which specifically binds to the mature form of the designated Nectin-4 that is expressed on the surface of certain mammalian cells.35NAI-5006826801V1Therapeutic Methods, Uses, and Compositions for Use

[0126] Provided herein are methods, compositions, and uses for treating patients, including human patients, in need of treatment with the antibodies, antigen-binding fragments thereof, and / or antibody conjugates disclosed herein.

[0127] In one aspect, provided herein is a method for treating a cancer in a patient, the method comprising administering to the patient a therapeutically effective amount of an anti-Nectin-4 antibody conjugate, wherein the antibody conjugate comprises an anti-Nectin-4 antibody, or antigen-binding fragment thereof. In specific embodiments, the anti-Nectin-4 antibody, or antigen-binding fragment thereof, is an anti-human Nectin-4 antibody, or antigen-binding fragment thereof. In specific embodiments, the anti-Nectin-4 antibody, or antigen-binding fragment thereof, of the disclosed methods is a composition of anti-Nectin-4 antibodies, or antigen-binding fragments thereof.

[0128] In one aspect, provided herein is a method for treating a cancer in a patient, the method comprising administering to the patient a therapeutically effective amount of an anti-Nectin-4 antibody conjugate, wherein the antibody conjugate comprises an anti-Nectin-4 antibody, or antigen-binding fragment thereof, and at least one therapeutic agent. In specific embodiments, the anti-Nectin-4 antibody, or antigen-binding fragment thereof, is an anti-human Nectin-4 antibody, or antigen-binding fragment thereof. In specific embodiments, the anti-Nectin-4 antibody, or antigen-binding fragment thereof, of the disclosed methods is a composition of anti-Nectin-4 antibodies, or antigen-binding fragments thereof.

[0129] In some embodiments of the method, the anti-Nectin-4 antibody, or antigen-binding fragment thereof, comprises:(a) the following six complementarity determining regions (CDRs) for heavy and light chains defined according to the Chothia numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 3;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 4;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 5; CDR-L1 comprising the sequence as set forth in SEQ ID NO: 6; CDR-L2 comprising the sequence as set forth in SEQ ID NO: 7; and CDR-L3 comprising the sequence as set forth in SEQ ID NO: 8; or(b) the following six CDRs for heavy and light chains defined according to the Abm numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 9;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 10;36NAI-5006826801V1CDR-H3 comprising the sequence as set forth in SEQ ID NO: 5;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 6;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 7; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 8; or(c) the following six CDRs for heavy and light chains defined according to the Kabat numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 11;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 12 or 63;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 5;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 6;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 7; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 8; or(d) the following six CDRs for heavy and light chains defined according to the IMGT numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 13;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 14;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 15;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 16;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 17; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 8.

[0130] In some embodiments of the method, the anti-Nectin-4 antibody, or antigen-binding fragment thereof, comprises:(a) the following six CDRs for heavy and light chains defined according to the Chothia numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 21 or 18;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 22;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 23;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 24;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 25; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 26; or(b) the following six CDRs for heavy and light chains defined according to the Abm numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 27 or 36;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 28;37NAI-5006826801V1CDR-H3 comprising the sequence as set forth in SEQ ID NO: 23;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 24;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 25; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 26; or(c) the following six CDRs for heavy and light chains defined according to the Kabat numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 29 or 44;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 30;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 23;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 24;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 25; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 26; or(d) the following six CDRs for heavy and light chains defined according to the IMGT numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 31 or 54;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 32;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 33;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 34;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 35; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 26.

[0131] In some embodiments of the method, the anti-Nectin-4 antibody, or antigen-binding fragment thereof, comprises:(a) the following six CDRs for heavy and light chains defined according to the Chothia numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 39;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 40;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 41;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 42;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 43; andCDR-L3 comprising SEQ ID NO: 8 or a variant thereof; or(b) the following six CDRs for heavy and light chains defined according to the Abm numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 45;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 46;38NAI-5006826801V1CDR-H3 comprising the sequence as set forth in SEQ ID NO: 41;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 42;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 43; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 8; or(c) the following six CDRs for heavy and light chains defined according to the Kabat numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 47;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 48 or 64;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 41;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 42;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 43; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 8; or(d) the following six CDRs for heavy and light chains defined according to the IMGT numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 49;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 50;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 51;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 52;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 53; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 8.

[0132] In some embodiments of the method, the anti-Nectin-4 antibody, or antigen-binding fragment thereof, comprises: a VH having the sequence as set forth in SEQ ID NO: 19 or SEQ ID NO: 57 and a VL having the sequence as set forth in SEQ ID NO: 20 or SEQ ID NO: 58; a VH having the sequence as set forth in SEQ ID NO: 1 or SEQ ID NO: 55 and a VL having the sequence as set forth in SEQ ID NO: 2 or SEQ ID NO: 56; or a VH having the sequence as set forth in SEQ ID NO: 37 or SEQ ID NO: 59 and a VL having the sequence as set forth in SEQ ID NO: 38 or SEQ ID NO: 60.

[0133] In some embodiments of the method, the anti-Nectin-4 antibody, or antigen-binding fragment thereof, comprises:(1) a VH having the sequence as set forth in SEQ ID NO: 57 and a VL having the sequence as set forth in SEQ ID NO: 58;(2) a VH having the sequence as set forth in SEQ ID NO: 1 and a VL having the sequence as set forth in SEQ ID NO: 2;39NAI-5006826801V1(3) a VH having the sequence as set forth in SEQ ID NO: 19 and a VL having the sequence as set forth in SEQ ID NO: 20;(4) a VH having the sequence as set forth in SEQ ID NO: 37 and a VL having the sequence as set forth in SEQ ID NO: 38;(5) a VH having the sequence as set forth in SEQ ID NO: 55 and a VL having the sequence as set forth in SEQ ID NO: 56; or(6) a VH having the sequence as set forth in SEQ ID NO: 59 and a VL having the sequence as set forth in SEQ ID NO: 60.

[0134] In some embodiments of the method, the anti-Nectin-4 antibody, or antigen-binding fragment thereof, further comprises a human immunoglobulin heavy chain constant region or a variant thereof, and a light chain constant region or a variant thereof, wherein the variant has a substitution, deletion, or addition of one or more amino acids compared to the wild-type sequence from which it is derived. In some embodiments of the method, the variant has a substitution, deletion, or addition of up to 5 amino acids compared to the wild-type sequence from which it was derived.

[0135] In some embodiments of the method, the anti-Nectin-4 antibody, or antigen-binding fragment thereof, further comprises a heavy chain constant region having the sequence as set forth in SEQ ID NO: 61 and a light chain constant region having the sequence as set forth in SEQ ID NO: 62.

[0136] In some embodiments of the method, the at least one therapeutic agent of the antibody conjugate is a cytotoxic agent.

[0137] In some embodiments of the method, the antibody conjugate has the structure shown in Formula (I),A-(L-D)y(i); wherein, A represents an antibody or antigen-binding fragment thereof described herein;L comprises an amino acid or a peptide consisting of 2-10 amino acids, or L is selectedfrom Vai, Cit, Phe, Lys, D-Val, Leu, Gly, Ala, Asn,40NAI-5006826801V1of position 1 and position 2 represents the position where L and A are connected, and the other represents the position where L and D are connected;D is selected from the group consisting of cytotoxic agents, therapeutic antibodies, radioisotopes, oligonucleotides and analogs thereof, biologically active peptides, protein toxins and enzymes; preferably, D is a molecule with anti-tumor biological activity;41NAI-5006826801V1y is an integer of 1 to 10.

[0138] In some embodiments of the method, the antibody conjugate is formed by connecting the (L-D) moiety of Formula (1) with a sulfhydryl group of A, for example, the sulfhydryl group of the side chain of a cysteine residue of A, through a thioether bond,Formula (1), wherein the antibody conjugate has the structure of Formula (2):42NAI-5006826801V1Formula (2), wherein y is an integer from 1 to 10, and A represents an antibody or antigen-binding fragment thereof described herein.

[0139] In some embodiments of the method, the molar ratio (DAR value) of therapeutic agent to the antibody, or antigen-binding fragment thereof, in the composition is a decimal or integer between 1 and 10. In some embodiments of the method, the DAR is a decimal or integer between 1 and 8.

[0140] In some embodiments of the method, the DAR value is 7.0, 7.03, 7.1, 7.12, 7.2, 7.3, 7.4, 7.5, 7.6, 7.7, 7.8, 7.9 or 8.0. In some embodiments of the method, the DAR value is 7.71.

[0141] In some embodiments of the method, the age of the patient is greater than or equal to 18 years of age.

[0142] Any suitable cancer may be treated using the methods, compositions, uses, and kits disclosed herein. In various embodiments, the anti-Nectin-4 antibody conjugate is used to treat a patient having a solid tumor and / or a cancer characterized by presence of a solid tumor. A solid tumor in various embodiments is an abnormal mass of tissue. Solid tumors may be benign, or malignant. Different types of solid tumors are named for the type of cells that form them. Examples of solid tumors are sarcomas, carcinomas, and lymphomas. Leukemias (cancers of the blood) generally do not form solid tumors. In various embodiments the solid tumor does not contain a cyst and / or a liquid area. Exemplary solid tumors include but are not limited to sarcomas and carcinomas such as fibrosarcoma, myxosarcoma, liposarcoma, chondrosarcoma, osteogenic sarcoma, chordoma, angiosarcoma, endotheliosarcoma, lymphangiosarcoma, lymphangioendotheliosarcoma, synovioma, mesothelioma, Ewing’s tumor, leiomyosarcoma, rhabdomyosarcoma, colon carcinoma, pancreatic cancer, breast cancer (e.g., triple-negative breast cancer (TNBC), urothelial carcinoma, head and neck squamous cell carcinoma, cervical cancer, endometrial carcinoma, lung cancer (e.g., non-small cell lung cancer (NSCLC) or small cell lung cancer (SCLC)), ovarian cancer, prostate cancer, squamous cell carcinoma, basal cell carcinoma, adenocarcinoma, sweat gland carcinoma, sebaceous gland carcinoma, papillary carcinoma, papillary adenocarcinomas, cystadenocarcinoma, medullary carcinoma,43NAI-5006826801V1bronchogenic carcinoma, renal cell carcinoma, hepatocellular carcinoma, bile duct carcinoma, choriocarcinoma, seminoma, embryonal carcinoma, Wilm’s tumor, cervical cancer, testicular tumor, bladder carcinoma, epithelial carcinoma, glioblastoma multiforme, astrocytoma, medulloblastoma, craniopharyngioma, ependymoma, pinealoma, hemangioblastoma, acoustic neuroma, oligodendroglioma, melanoma, neuroblastoma, and retinoblastoma.

[0143] In some embodiments of the method, the cancer is selected from the group consisting of urothelial carcinoma, head and neck squamous cell carcinoma, cervical cancer, endometrial carcinoma, and triple-negative breast cancer. In some embodiments of the method, the cancer is selected from the group consisting of cervical cancer and endometrial carcinoma.

[0144] In some embodiments of the method, the cancer is (i) locally recurrent unresectable and / or (ii) metastatic. In some embodiments of the method, the patient was previously treated for the cancer. In some embodiments of the method, the patient was previously treated with a chemotherapy, an anti-PD-l / anti-PD-Ll therapy, tisotumab vedotin, and / or enfortumab vedotin.

[0145] Any suitable dose or dosing regimen of the anti-Nectin-4 antibody, antigen binding fragment thereof, or antibody conjugate may be used. In some embodiments of the method, the anti-Nectin-4 antibody conjugate is administered at a dose of about 2 mg to about 700 mg. For example, the subject or patient is administered about 2 mg, about 7 mg, about 20 mg, about 30 mg, about 70 mg, about 200 mg, or about 700 mg. In various embodiments of the method, the patient is administered 2 mg of the anti-Nectin-4 antibody conjugate. In various embodiments of the method, the patient is administered 7 mg of the anti-Nectin-4 antibody conjugate. In various embodiments of the method, the patient is administered 20 mg of the anti-Nectin-4 antibody conjugate. In various embodiments of the method, the patient is administered 30 mg of the anti- Nectin-4 antibody conjugate. In various embodiments of the method, the patient is administered 70 mg of the anti-Nectin-4 antibody conjugate. In various embodiments of the method, the patient is administered 200 mg of the anti-Nectin-4 antibody conjugate. In various embodiments of the method, the patient is administered 700 mg of the anti-Nectin-4 antibody conjugate. In various embodiments of the method, the patient is administered a dose of about 200 mg to about 700 mg of the anti-Nectin-4 antibody conjugate. In various embodiments of the method, the patient is administered a dose of about 30 mg to about 700 mg of the anti-Nectin-4 antibody conjugate. In various embodiments of the method, the patient is administered a dose of about 30 mg, about 200 mg, or about 200 mg to about 700 mg of the anti-Nectin-4 antibody conjugate. In various embodiments of the method, the patient is administered a dose of about 30 mg to about 200 mg of the anti-Nectin-4 antibody conjugate. In various embodiments of the method, the44NAI-5006826801V1patient is administered a dose of about 70 mg to about 200 mg of the anti-Nectin-4 antibody conjugate.

[0146] In some embodiments of the method, the anti-Nectin-4 antibody conjugate is administered to the patient via intravenous infusion. In some embodiments of the method, the anti-Nectin-4 antibody conjugate is administered to the patient at a dose of about 1 mg / kg to about 6 mg / kg. In some embodiments of the method, the anti-Nectin-4 antibody conjugate is administered to the patient at a dose of about 1 mg / kg. In some embodiments of the method, the anti-Nectin-4 antibody conjugate is administered to the patient at a dose of about 2 mg / kg. In some embodiments of the method, the anti-Nectin-4 antibody conjugate is administered to the patient at a dose of about 3 mg / kg. In some embodiments of the method, the anti-Nectin-4 antibody conjugate is administered to the patient at a dose of about 4 mg / kg. In some embodiments of the method, the anti-Nectin-4 antibody conjugate is administered to the patient at a dose of about 5 mg / kg. In some embodiments of the method, the anti-Nectin-4 antibody conjugate is administered to the patient at a dose of about 6 mg / kg.

[0147] In some embodiments of the method, the anti-Nectin-4 antibody conjugate is administered to the patient every week (Q1W). In some embodiments of the method, the anti- Nectin-4 antibody conjugate is administered to the patient every two weeks (Q2W). In some embodiments of the method, the anti-Nectin-4 antibody conjugate is administered to the patient every three weeks (Q3W). In some embodiments of the method, the anti-Nectin-4 antibody conjugate is administered to the patient every four weeks (Q4W).

[0148] In another aspect, provided herein is a use of an anti-Nectin-4 antibody conjugate in the manufacture of a medicament for treating a cancer in a patient, wherein the antibody conjugate comprises an anti-Nectin-4 antibody, or antigen-binding fragment thereof, and at least one therapeutic.

[0149] In some embodiments of the use, the anti-Nectin-4 antibody, or antigen-binding fragment thereof, comprises:(a) the following six complementarity determining regions (CDRs) for heavy and light chains defined according to the Chothia numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 3; CDR-H2 comprising the sequence as set forth in SEQ ID NO: 4; CDR-H3 comprising the sequence as set forth in SEQ ID NO: 5; CDR-L1 comprising the sequence as set forth in SEQ ID NO: 6; CDR-L2 comprising the sequence as set forth in SEQ ID NO: 7; and CDR-L3 comprising the sequence as set forth in SEQ ID NO: 8; or45NAI-5006826801V1(b) the following six CDRs for heavy and light chains defined according to the Abm numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 9;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 10;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 5;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 6;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 7; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 8; or(c) the following six CDRs for heavy and light chains defined according to the Kabat numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 11;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 12 or 63;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 5;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 6;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 7; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 8; or(d) the following six CDRs for heavy and light chains defined according to the IMGT numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 13;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 14;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 15;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 16;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 17; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 8.

[0150] In some embodiments of the use, the anti-Nectin-4 antibody, or antigen-binding fragment thereof, comprises:(a) the following six CDRs for heavy and light chains defined according to the Chothia numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 21 or 18;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 22;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 23;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 24;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 25; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 26; or46NAI-5006826801V1(b) the following six CDRs for heavy and light chains defined according to the Abm numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 27 or 36;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 28;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 23;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 24;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 25; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 26; or(c) the following six CDRs for heavy and light chains defined according to the Kabat numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 29 or 44;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 30;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 23;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 24;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 25; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 26; or(d) the following six CDRs for heavy and light chains defined according to the IMGT numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 31 or 54;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 32;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 33;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 34;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 35; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 26.

[0151] In some embodiments of the use, the anti-Nectin-4 antibody, or antigen-binding fragment thereof, comprises:(a) the following six CDRs for heavy and light chains defined according to the Chothia numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 39;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 40;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 41;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 42;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 43; andCDR-L3 comprising SEQ ID NO: 8 or a variant thereof; or47NAI-5006826801V1(b) the following six CDRs for heavy and light chains defined according to the Abm numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 45;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 46;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 41;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 42;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 43; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 8; or(c) the following six CDRs for heavy and light chains defined according to the Kabat numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 47;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 48 or 64;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 41;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 42;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 43; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 8; or(d) the following six CDRs for heavy and light chains defined according to the IMGT numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 49;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 50;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 51;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 52;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 53; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 8.

[0152] In some embodiments of the use, the anti-Nectin-4 antibody, or antigen-binding fragment thereof, comprises: a VH having the sequence as set forth in SEQ ID NO: 19 or SEQ ID NO: 57 and a VL having the sequence as set forth in SEQ ID NO: 20 or SEQ ID NO: 58; a VH having the sequence as set forth in SEQ ID NO: 1 or SEQ ID NO: 55 and a VL having the sequence as set forth in SEQ ID NO: 2 or SEQ ID NO: 56; or a VH having the sequence as set forth in SEQ ID NO: 37 or SEQ ID NO: 59 and a VL having the sequence as set forth in SEQ ID NO: 38 or SEQ ID NO: 60.

[0153] In some embodiments of the use, the anti-Nectin-4 antibody, or antigen-binding fragment thereof, comprises:48NAI-5006826801V1(1) a VH having the sequence as set forth in SEQ ID NO: 57 and a VL having the sequence as set forth in SEQ ID NO: 58;(2) a VH having the sequence as set forth in SEQ ID NO: 1 and a VL having the sequence as set forth in SEQ ID NO: 2;(3) a VH having the sequence as set forth in SEQ ID NO: 19 and a VL having the sequence as set forth in SEQ ID NO: 20;(4) a VH having the sequence as set forth in SEQ ID NO: 37 and a VL having the sequence as set forth in SEQ ID NO: 38;(5) a VH having the sequence as set forth in SEQ ID NO: 55 and a VL having the sequence as set forth in SEQ ID NO: 56; or(6) a VH having the sequence as set forth in SEQ ID NO: 59 and a VL having the sequence as set forth in SEQ ID NO: 60.

[0154] In some embodiments of the use, the anti-Nectin-4 antibody, or antigen-binding fragment thereof, further comprises a human immunoglobulin heavy chain constant region or a variant thereof, and a light chain constant region or a variant thereof, wherein the variant has a substitution, deletion, or addition of one or more amino acids compared to the wild-type sequence from which it is derived. In some embodiments of the use, the variant has a substitution, deletion, or addition of up to 5 amino acids compared to the wild-type sequence from which it was derived. In some embodiments of the use, the anti-Nectin-4 antibody, or antigen-binding fragment thereof, further comprises a heavy chain constant region having the sequence as set forth in SEQ ID NO: 61 and a light chain constant region having the sequence as set forth in SEQ ID NO: 62.

[0155] In some embodiments of the use, the at least one therapeutic agent of the antibody conjugate is a cytotoxic agent.

[0156] In some embodiments of the use, the antibody conjugate has the structure shown in Formula (I),A-(L-D)y(i); wherein, A represents the antibody or antigen-binding fragment thereof as described herein;49NAI-5006826801V1L comprises an amino acid or a peptide consisting of 2-10 amino acids, or L is selectedfrom Vai, Cit, Phe, Lys, D-Val, Leu, Gly, Ala, Asn,of position 1 and position 2 represents the position where L and A are connected, and the other represents the position where L and D are connected;D is selected from the group consisting of cytotoxic agents, therapeutic antibodies, radioisotopes, oligonucleotides and analogs thereof, biologically active peptides, protein toxins and enzymes; preferably, D is a molecule with anti-tumor biological activity;50NAI-5006826801V1y is an integer of 1 to 10.

[0157] In some embodiments of the use, the antibody conjugate is formed by connecting the (L-D) moiety of Formula (1) with a sulfhydryl group of A, for example, the sulfhydryl group of the side chain of a cysteine residue of A, through a thioether bond,Formula (1), wherein the antibody conjugate has the structure of Formula (2):51NAI-5006826801V1Formula (2), wherein y is an integer from 1 to 10, and A represents the antibody or antigen-binding fragment thereof as described herein.

[0158] In some embodiments of the use, the molar ratio (DAR value) of therapeutic agent to the antibody, or antigen-binding fragment thereof, in the composition is a decimal or integer between 1 and 10. In some embodiments of the use, the DAR is a decimal or integer between 1 and 8.

[0159] In some embodiments of the use, the DAR value is 7.0, 7.03, 7.1, 7.12, 7.2, 7.3, 7.4, 7.5, 7.6, 7.7, 7.8, 7.9 or 8.0. In some embodiments of the use, the DAR value is 7.71.

[0160] In some embodiments of the use, the age of the patient is greater than or equal to 18 years of age.

[0161] Any suitable cancer may be treated using the methods, compositions, uses, and kits disclosed herein. In various embodiments, the anti-Nectin-4 antibody conjugate is used to treat a patient having a solid tumor and / or a cancer characterized by presence of a solid tumor. A solid tumor in various embodiments is an abnormal mass of tissue. Solid tumors may be benign, or malignant. Different types of solid tumors are named for the type of cells that form them. Examples of solid tumors are sarcomas, carcinomas, and lymphomas. Leukemias (cancers of the blood) generally do not form solid tumors. In various embodiments the solid tumor does not contain a cyst and / or a liquid area. Exemplary solid tumors include but are not limited to sarcomas and carcinomas such as fibrosarcoma, myxosarcoma, liposarcoma, chondrosarcoma, osteogenic sarcoma, chordoma, angiosarcoma, endotheliosarcoma, lymphangiosarcoma, lymphangioendotheliosarcoma, synovioma, mesothelioma, Ewing’s tumor, leiomyosarcoma, rhabdomyosarcoma, colon carcinoma, pancreatic cancer, breast cancer (e.g., triple-negative breast cancer (TNBC), urothelial carcinoma, head and neck squamous cell carcinoma, cervical cancer, endometrial carcinoma, lung cancer (e.g., non-small cell lung cancer (NSCLC) or small cell lung cancer (SCLC)), ovarian cancer, prostate cancer, squamous cell carcinoma, basal cell carcinoma, adenocarcinoma, sweat gland carcinoma, sebaceous gland carcinoma, papillary carcinoma, papillary adenocarcinomas, cystadenocarcinoma, medullary carcinoma,52NAI-5006826801V1bronchogenic carcinoma, renal cell carcinoma, hepatocellular carcinoma, bile duct carcinoma, choriocarcinoma, seminoma, embryonal carcinoma, Wilm’s tumor, cervical cancer, testicular tumor, bladder carcinoma, epithelial carcinoma, glioblastoma multiforme, astrocytoma, medulloblastoma, craniopharyngioma, ependymoma, pinealoma, hemangioblastoma, acoustic neuroma, oligodendroglioma, melanoma, neuroblastoma, and retinoblastoma.

[0162] In some embodiments of the use, the cancer is selected from the group consisting of urothelial carcinoma, head and neck squamous cell carcinoma, cervical cancer, endometrial carcinoma, and triple-negative breast cancer. In some embodiments of the use, the cancer is selected from the group consisting of cervical cancer and endometrial carcinoma.

[0163] In some embodiments of the use, the cancer is (i) locally recurrent unresectable and / or (ii) metastatic. In some embodiments of the use, the patient was previously treated for the cancer. In some embodiments of the use, the patient was previously treated with a chemotherapy, an anti-PD-l / anti-PD-Ll therapy, tisotumab vedotin, and / or enfortumab vedotin.

[0164] Any suitable dose or dosing regimen of the anti-Nectin-4 antibody conjugate may be used. In some embodiments of the use, the anti-Nectin-4 antibody conjugate is administered at a dose of about 2 mg to about 700 mg. For example, the subject or patient is administered about 2 mg, about 7 mg, about 20 mg, about 30 mg, about 70 mg, about 200 mg, or about 700 mg. In various embodiments of the use, the patient is administered 2 mg of the anti-Nectin-4 antibody conjugate. In various embodiments of the use, the patient is administered 7 mg of the anti- Nectin-4 antibody conjugate. In various embodiments of the use, the patient is administered 20 mg of the anti-Nectin-4 antibody conjugate. In various embodiments of the use, the patient is administered 30 mg of the anti-Nectin-4 antibody conjugate. In various embodiments of the use, the patient is administered 70 mg of the anti-Nectin-4 antibody conjugate. In various embodiments of the use, the patient is administered 200 mg of the anti-Nectin-4 antibody conjugate. In various embodiments of the use, the patient is administered 700 mg of the anti- Nectin-4 antibody conjugate. In various embodiments of the use, the patient is administered a dose of about 200 mg to about 700 mg of the anti-Nectin-4 antibody conjugate. In various embodiments of the use, the patient is administered a dose of about 30 mg to about 700 mg of the anti-Nectin-4 antibody conjugate. In various embodiments of the use, the patient is administered a dose of about 30 mg, about 200 mg, or about 200 mg to about 700 mg of the anti-Nectin-4 antibody conjugate. In various embodiments of the use, the patient is administered a dose of about 30 mg to about 200 mg of the anti-Nectin-4 antibody conjugate. In various embodiments of the use, the patient is administered a dose of about 70 mg to about 200 mg of the anti-Nectin-4 antibody conjugate.53NAI-5006826801V1

[0165] In some embodiments of the use, the anti-Nectin-4 antibody conjugate is administered to the patient via intravenous infusion. In some embodiments of the use, the anti-Nectin-4 antibody conjugate is administered to the patient at a dose of about 1 mg / kg to about 6 mg / kg. In some embodiments of the use, the anti-Nectin-4 antibody conjugate is administered to the patient at a dose of about 1 mg / kg. In some embodiments of the use, the anti-Nectin-4 antibody conjugate is administered to the patient at a dose of about 2 mg / kg. In some embodiments of the use, the anti-Nectin-4 antibody conjugate is administered to the patient at a dose of about 3 mg / kg. In some embodiments of the use, the anti-Nectin-4 antibody conjugate is administered to the patient at a dose of about 4 mg / kg. In some embodiments of the use, the anti-Nectin-4 antibody conjugate is administered to the patient at a dose of about 5 mg / kg. In some embodiments of the use, the anti-Nectin-4 antibody conjugate is administered to the patient at a dose of about 6 mg / kg.

[0166] In some embodiments of the use, the anti-Nectin-4 antibody conjugate is administered to the patient every week (Q1W). In some embodiments of the use, the anti-Nectin-4 antibody conjugate is administered to the patient every two weeks (Q2W). In some embodiments of the use, the anti-Nectin-4 antibody conjugate is administered to the patient every three weeks (Q3W). In some embodiments of the use, the anti-Nectin-4 antibody conjugate is administered to the patient every four weeks (Q4W).

[0167] In some embodiments of the method or use, the antibody conjugate or medicament is administered at least twice.Additional Therapeutic Agents

[0168] In particular embodiments, the antibodies, antigen-binding fragments thereof, or antibody conjugates disclosed herein may be used alone, or in association with other, additional therapeutic agents and / or therapeutic procedures, for treating or preventing any disease such as cancer, e.g., as discussed herein, in a subject in need of such treatment or prevention. Compositions, e.g., pharmaceutical compositions comprising a pharmaceutically acceptable carrier, comprising such antibodies and fragments thereof in association with further therapeutic agents are also part of the present disclosure.

[0169] Therefore, the present disclosure provides a method or use of treating cancer in a subject (e.g., a human subject), comprising administering to the subject an effective amount of an antibody, antigen-binding fragment thereof, or antibody conjugate (e.g., an anti-Nectin-4 antibody, antigen-binding fragment thereof, or antibody conjugates) disclosed herein, optionally in association with an additional therapeutic agent or therapeutic procedure. The present disclosure also provides a method of treating cancer in a human subject, comprising54NAI-5006826801V1administering to the subject an effective amount of an antibody, antigen-binding fragment thereof, or antibody conjugate disclosed herein, optionally in association with an additional therapeutic agent or therapeutic procedure. The present disclosure also provides a method of increasing the activity of an immune cell, comprising administering to a subject in need thereof an effective amount of an antibody, antigen-binding fragment thereof, or antibody conjugate disclosed herein. In another embodiment, the present disclosure provides an antibody or antigen binding fragment thereof of the invention, for use in treatment of cancer as a monotherapy; increasing the activity of an immune cell; or treatment of cancer in combination with one or more additional therapeutic agents. In a further embodiment, the present disclosure provides use of the antibody or antigen binding fragment thereof of the invention for the manufacture of a medicament for increasing immune cell activation; treating cancer as a monotherapy; or treating cancer in combination with one or more additional therapeutic agents. In another embodiment, the present disclosure provides a combination of an antibody or antigen binding fragment thereof of the invention and a further therapeutic agent for the treatment of cancer; increasing the activity of an immune cell; or treatment of another disorder.

[0170] In other embodiments, the disclosure provides a method of treating cancer in a human subject, comprising administering to the subject an effective amount of an antibody or antigen binding fragment thereof disclosed herein, or an expression vector or a host cell according to the disclosure, optionally in association with one or more additional therapeutic agents and / or therapeutic procedures.

[0171] In some embodiments, the antibodies, antigen-binding fragments thereof, or antibody conjugates disclosed herein (e.g., an anti-Necti-4 antibody, antigen-binding fragment thereof, or antibody conjugate) may be used in association with a tumor vaccine. Examples of tumor vaccines include but are not limited to vaccines for Human Papillomavirus (HPV) infection- caused cancer such as Gardasil®, Gardasil® and Cervarix®; vaccines that prevent hepatitis B virus-caused liver cancer such as Engerix-B® and Recombivax HB®; oncolytic virus therapy that triggers immune response such as Imlygic®; DNA vaccines such as Synchotrope MA2M plasmid DNA vaccine and ZYC101; mammaglobin-a DNA vaccine (see Clinical Cancer Res. 2014 20(23):5964-75); vector based vaccines such as PSA-TRICOM (prostvac), PANVAC-VF, Listeria monocytogenes-based PSA vaccine (see Therapeutic Advances in Vaccines, 2014, 2(5) 137- 148), ZzVerzri-mesothelin Adeno-CEA; allogeneic vaccines such as GV AX, BLP-25 (anti- Ankara-mucin 1), Belagenpumatucel-L, TG4010, CIMAvax epidermal growth factor vaccine, NY-ESO, GM.CD40L-CCL21; autologous vaccines such as:Adeno-CD40L, BCG, INGN-225, dendritic cell vaccines such as Provenge® (Sipuleucel-T), rF-CEA-MUCl -TRICOM (panvac-55NAI-5006826801V1DC); antigen vaccines such as MUC-1 (stimuvax), NY-ESO-1, GP-100, MAGE- A3 (melanoma antigen encoding gene A3), or INGN-225 (see Pharmacology & Therapeutics 153 (2015) 1-9).

[0172] In particular embodiments, the anti-Nectin-4 antibodies, antigen-binding fragments thereof, or antibody conjugates disclosed herein may be used in association with radiation therapy.

[0173] In particular embodiments, the anti-Nectin-4 antibodies, antigen-binding fragments thereof, or antibody conjugates disclosed herein may be used in association with targeted therapies. Examples of targeted therapies include: hormone therapies, signal transduction inhibitors (e.g., EGFR inhibitors, such as cetuximab (Erbitux®) and erlotinib (Tarceva®)); HER2 inhibitors (e.g., trastuzumab (Herceptin®) and pertuzumab (Perjeta®)); BCR-ABL inhibitors (such as imatinib (Gleevec®) and dasatinib (Sprycel®)); ALK inhibitors (such as crizotinib (Xalkori®) and ceritinib (Zykadia®)); BRAF inhibitors (such as vemurafenib (Zelboraf®) and dabrafenib (Tafinlar®)), gene expression modulators, apoptosis inducers (e.g., bortezomib (Velcade®) and carfilzomib (Kyprolis®)), angiogenesis inhibitors, e.g., bevacizumab (Avastin®) and ramucirumab (Cyramza®), monoclonal antibodies attached to toxins (e.g., brentuximab vedotin (Adcetris®) and ado-trastuzumab emtansine (Kadcyla®)).

[0174] In particular embodiments, the anti-Nectin-4 antibodies, antigen-binding fragments thereof, or antibody conjugates may be used in association with an anti-cancer therapeutic agent or immunomodulatory drug such as an immunomodulatory receptor inhibitor, e.g., an antibody, antigen-binding fragment thereof, or antibody conjugate that specifically binds to the receptor.

[0175] In an embodiment, an anti-Nectin-4 antibody, antigen-binding fragment thereof, or antibody conjugate of the disclosure is used in association with one or more of: an anti-PD-1 antibody (e.g., pembrolizumab (KEYTRUDA®), nivolumab, pidilizumab (CT-011)), an anti- PD-L1 antibody (e.g., BMS-936559, Durvalumab, MSB0010718C or MPDL3280A), an anti- PD-L2 antibody, anti-TIGIT antibody, an anti-CTLA4 antibody, an anti-CSl antibody (e.g., elotuzumab), an anti-KIR2DLl / 2 / 3 antibody (e.g., lirilumab), an anti-CD137 antibody (e.g., urelumab), an anti-GITR antibody (e.g., TRX518), an anti-ILTl antibody, an anti-ILT2 antibody, an anti-ILT3 antibody, an anti-ILT4 antibody, an anti-ILT5 antibody, an anti-ILT6 antibody, an anti-ILT7 antibody, an anti-ILT8 antibody, an anti-CD40 antibody, an anti-OX40 antibody, an anti-ICOS, an anti-SIRPa, an anti-KIR2DLl antibody, an anti-KIR2DL2 / 3 antibody, an anti-KIR2DL4 antibody, an anti-KIR2DL5A antibody, an anti-KIR2DL5B antibody, an anti-KIR3DLl antibody, an anti-KIR3DL2 antibody, an anti-KIR3DL3 antibody, an anti-NKG2A antibody, an anti-NKG2C antibody, an anti-NKG2E antibody, an anti -4- IBB56NAI-5006826801V1antibody (e.g., PF-05082566), an anti-TSLP antibody, an anti-IL-10 antibody, IL-10 or PEGylated IL-10, or any small organic molecule inhibitor of such targets.

[0176] In an embodiment, an anti-Nectin-4 antibody, antigen-binding fragment thereof, or antibody conjugate of the disclosure is used in association with one or more of an inhibitor (e.g., a small organic molecule or an antibody or antigen-binding fragment thereof) such as: an MTOR (mammalian target of rapamycin) inhibitor, a cytotoxic agent, a platinum agent, an EGFR inhibitor, a VEGF inhibitor, a microtubule stabilizer, a taxane, a CD20 inhibitor, a CD52 inhibitor, a CD30 inhibitor, a RANK (Receptor activator of nuclear factor kappa-B) inhibitor, a STING agonist, a CXCR2 antagonist, a RANKL (Receptor activator of nuclear factor kappa-B ligand) inhibitor, an ERK inhibitor, a MAP Kinase inhibitor, an AKT inhibitor, a MEK inhibitor, a PARP inhibitor, a PI3K inhibitor, a HER1 inhibitor, a HER2 inhibitor, a HER3 inhibitor, a HER4 inhibitor, a Bcl2 inhibitor, a CD22 inhibitor, a CD79b inhibitor, an ErbB2 inhibitor, or a farnesyl protein transferase inhibitor.

[0177] In some examples, the anti-Nectin-4 antibody or antigen binding fragment thereof is co-administered with one or more chemotherapeutic agents. Any suitable chemotherapeutic agent, including any chemotherapeutic agent or combination thereof disclosed herein, may be used. In some examples, the one or more chemotherapeutic agents comprise nab-paclitaxel. In some examples, the nab-paclitaxel is administered to the patient via intravenous infusion. In some examples, the nab-paclitaxel is administered to the patient at a dose of about 100 mg / m2In some examples, the nab-paclitaxel is administered to the patient on a 28-day cycle, and the nab-paclitaxel is administered to the patient on Day 1, 8, and 15 of the 28-day cycle. In some examples, the nab-paclitaxel is administered to the patient on a 28-day cycle, and the nab- paclitaxel is administered to the patient at a dose of about 100 mg / m2on Day 1, 8, and 15 of the 28-day cycle.

[0178] In an embodiment, an anti-Nectin-4 antibody, antigen-binding fragment thereof, or antibody conjugate of the disclosure is used in association with any one or more of: 13-cis- retinoic acid, 3-[5- (methyl sulfonylpiperadinemethylj-indolyl] -quinol one, 4-hydroxytamoxifen, 5- deooxyuridine, 5'-deoxy-5-fluorouridine, 5 -fluorouracil, 6-mecaptopurine, 7- hydroxystaurosporine, A-443654, abirateroneacetate, ABT-578, acolbifene, ADS- 100380, ALT-110, altretamine, amifostine, aminoglutethimide, amrubicin, Amsacrine, anagrelide, anastrozole, angiostatin, AP-23573, ARQ-197, arzoxifene, AS-252424, AS- 605240, asparaginase, AT-9263, atrasentan, axitinib, AZDI 152, Bacillus Calmette-Guerin (BCG) vaccine, batabulin, BC-210, besodutox, bevacizumab, bicalutamide, Biol 11, BIO140, bleomycin, BMS-214662, BMS-247550, BMS-275291, BMS-310705, bortezimib, buserelin,57NAI-5006826801V1busulfan, calcitriol, camptothecin, canertinib, capecitabine, carboplatin, carmustine, CC8490, cediranib, CG-1521, CG-781, chlamydocin, chlorambucil, chlorotoxin, cilengitide, cimitidine, cisplatin, cladribine, clodronate, COL-3, CP-724714, cyclophosphamide, cyproterone, cyproteroneacetate, cytarabine, cytosinearabinoside, dacarbazine, dacinostat, dactinomycin, dalotuzumab, danusertib, dasatanib, daunorubicin, decatanib, deguelin, denileukin, deoxycoformycin, depsipeptide, diarylpropionitrile, diethylstilbestrol, diftitox, docetaxel, dovitinib, doxorubicin, droloxifene, edotecarin, yttrium-90 labeled-edotreotide, edotreotide, EKB-569, EMD121974, endostatin, enzalutamide, enzastaurin, epirubicin, epithilone B, ERA- 923, cetuximab (Erbitux®), erlotinib, estradiol, estramustine, etoposide, everolimus, exemestane, ficlatuzumab, finasteride, flavopiridol, floxuridine, fludarabine, fludrocortisone, fluoxymesterone, flutamide, FOLFOX regimen, fulvestrant, galeterone, gefitinib, gemcitabine, gimatecan, goserelin, goserelin acetate, gossypol, GSK461364, GSK690693, HMR-3339, hydroxyprogesteronecaproate, hydroxyurea, IC87114, idarubicin, idoxyfene, ifosfamide, IM862, imatinib, IMC-1C11, INCB24360, INOlOOl, interferon, interleukin- 12, ipilimumab, irinotecan, JNJ-16241199, ketoconazole, KRX-0402, lapatinib, lasofoxifene, letrozole, leucovorin, leuprolide, leuprolide acetate, levamisole, liposome entrapped paclitaxel, lomustine, lonafarnib, lucanthone, LY292223, LY292696, LY293646, LY293684, LY294002, LY317615, marimastat, mechlorethamine, medroxyprogesteroneacetate, megestrolacetate, melphalan, mercaptopurine, mesna, methotrexate, mithramycin, mitomycin, mitotane, mitoxantrone, tozasertib, MLN8054, neovastat, Neratinib, neuradiab, nilotinib, nilutimide, nolatrexed, NVP- BEZ235, oblimersen, octreotide, ofatumumab, olaparib, oregovomab, orteronel, oxaliplatin, paclitaxel, palbociclib, pamidronate, panitumumab, pazopanib, PD0325901, PD 184352, PEG- interferon, pemetrexed, pentostatin, perifosine, phenylalanine mustard, PI-103, pictilisib, PIK- 75, pipendoxifene, PKL166, plicamycin, porfimer, prednisone, procarbazine, a progestin, PX- 866, R-763, raloxifene, raltitrexed, razoxin, ridaforolimus, rituximab, romidepsin, RTA744, rubitecan, scriptaid, Sdxl02, seliciclib, selumetinib, semaxanib, SF1126, sirolimus, SN36093, sorafenib, spironolactone, squalamine, SR13668, streptozocin, SU6668, suberoylanalide hydroxamic acid, sunitinib, synthetic estrogen, talampanel, talimogene laherparepvec, tamoxifen, temozolomide, temsirolimus, teniposide, tesmilifene, testosterone, tetrandrine, TGX- 221, thalidomide, thioguanine, thiotepa, ticilimumab, tipifarnib, tivozanib, TKI-258, TLK286, topotecan, toremifene citrate, trabectedin, trastuzumab, tretinoin, trichostatin A, triciribinephosphate monohydrate, triptorelin pamoate, TSE-424, uracil mustard, valproic acid, valrubicin, vandetanib, vatalanib, VEGF trap, vinblastine, vincristine, vindesine, vinorelbine,58NAI-5006826801V1vitaxin, vitespan, vorinostat, VX-745, wortmannin, Xr311, zanolimumab, ZK186619, ZK- 304709, ZM336372, or ZSTK474.

[0179] Additional non-limiting examples of suitable anti-cancer agents that may be used in combination with an anti-Nectin-4 antibody, antigen-binding fragment thereof, or antibody conjugate include cytostatic agents, cytotoxic agents, targeted therapeutic agents (e.g., small molecules, biologies, siRNA and microRNA) against cancer and neoplastic diseases, antimetabolites (such as methotrexate, 5-fluorouracil, gemcitabine, fludarabine, capecitabine); alkylating agents, such as temozolomide, cyclophosphamide; DNA interactive and DNA damaging agents, such as cisplatin, oxaliplatin, and doxorubicin; Ionizing irradiation, such as radiation therapy; topoisomerase II inhibitors, such as etoposide and doxorubicin; topoisomerase I inhibitors, such as irinotecan and topotecan; tubulin interacting agents, such as paclitaxel, docetaxel, and nab-paclitaxel (Abraxane®), epothilones; kinesin spindle protein inhibitors; spindle checkpoint inhibitors; poly(ADP-ribose) polymerase (PARP) inhibitors, such as olaparib, niraparib and veliparib; matrix metalloprotease (MMP) inhibitors; protease inhibitors, such as cathepsin D and cathepsin K inhibitors; proteosome or ubiquitination inhibitors, such as bortezomib; adenoviral-p53; Bcl-2 inhibitors, such as ABT-263; heat shock protein (HSP) modulators, such as geldanamycin and 17-AAG; histone deacetylase (HD AC) inhibitors, such as vorinostat (SAHA); sex hormone modulating agents, such as anti-estrogens (e.g., tamoxifen and fulvestrant), selective estrogen receptor modulators (SERM) (e.g., raloxifene), antiandrogens (e.g., bicalutamide, flutamide), and luteinizing hormone-releasing hormone (LHRH) agonists (e.g., leuprolide); 5a-reductase inhibitors, such as finasteride; cytochrome P450 C17 lyase (CYP450cl7, also called 17aC); aromatase inhibitors, such as letrozole, anastrozole, and exemestane; EGFR kinase inhibitors, such as gefitinib, erlotinib, and lapatinib; multi-targeted kinases (serine / threonine and / or tyrosine kinase) inhibitors; ABL kinase inhibitors, such as imatinib, nilotinib, and dasatinib; VEGFR-1, VEGFR-2, PDGFR, KDR, FLT, c-Kit, Tie2, Raf, MEK or ERK inhibitors, such as sunitinib, sorafenib, vandetanib, pazopanib, PLX-4032, axitinib, PTK787, or GSK-1120212; Polo-like kinase inhibitors; Aurora kinase inhibitors; JAK inhibitors; c-MET kinase inhibitors; PI3K and mTOR inhibitors, such as GDC-0941, BEZ-235, BKM-120 and AZD-8055; rapamycin and its analogs, such as temsirolimus, everolimus, and deforolimus; a STING (Stimulator of Interferon Genes) agonist; a CXCR (CXC Chemokine Receptor) inhibitor; a CXCR2 antagonist, other anti-cancer (also known as anti -neoplastic) agents including but are not limited to ara-C, adriamycin, cytoxan, carboplatin, uracil mustard, clormethine, ifosfsmide, melphalan, chlorambucil, pipobroman, triethylenemelamine, triethylenethiophosphoramine, busulfan, carmustine, lomustine, streptozocin, dacarbazine,59NAI-5006826801V1floxuridine, cytarabine, 6-mercaptopurine, 6-thioguanine, fludarabine phosphate, pentostatine, vinblastine, vincristine, vindesine, vinorelbine, navelbine, bleomycin, dactinomycin, daunorubicin, doxorubicin, epirubicin, teniposide, cytarabine, pemetrexed, idarubicin, mithramycin, deoxycoformycin, mitomycin-C, L-asparaginase, teniposide, ethinylestradiol, diethylstilbestrol, testosterone, prednisone, fluoxymesterone, dromostanolone propionate, testolactone, megestrolacetate, methylprednisolone, methyltestosterone, prednisolone, triamcinolone, chi orotriani sene, hydroxyprogesterone, aminoglutethimide, estramustine, flutamide medroxyprogesteroneacetate, toremifene, goserelin, hydroxyurea, amsacrine, procarbazine, mitotane, mitoxantrone, levamisole, drolloxafine, hexamethylmelamine, tositumomab (Bexxar®), ibritumomab tiuxetan, arsenic trioxide, porfimer sodium, thiotepa, altretamine, liposomal doxorubicin, denileukin diftitox (Ontak®), cytarabine liposome injection (Depocyt®), darbepoetin alfa (Aranesp®), filgrastim (Neupogen®), pegfilgrastim (Neulasta®), or palifermin (Kepivance); famesyl protein transferase inhibitors, such as SARASAR™, tipifarnib; interferons, such as Intron A, Peg-Intron; anti-ErbBl antibodies, such as cetuximab, panitumumab; anti-ErbB2 antibodies, such as trastuzumab; anti-CD52 antibodies, such as alemtuzumab, anti-CD20 antibodies, such as rituximab; anti-CD33 antibodies, such as gemtuzumab ozogamicin; anti-VEGF antibodies, such as bevacizumab (Avastin®), TRAIL ligands, such as lexatumumab, mapatumumab, and AMG-655; anti-CTLA-4 antibodies, such as ipilimumab; antibodies against CTA1, CEA, CD5, CD19, CD22, CD30, CD44, CD44V6, CD55, CD56, EpCAM, FAP, MHCII, HGF, IL-6, MUC1, PSMA, TAL6, TAG-72, TRAILR, VEGFR, IGF-2, FGF; or anti-IGF-lR antibodies, such as dalotuzumab and robatumumab (SCH 717454).

[0180] Examples of cytotoxic / cytostatic agents that may be used in association with an anti- Nectin-4 antibody or an antigen-binding fragment thereof include, but are not limited to, platinum coordinator compounds, sertenef, cachectin, ifosfamide, tasonermin, lonidamine, carboplatin, altretamine, prednimustine, dibromodulcitol, ranimustine, fotemustine, nedaplatin, oxaliplatin, temozolomide, heptaplatin, estramustine, improsulfan tosilate, trofosfamide, nimustine, dibrospidium chloride, pumitepa, lobaplatin, satraplatin, profiromycin, cisplatin, irofulven, dexifosfamide, cis-aminedichloro(2-methyl-pyridine)platinum, benzylguanine, glufosfamide, GPX100, (trans, trans, trans)-bis-mu-(hexane-l,6-diamine)-mu-[diamine- platinum(II)]bis[diamine(chloro)platinum (II)]tetrachloride, diarizidinylspermine, arsenic trioxide, 1-(1 l-dodecylamino-10-hydroxyundecyl)-3,7-dimethylxanthine, zorubicin, idarubicin, daunorubicin, bisantrene, mitoxantrone, pirarubicin, pinafide, valrubicin, amrubicin, antineoplaston, 3 ’ -deamino-3 ’ -morpholino- 13 -deoxo- 10-hy droxy carminomy cin, annamy cin,60NAI-5006826801V1galarubicin, elinafide, MEN10755, 4-demethoxy-3-deamino-3-aziridinyl-4-methylsulphonyl- daunorubicin (see international patent publication no. W02000 / 050032).

[0181] An example of a hypoxia activatable compound that may be used in association with an anti-Nectin-4 antibody or an antigen-binding fragment thereof include, but are not limited to, is tirapazamine.

[0182] Examples of proteosome inhibitors that may be used in association with an anti-Nectin- 4 antibody or an antigen-binding fragment thereof include, but are not limited to, include but are not limited to lactacystin and bortezomib (Velcade®).

[0183] Examples of microtubule inhibitors / microtubule-stabilizing agents that may be used in association with an anti-Nectin-4 antibody or an antigen-binding fragment thereof include taxanes in general. Specific compounds include paclitaxel (Taxol®), nab-paclitaxel (Abraxane®), vindesine sulfate, 3’,4’-didehydro-4’-deoxy-8’-norvincaleukoblastine, docetaxel (Taxotere®), rhizoxin, dolastatin, mivobulin isethionate, auristatin, cemadotin, RPR109881, BMS 184476, vinflunine, cryptophycin, 2,3,4,5,6-pentafluoro-N-(3-fluoro-4-methoxyphenyl) benzene sulfonamide, anhydrovinblastine, N,N-dimethyl-L-valyl-L-valyl-N-methyl-L-valyl-L- prolyl-L-proline-t-butylamide, TDX258, the epothilones (see for example U.S. Pat. Nos. 6,284,781 and 6,288,237) and BMS188797.

[0184] Some examples of topoisomerase inhibitors that may be used in association with an anti-Nectin-4 antibody or an antigen-binding fragment thereof include, but are not limited to, are topotecan, hycaptamine, irinotecan, rubitecan, 6-ethoxypropionyl-3’,4’-O-exo-benzylidene- chartreusin, 9-methoxy-N,N-dimethyl-5-nitropyrazolo[3,4,5-kl]acridine-2-(6H) propanamine, 1- amino-9-ethyl-5-fluoro-2,3-dihydro-9-hydroxy-4-methyl-lH,12H-benzo[de]pyrano[3’,4’ :b,7]- indolizino[l,2b]quinoline-10,13(9H,15H)dione, lurtotecan, 7-[2-(N-isopropylamino)ethyl]- (20S)camptothecin, BNP1350, BNPI1100, BN80915, BN80942, etoposide phosphate, teniposide, sobuzoxane, 2’-dimethylamino-2’-deoxy-etoposide, GL331, N-[2- (dimethylamino)ethyl]-9-hydroxy-5,6-dimethyl-6H-pyrido[4,3-b]carbazole-l-carboxamide, asulacrine, (5a, 5aB, 8aa,9b)-9-[2-[N-[2-(dimethylamino)ethyl]-N-methylamino]ethyl]-5-[4- hydro0xy-3,5-dimethoxyphenyl]-5,5a,6,8,8a,9-hexohydrofuro(3’,4’:6,7)naphtho(2,3-d)-l,3- dioxol-6-one, 2,3-(methylenedioxy)-5-methyl-7-hydroxy-8-methoxybenzo[c]- phenanthridinium, 6,9-bis[(2-aminoethyl)amino]benzo[g]isoguinoline-5, 10-dione, 5-(3- aminopropylamino)-7,10-dihydroxy-2-(2-hydroxyethylaminomethyl)-6H-pyrazolo[4,5,l- de]acridin-6-one, N-[l-[2(diethylamino)ethylamino]-7-methoxy-9-oxo-9H-thioxanthen-4- ylmethyl]formamide, N-(2-(dimethylamino)ethyl)acridine-4-carboxamide, 6-[[2- (dimethylamino)ethyl]amino]-3-hydroxy-7H-indeno[2,l-c] quinolin-7-one, and dimesna.61NAI-5006826801V1

[0185] Examples of inhibitors of mitotic kinesins, and in particular the human mitotic kinesin KSP, that may be used in association with an anti-Nectin-4 antibody or an antigen-binding fragment thereof, are described in W003 / 039460, W003 / 050064, W003 / 050122, WO03 / 049527, WO03 / 049679, WO03 / 049678, WO04 / 039774, WO03 / 079973, W003 / 099211, W003 / 105855, W003 / 106417, W004 / 037171, W004 / 058148, W004 / 058700, WO04 / 126699, W005 / 018638, WO05 / 019206, WO05 / 019205, WO05 / 018547, W005 / 017190, US2005 / 0176776.

[0186] Examples of histone deacetylase inhibitors that may be used in association with an anti- Nectin-4 antibody or an antigen-binding fragment thereof include, but are not limited to, SAHA, TSA, oxamflatin, PXD101, MG98 and scriptaid. Further reference to other histone deacetylase inhibitors may be found in Miller, T.A. et al. J. Med. Chem. 46(24):5097-5116 (2003).

[0187] Examples of inhibitors of kinases involved in mitotic progression that may be used in association with an anti-Nectin-4 antibody or an antigen-binding fragment thereof include, but are not limited to, inhibitors of aurora kinase, inhibitors of Polo-like kinases (PLK; in particular inhibitors of PLK-1), inhibitors of Bub-1 and inhibitors of Bub-Rl. An example of an aurora kinase inhibitor is VX-680.

[0188] Antiproliferative agents that may be used in association with an anti-Nectin-4 antibody or an antigen-binding fragment thereof includes antisense RNA and DNA oligonucleotides such as G3139, ODN698, RVASKRAS, GEM231, and INX3001, and antimetabolites such as enocitabine, carmofur, tegafur, pentostatin, doxifluridine, trimetrexate, fludarabine, capecitabine, galocitabine, cytarabine ocfosfate, fosteabine sodium hydrate, raltitrexed, paltitrexid, emitefur, tiazofurin, decitabine, nolatrexed, pemetrexed, nelzarabine, 2’-deoxy-2’- methylidenecytidine, 2’ -fluoromethylene-2’ -deoxy cytidine, N-[5-(2,3-dihydro- benzofuryl)sulfonyl]-N’-(3,4-dichlorophenyl)urea, N6-[4-deoxy-4-[N2-[2(E),4(E)- tetradecadienoyl]glycylamino]-L-glycero-B-L-manno-heptopyranosyl]adenine, aplidine, ecteinascidin, troxacitabine, 4-[2-amino-4-oxo-4,6,7,8-tetrahydro-3H-pyrimidino[5,4- b][l,4]thiazin-6-yl-(S)-ethyl]-2,5-thienoyl-L-glutamic acid, aminopterin, 5-flurouracil, alanosine, 1 l-acetyl-8-(carbamoyloxymethyl)-4-formyl-6-methoxy-14-oxa-l,l 1- diazatetracyclo(7.4.1.0.0)-tetradeca-2,4,6-trien-9-yl acetic acid ester, swainsonine, lometrexol, dexrazoxane, methioninase, 2’-cyano-2’-deoxy-N4-palmitoyl-l-B-D-arabino furanosyl cytosine, 3-aminopyridine-2-carboxaldehyde thiosemicarbazone and trastuzumab.

[0189] Examples of angiogenesis inhibitors that may be used in association with an anti- Nectin-4 antibody or an antigen-binding fragment thereof include, but are not limited to, tyrosine kinase inhibitors, such as inhibitors of the tyrosine kinase receptors Fit- 1 (VEGFR1)62NAI-5006826801V1and Flk-1 / KDR (VEGFR2), inhibitors of epidermal-derived, fibroblast-derived, or platelet derived growth factors, MMP (matrix metalloprotease) inhibitors, integrin blockers, interferon- a, interleukin- 12, pentosan poly sulfate, cyclooxygenase inhibitors, including nonsteroidal antiinflammatories (NSAIDs) like aspirin and ibuprofen as well as selective cyclooxy -genase-2 inhibitors like celecoxib and rofecoxib (PNAS, Vol. 89, p. 7384 (1992); JNCI, Vol. 69, p. 475 (1982); Arch. Opthalmol., Vol. 108, p.573 (1990); Anat. Rec., Vol. 238, p. 68 (1994); FEBS Letters, Vol. 372, p. 83 (1995); Clin, Orthop. Vol. 313, p. 76(1995); J. Mol. Endocrinol., Vol. 16, p.107 (1996); Jpn. J. Pharmacol., Vol. 75, p. 105 (1997); Cancer Res., Vol. 57, p. 1625 (1997); Cell, Vol. 93, p. 705 (1998); Inti. J. Mol. Med., Vol. 2, p. 715 (1998); J. Biol. Chem., Vol. 274, p. 9116 (1999)), steroidal anti-inflammatories (such as corticosteroids, mineralocorticoids, dexamethasone, prednisone, prednisolone, methylpred, betamethasone), carboxyamidotriazole, combretastatin A-4, squalamine, 6-O-chloroacetyl-carbonyl)-fumagillol, thalidomide, angiostatin, troponin-1, angiotensin II antagonists (see Fernandez et al., J. Lab. Clin. Med. 105: 141-145 (1985)), and antibodies to VEGF (see, Nature Biotechnology, Vol. 17, pp.963-968 (October 1999); Kim et al., Nature, 362, 841-844 (1993); WO 00 / 44777; and WO 00 / 61186).

[0190] Other examples of angiogenesis inhibitors include, but are not limited to, endostatin, ukrain, ranpimase, IM862, 5-methoxy-4-[2-methyl-3-(3-methyl-2-butenyl)oxiranyl]-l- oxaspiro[2,5]oct-6-yl(chloroacetyl)carbamate, acetyldinanaline, 5-amino-l-[[3,5-dichloro-4-(4- chlorobenzoyl)phenyl]methyl]-lH-l,2,3-triazole-4-carboxamide,CM101, squalamine, combretastatin, RPI4610, NX31838, sulfated mannopentaose phosphate, 7,7-(carbonyl- bis[imino-N-methyl-4,2-pyrrolocarbonylimino[N-methyl-4,2-pyrrole]-carbonylimino]-bis- (1,3- naphthalene disulfonate), and 3-[(2,4-dimethylpyrrol-5-yl)methylene]-2-indolinone (SU5416).

[0191] Some specific examples of tyrosine kinase inhibitors include N- (trifluoromethylphenyl)-5-methylisoxazol-4-carboxamide, 3-[(2,4-dimethylpyrrol-5- yl)methylidenyl)indolin-2-one, 17-(allylamino)-17-demethoxygeldanamycin, 4-(3-chloro-4- fluorophenylamino)-7-methoxy-6-[3-(4-morpholinyl)propoxyl]quinazoline, N-(3- ethynylphenyl)-6,7-bis(2-methoxyethoxy)-4-quinazolinamine, BIBX1382, 2,3,9,10,11,12- hexahydro-10-(hydroxymethyl)-10-hydroxy-9-methyl-9,12-epoxy-lH-diindolo[l,2,3- fg:3’,2’,r-kl]pyrrolo[3,4-i][l,6]benzodiazocin-l-one, SH268, genistein, STI571, CEP2563, 4- (3-chlorophenylamino)-5,6-dimethyl-7H-pyrrolo[2,3-d]pyrimidinemethane sulfonate, 4-(3- bromo-4-hydroxyphenyl)amino-6,7-dimethoxyquinazoline, 4-(4’-hydroxyphenyl)amino-6,7- dimethoxyquinazoline, SU6668, STI571A, N-4-chlorophenyl-4-(4-pyridylmethyl)-l- phthalazinamine, and EMD121974.63NAI-5006826801V1

[0192] In some examples, anti-Nectin-4 antibodies or antigen binding fragments may be used in association with PPAR-y (i.e., PPAR-gamma) agonists or PPAR-6 (i.e., PPAR-delta) agonists. PPAR-y and PPAR-6 are the nuclear peroxisome proliferator-activated receptors y and 6. The expression of PPAR-y on endothelial cells and its involvement in angiogenesis has been reported in the literature (see Agarwal J. Cardiovasc. Pharmacol. 1998; 31 :909-913; J. Biol. Chem. 1999;274:9116-9121; Murata et al., Invest. Ophthalmol Vis. Sci. 2000; 41 :2309-2317). PPAR-y agonists have been shown to inhibit the angiogenic response to VEGF in vitro; both troglitazone and rosiglitazone maleate inhibit the development of retinal neovascularization in mice. (Arch. Ophthamol. 2001; 119:709-717). Examples of PPAR-y agonists and PPAR-y / a agonists include, but are not limited to, Lynparza®, Rucaparib®, Talazoparib®, niraparib, Veliparib®, thiazolidinediones (such as DRF2725, CS-011, troglitazone, rosiglitazone, and pioglitazone), fenofibrate, gemfibrozil, clofibrate, GW2570, SB219994, AR-H039242, JTT-501, MCC-555, GW2331, GW409544, NN2344, KRP297, NP0110, DRF4158, NN622, GI262570, PNU182716, DRF552926, 2-[(5,7-dipropyl-3-trifluoromethyl-l,2-benzisoxazol-6-yl)oxy]-2- methylpropionic acid, and 2(R)-7-(3-(2-chloro-4-(4-fluorophenoxy) phenoxy )propoxy)-2- ethylchromane-2-carboxylic acid.

[0193] Anti-Nectin-4 antibodies, antigen-binding fragments thereof, or antibody conjugates may also be used for treating or preventing breast cancer in association with aromatase inhibitors. Examples of aromatase inhibitors include but are not limited to: anastrozole, letrozole and exemestane.

[0194] Anti-Nectin-4 antibodies, antigen-binding fragments thereof, or antibody conjugates may also be used in association with one or more of the following chemotherapeutic agents: abarelix (Plenaxis depot®); aldesleukin (Prokine®); Aldesleukin (Proleukin®); alitretinoin (Panretin®); allopurinol (Zyloprim®); altretamine (Hexalen®); amifostine (Ethyol®); anastrozole (Arimidex®); arsenic trioxide (Trisenox®); asparaginase (Elspar®); azacitidine (Vidaza®); bendamustine hydrochloride (Treanda®); bexarotene capsules (Targretin®); bexarotene gel (Targretin®); bleomycin (Blenoxane®); bortezomib (Velcade®); brefeldin A; busulfan intravenous (Busulfex®); busulfan oral (Myleran®); calusterone (Methosarb®); capecitabine (Xeloda®); carboplatin (Paraplatin®); carmustine (BCNU®, BiCNU®); carmustine (Gliadel®); carmustine with Polifeprosan 20 Implant (Gliadel Wafer®); celecoxib (Celebrex®); chlorambucil (Leukeran®); cisplatin (Platinol®); cladribine (Leustatin®, 2- CdA®); clofarabine (Clolar®); cyclophosphamide (Cytoxan®, Neosar®); cyclophosphamide (Cytoxan Injection®); cyclophosphamide (Cytoxan Tablet®); cytarabine (Cytosar-U®); cytarabine liposomal (DepoCyt®); dacarbazine (DTIC-Dome®); dactinomycin, actinomycin D64NAI-5006826801V1(Cosmegen®); dalteparin sodium injection (Fragmin®); darbepoetin alfa (Aranesp®); dasatinib (Sprycel®); daunorubicin liposomal (DanuoXome®); daunomycin (Daunorubicin®); daunorubicin, daunomycin (Cerubidine®); degarelix (Firmagon®); Denileukin diftitox (Ontak®); dexrazoxane (Zinecard®); dexrazoxane hydrochloride (Totect®); didemnin B; 17- DMAG; docetaxel (Taxotere®); doxorubicin (Adriamycin PFS®); doxorubicin (Adriamycin®, Rubex®); doxorubicin (Adriamycin PFS Injection®); doxorubicin liposomal (Doxil®); dromostanolone propionate (Dromostanolone ®); dromostanolone propionate (Masterone Injection®); eculizumab injection (Soliris®); Elliott's B Solution (Elliott's B Solution®); eltrombopag (Promacta®); epirubicin (Ellence®); Epoetin alfa (epogen®); erlotinib (Tarceva®); estramustine (Emcyt®); ethinyl estradiol; etoposide phosphate (Etopophos®); etoposide, VP- 16 (Vepesid®); everolimus tablets (Afinitor®); exemestane (Aromasin®); ferumoxytol (Feraheme Injection®); floxuridine (intraarterial) (FUDR®); fludarabine (Fludara®); fluorouracil, 5-FU (Adrucil®); fulvestrant (Faslodex®); gefitinib (Iressa®); geldanamycin; gemcitabine (Gemzar®); gemtuzumab ozogamicin (Mylotarg®); goserelin acetate (Zoladex Implant®); goserelin acetate (Zoladex®); histrelin acetate (Histrelin implant®); hydroxyurea (Hydreaidarubicin (Idamycin®); ifosfamide (IFEX®); imatinib mesylate (Gleevec®); interferon alfa 2a (Roferon A®); Interferon alfa-2b (Intron A®); iobenguane I123injection (AdreView®); irinotecan (Camptosar®); ixabepilone (Ixempra®); lapatinib tablets (Tykerb®); lenalidomide (Revlimid®); letrozole (Femara®); leucovorin (Wellcovorin®, Leucovorin®); Leuprolide Acetate (Eligard®); levamisole (Ergamisol®); lomustine, CCNU (CeeBU®); meclorethamine, nitrogen mustard (Mustargen®); megestrol acetate (Megace®); melphalan, L-PAM (Alkeran®); mercaptopurine, 6-MP (Purinethol®); mesna (Mesnex®); mesna (Mesnex tabs®); methotrexate (Methotrexate®); methoxsalen (Uvadex®); 8-methoxypsoralen; mitomycin C (Mutamycin®); mitotane (Lysodren®); mitoxantrone (Novantrone®); mitramycin; nandrolone phenpropionate (Durabolin-50®); nelarabine (Arranon®); nilotinib (Tasigna®); Nofetumomab (Verluma®); ofatumumab (Arzerra®); Oprelvekin (Neumega®); oxaliplatin (Eloxatin®); paclitaxel (Paxene®); paclitaxel (Taxol®); paclitaxel protein-bound particles (Abraxane®); palifermin (K epivance®); pamidronate (Aredia®); panitumumab (Vectibix®); pazopanib tablets (Votrienttm®); pegademase (Adagen (Pegademase Bovine)®); pegaspargase (Oncaspar®); pemetrexed disodium (Alimta®); pentostatin (Nipent®); pipobroman (Vercyte®); plerixafor (Mozobil®); plicamycin, mithramycin (Mithracin®); porfimer sodium (Photofrin®); pralatrexate injection (Folotyn®); procarbazine (Matulane®); quinacrine (Atabrine®); rapamycin; rasburicase (Elitek®); raloxifene hydrochloride (Evista®); romidepsin (Istodax®); romiplostim (Nplate®);65NAI-5006826801V1sargramostim (Leukine®); sargramostim (Prokine®); sorafenib (Nexavar®); streptozocin (Zanosar®); sunitinib maleate (Sutent®); talc (Sclerosol®); tamoxifen (Nolvadex®); temozolomide (Temodar®); temsirolimus (Torisel®); teniposide, VM-26 (Vumon®); testolactone (Teslac®); thioguanine, 6-TG (Thioguanine®); thiopurine; thiotepa (Thioplex®); topotecan (Hycamtin®); toremifene (Fareston®); Tositumomab (Bexxar®); Tositumomab / I-131 tositumomab (Bexxar®); trans-retinoic acid; tretinoin, ATRA (Vesanoid®); triethylenemelamine; uracil mustard (Uracil Mustard Capsules®); valrubicin (Valstar®); vinblastine (Velban®); vincristine (Oncovin®); vinorelbine (Navelbine®); vorinostat (Zolinza®); wortmannin; and zoledronate (Zometa®).

[0195] In an embodiment, an anti-Nectin-4 antibody, antigen-binding fragment thereof, or antibody conjugate of the disclosure is used in association with anti-cancer radiation therapy. For example, in an embodiment, the radiation therapy is external beam therapy (EBT): a method for delivering a beam of high-energy X-rays to the location of the tumor. The beam is generated outside the patient (e.g., by a linear accelerator) and is targeted at the tumor site. These X-rays can destroy the cancer cells and careful treatment planning allows the surrounding normal tissues to be spared. No radioactive sources are placed inside the patient's body. In an embodiment, the radiation therapy is proton beam therapy: a type of conformal therapy that bombards the diseased tissue with protons instead of X-rays. In an embodiment, the radiation therapy is conformal external beam radiation therapy: a procedure that uses advanced technology to tailor the radiation therapy to an individual's body structures. In an embodiment, the radiation therapy is brachytherapy: the temporary placement of radioactive materials within the body, usually employed to give an extra dose or boost of radiation to an area.

[0196] In an embodiment, an anti-Nectin-4 antibody, antigen-binding fragment thereof, or antibody conjugate of the disclosure is used in association with a surgical procedure such as surgical tumorectomy.

[0197] In some examples, the patient is infused with autologous T cells expanded ex vivo with anti-Nectin-4 specific antibodies, antigen-binding fragments thereof, or antibody conjugates. In another embodiment, the patient is administered autologous T cells in combination with the anti- Nectin-4 specific antibodies, antigen-binding fragments thereof, or antibody conjugates. In yet another embodiment, the patient is vaccinated with a cancer vaccine, and infused with autologous T cells expanded ex vivo with anti-Nectin-4 specific a antibodies, antigen-binding fragments thereof, or antibody conjugates. The autologous T-cells can be autologous infiltrating lymphocytes, T-cells transduced with high affinity T-cell receptors against tumor antigens or T cells transduced with chimeric antigen receptors composed of hybrid immunoglobulin lights66NAI-5006826801V1chains with endo-domains of T-cell signaling molecules. See Kalos M. and June C. H., Immunity, 39, 2013, p49-60; Wu R. et al., Cancer J. 2012; 18(2): 160-175; and June, C. H, J. Clin. Invest. 117: 1466-1476 (2007).Anti-Nectin-4 Antibodies and Antigen-binding Fragments

[0198] The present disclosure provides anti-Nectin-4 antibodies and antigen-binding fragments thereof having specified structural and / or functional features, methods of use of the antibodies or antigen-binding fragments thereof in the treatment or prevention of disease (e.g., cancers expressing Nectin-4), and antibody conjugates and comprising the antibodies or antigenbinding fragments thereof for use in the treatment or prevention of disease (e.g., cancers expressing Nectin-4).

[0199] Any suitable anti-Nectin-4 antibody (e.g., enfortumab) may be used in the methods, compositions, uses, and kits disclosed herein. In some examples, the anti-Nectin-4 antibody or antigen-binding fragment thereof that is discussed herein comprises a heavy chain variable region (VH) comprising 3 heavy chain CDRs (CDRH): CDR-H1, CDR-H2 and CDR-H3. The CDR-H1, CDR-H2 and CDR-H3 are selected from the CDRH group in Table 1.Table 1. Heavy chain CDRs of anti-Nectin-4 antibodies67NAI-5006826801V1

[0200] In some examples, the anti-Nectin-4 antibody or antigen-binding fragment thereof that is discussed herein comprises a light chain variable region (VL) comprising 3 light chain CDRs (CDRL): CDR-L1, CDR-L2 and CDR-L3. The CDR-L1, CDR-L2 and CDR-L3 are selected from the CDRL group in Table 2.Table 2. Light chain CDRs of anti-Nectin-4 antibody68NAI-5006826801V1

[0201] There are several methods available for mapping antibody epitopes on target antigens, including: H / D-Ex Mass spec, X-ray crystallography, pepscan analysis, alanine scanning, hydroxyl radical foot printing and site directed mutagenesis. For example, HDX (Hydrogen Deuterium Exchange) coupled with proteolysis and mass spectrometry can be used to determine the epitope of an antibody on a specific antigen Y. HDX-MS relies on the accurate measurement and comparison of the degree of deuterium incorporation by an antigen when incubated in D2O on its own and in presence of its antibody at various time intervals. Deuterium is exchanged with hydrogen on the amide backbone of the proteins in exposed areas whereas regions of the antigen bound to the antibody will be protected and will show less or no exchange after analysis by LC-MS / MS of proteolytic fragments. In one embodiment, the epitope is determined by solving the X-ray crystal structure of a complex between Nectin-4 or a fragment thereof and an anti-Nectin-4 antibody or fragment thereof and identifying one or more Nectin-4 residues within 4 A of the anti-Nectin-4 antibody residues. In another embodiment, the epitope includes for example, Nectin-4 residues that have van der Waals, polar interaction, salt bridge or hydrogen bond contact with the anti-Nectin-4 antibody residues. In another embodiment, the epitope is determined by mutagenesis (for example Alanine scanning) of Nectin-4 residues and analyzing the loss of binding to the anti-Nectin-4 antibody as a result of the mutagenesis.

[0202] Also provided are isolated polypeptides comprising the VL domains of the anti-Nectin- 4 antibodies of the disclosure (e.g., SEQ ID NOS: 2, 20, 38, 56, 58, and 60), and isolated polypeptides comprising the VH domains of the anti-Nectin-4 antibodies of the disclosure (e.g., SEQ ID NOS: 1, 19, 37, 55, 57, and 59) having up to 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 or more amino acid substitutions. In some examples, the substitutions are in the framework regions. In other examples, the substitutions are in the CDR regions. In other examples, the substations are in the CDR regions and the framework regions.

[0203] In another embodiment, an antibody or antigen-binding fragment thereof that binds Nectin-4 and has VL domains and VH domains with at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, 80% or 75% sequence identity to one or more of the VL domains or VH domains described herein and exhibits specific binding to Nectin-4 may also be utilized in the methods and uses described herein. In another embodiment, the binding antibody or antigen-binding fragment thereof of the present disclosure comprises VL and VH domains (with and without69NAI-5006826801V1signal sequence) having up to 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25 or more amino acid substitutions, and exhibits specific binding to Nectin-4.

[0204] Also included in the present disclosure are polypeptides, e.g., immunoglobulin polypeptides, comprising amino acid sequences that are at least about 75% identical, 80% identical, more preferably at least about 90% identical and most preferably at least about 95% identical (e.g., 95%, 96%, 97%, 98%, 99%, 100%) to the amino acid sequences of the antibodies provided herein when the comparison is performed by a BLAST algorithm wherein the parameters of the algorithm are selected to give the largest match between the respective sequences over the entire length of the respective reference sequences (e.g., expect threshold: 10; word size: 3; max matches in a query range: 0; BLOSUM 62 matrix; gap costs: existence 11, extension 1; conditional compositional score matrix adjustment).Polynucleotides

[0205] The present disclosure further provides polynucleotides encoding any of the polypeptides or immunoglobulin chains of antibodies (e.g., anti-Nectin-4 antibodies) and antigen-binding fragments thereof of the disclosure. For example, the present disclosure includes polynucleotides encoding an anti-Nectin-4 antibody or antigen-binding fragment thereof having one or more of the amino acids described in Tables 1 and 2.

[0206] In one embodiment, an isolated polynucleotide, for example DNA, encoding one or more of the polypeptide chains of the isolated antibodies or antigen-binding fragments thereof set forth herein is provided. In one embodiment, the isolated polynucleotide encodes an antibody or antigen- binding fragment thereof comprising at least one mature immunoglobulin light chain variable (VL) domain according to the disclosure and / or at least one mature immunoglobulin heavy chain variable (VH) domain according to the disclosure. In some embodiments the isolated polynucleotide encodes both a light chain and a heavy chain on a single polynucleotide molecule, and in other embodiments the light and heavy chains are encoded on separate polynucleotide molecules. In another embodiment the polynucleotide further encodes a signal sequence.

[0207] The present disclosure also provides vectors, e.g., expression vectors, such as plasmids, comprising the isolated polynucleotides of the disclosure, wherein the polynucleotide is operably linked to control sequences that are recognized by a host cell when the host cell is transfected with the vector. Also provided are host cells comprising a vector or vectors of the present disclosure and methods for producing an antibody or antigen-binding fragment thereof or polypeptide disclosed herein comprising culturing a host cell harboring one or more expression vectors or one or more nucleic acids encoding the immunoglobulin chains of the70NAI-5006826801V1antibody or antigen-binding fragment thereof in culture medium, and isolating the antigen or antigen-binding fragment thereof from the host cell or culture medium.Methods of Making Antibodies and Antigen-binding Fragments Thereof

[0208] Methods for making antibodies (e.g., anti-Nectin-4 antibodies) and antigen-binding fragments thereof are described herein. For example, the disclosure provides methods comprising culturing a hybridoma cell that expresses the antibody or fragment thereof under conditions favorable to such expression and, optionally, isolating the antibody or fragment thereof from the hybridoma and / or the growth medium (e.g., cell culture medium).

[0209] The antibodies disclosed herein may also be produced recombinantly (e.g., in an E. colU l expression system, a mammalian cell expression system, or a lower eukaryote expression system). Nucleic acids encoding the antibody immunoglobulin molecules of the disclosure (e.g., VH and / or VL) may be inserted into a pET -based plasmid and expressed in the E. colUTl system. For example, methods for expressing an antibody or antigen-binding fragment thereof or immunoglobulin chain thereof in a host cell (e.g., bacterial host cell such as E.coli such as BL21 or BL21DE3) are described herein. In various embodiments, the methods comprise expressing T7 RNA polymerase in the cell which also includes a polynucleotide encoding an immunoglobulin chain that is operably linked to a T7 promoter. For example, a bacterial host cell, such as an E. coli cell, includes a polynucleotide encoding the T7 RNA polymerase gene operably linked to a lac promoter and expression of the polymerase and the chain is induced by incubation of the host cell with isopropyl-beta-D-thiogalactopyranoside (IPTG).

[0210] There are several methods by which to produce recombinant antibodies which are known in the art. One example of a method for recombinant production of antibodies is disclosed in U.S. Patent No. 4,816,567.

[0211] Transformation can be by any known method for introducing polynucleotides into a host cell. Methods for introduction of heterologous polynucleotides into mammalian cells are well known in the art and include dextran-mediated transfection, calcium phosphate precipitation, polybrene-mediated transfection, protoplast fusion, electroporation, encapsulation of the polynucleotide(s) in liposomes, biolistic injection and direct microinjection of the DNA into nuclei. In addition, nucleic acid molecules may be introduced into mammalian cells by viral vectors. Methods of transforming cells are well known in the art. See, for example, U.S. Patent Nos. 4,399,216; 4,912,040; 4,740,461 and 4,959,455.

[0212] Described herein are recombinant methods for making an antibody or antigen-binding fragment thereof of the present disclosure, or an immunoglobulin chain thereof, comprising71NAI-5006826801V1introducing a polynucleotide encoding one or more immunoglobulin chains of the antibody or fragment thereof (e.g., heavy and / or light immunoglobulin chain); culturing the host cell (e.g., CHO or Pichia (e.g., Pichia pastoris)) under condition favorable to such expression and, optionally, isolating the antibody or fragment thereof or chain from the host cell and / or medium in which the host cell is grown. Antibodies can also be synthesized by any of the methods set forth in U.S. Patent No. 6,331,415.

[0213] Eukaryotic and prokaryotic host cells, including mammalian cells as hosts for expression of the antibodies or fragments thereof or immunoglobulin chains disclosed herein are well known in the art and include many immortalized cell lines available from the American Type Culture Collection (ATCC). These include, inter alia, Chinese hamster ovary (CHO) cells, NSO, SP2 cells, HeLa cells, baby hamster kidney (BHK) cells, monkey kidney cells (COS), human hepatocellular carcinoma cells (e.g., Hep G2), A549 cells, 3T3 cells, HEK-293 cells and a number of other cell lines. Mammalian host cells include human, mouse, rat, dog, monkey, pig, goat, bovine, horse and hamster cells. Cell lines may be selected by determining which cell lines have high expression levels. Other cell lines that may be used are insect cell lines, such as Sf9 cells, amphibian cells, bacterial cells, plant cells and fungal cells. Fungal cells include yeast and filamentous fungus cells including, for example, Pichia sp. (e.g., Pichia pastoris, Pichia fmlandica, Pichia trehalophila, Pichia koclamae, Pichia membranaefaciens, Pichia minuta (Ogataea minuta, Pichia lindneri), Pichia opuntiae, Pichia thermotolerans, Pichia salictaria, Pichia guercuum, Pichia pijperi, Pichia stiptis, o Pichia methanolica), Saccharomyces sp.(e.g., Saccharomyces cerevisiae), Hansenula polymorpha, Kluyveromyces sp. (e.g., Kluyveromyces lachs), Candida sp. (e.g., Candida albicans), Aspergillus sp. (e.g., Aspergillus nidulans, Aspergillus niger, or Aspergillus oryzae), Trichoderma reesei, Chrysosporium lucknowense, Fusarium sp. (e.g., Fusarium gramineum ox Fusarium venenatum), Physcomitrella patens and Neurospora crassa.

[0214] When recombinant expression vector(s) encoding the heavy chain or antigen-binding portion or fragment thereof, the light chain or antigen-binding fragment thereof, or both, are introduced into host cells, the antibodies are produced by culturing the host cells for a period of time sufficient to allow for expression of the antibody or fragment thereof or chain in the host cells or secretion of the into the culture medium in which the host cells are grown.

[0215] Antibodies and antigen-binding fragments thereof and immunoglobulin chains can be recovered from the culture medium using standard protein purification methods. Further, expression of antibodies and antigen-binding fragments thereof and immunoglobulin chains (or other moieties therefrom) are described herein. The expression for examples is achieved from72NAI-5006826801V1production cell lines that can be enhanced using a number of known techniques. For example, the glutamine synthetase gene expression system (the GS system) is a common approach for enhancing expression under certain conditions. The GS system is discussed in European Patent Nos. 0 216 846, 0 256 055, and 0 323 997 and European Patent Application No. 89303964.4. The mammalian host cells (e.g., CHO) may lack a glutamine synthetase gene and are grown in the absence of glutamine in the medium wherein, however, the polynucleotide encoding the immunoglobulin chain comprises a glutamine synthetase gene which complements the lack of the gene in the host cell.

[0216] In general, glycoproteins produced in a particular cell line or transgenic animal will have a glycosylation pattern that is characteristic for glycoproteins produced in the cell line (e.g., CHO cells) or transgenic animal. Therefore, the particular glycosylation pattern of an antibody will depend on the particular cell line or transgenic animal used to produce the antibody. However, all antibodies encoded by the nucleic acid molecules provided herein, or comprising the amino acid sequences provided herein, are included in the present disclosure, independent of the glycosylation pattern the antibodies may have. Similarly, in particular embodiments, antibodies with a glycosylation pattern comprising only non-fucosylated N- glycans may be advantageous, because these antibodies have been shown to typically exhibit more potent efficacy than their fucosylated counterparts both in vitro and in vivo (See for example, Shinkawa et al., J. Biol. Chem. 278: 3466-3473 (2003); U.S. Patent Nos. 6,946,292 and 7,214,775). These antibodies with non-fucosylated N-glycans are not likely to be immunogenic because their carbohydrate structures are a normal component of the population that exists in human serum IgG.

[0217] Bispecific antibodies can be produced by a variety of methods including fusion of hybridomas or linking of Fab’ fragments. See, e.g., Songsivilai, et al., (1990) Clin. Exp. Immunol. 79: 315-321, Kostelny, et al., (1992) J Immunol. 148: 1547- 1553. In addition, bispecific antibodies may be formed as “diabodies” (Holliger et al., (1993) PNAS USA 90:6444-6448) or as “Janusins” (Traunecker et al., (1991) EMBO J. 10:3655-3659 and Traunecker, et al., (1992) Int. J. Cancer Suppl. 7:51-52).Antibody Engineering

[0218] Further included are embodiments in which the antibodies (e.g., anti-Nectin-4 antibodies) and antigen-binding fragments thereof are engineered antibodies.

[0219] In certain embodiments, the antibodies and antigen-binding fragments thereof are engineered to include modifications in the framework and / or CDRs to improve their properties. Such engineered changes can be based on molecular modeling. A molecular model for the73NAI-5006826801V1variable region for the parental (non-human) antibody sequence can be constructed to understand the structural features of the antibody and used to identify potential regions on the antibody that can interact with the antigen. Conventional CDRs are based on alignment of immunoglobulin sequences and identifying variable regions. Kabat et al., (1991) Sequences of Proteins of Immunological Interest, Kabat, et al.; National Institutes of Health, Bethesda, Md.; 5th ed.; NIH Publ. No. 91-3242; Kabat (1978) Adv. Prot. Chem. 32: 1-75; Kabat, et al., (1977) J. Biol. Chem. 252:6609-6616. Chothia and coworkers carefully examined conformations of the loops in crystal structures of antibodies and proposed hypervariable loops. Chothia, et al., (1987) J Mol. Biol. 196:901-917 or Chothia, et al., (1989) Nature 342:878-883. There are variations between regions classified as CDRs and hypervariable loops. Later studies (Raghunathan et al., (2012) J. Mol Recog. 25, 3, 103-113) analyzed several antibody-antigen crystal complexes and observed that the antigen binding regions in antibodies do not necessarily conform strictly to the CDR residues or hypervariable loops. The molecular model for the variable region of the non-human antibody can be used to guide the selection of regions that can potentially bind to the antigen. In practice, the potential antigen binding regions based on model differ from the conventional CDRs or hypervariable loops. Commercial scientific software such as MOE (Chemical Computing Group) can be used for molecular modeling. Human frameworks can be selected based on best matches with the non-human sequence both in the frameworks and in the CDRs. For FR4 (framework 4) in VH, VJ regions for the human germlines are compared with the corresponding non-human region. In the case of FR4 (framework 4) in VL, J-kappa and J-Lambda regions of human germline sequences are compared with the corresponding non- human region. Once suitable human frameworks are identified, the CDRs are grafted into the selected human frameworks. In some cases, the certain residues in the VL-VH interface can be retained as in the non-human (parental) sequence. Molecular models can also be used for identifying residues that can potentially alter the CDR conformations and hence binding to antigen. In some cases, these residues are retained as in the non-human (parental) sequence. Molecular models can also be used to identify solvent exposed amino acids that can result in unwanted effects such as glycosylation, deamidation and oxidation. Developability filters can be introduced early on in the design stage to eliminate / minimize these potential problems.

[0220] In some examples, the engineered antibodies include modifications to framework residues within the variable domains of the antibody, e.g., to improve the properties of the antibody or fragment thereof. Typically, such framework modifications are made to decrease the immunogenicity of the antibody or fragment thereof. This is usually accomplished by replacing non-CDR residues in the variable domains (i.e., framework residues) in a parental (e.g., rodent)74NAI-5006826801V1antibody or fragment thereof with analogous residues from the immune repertoire of the species in which the antibody is to be used, e.g., human residues in the case of human therapeutics. In some embodiments, it is desirable to increase the affinity, or alter the specificity of an engineered (e.g., humanized) antibody. One approach is to “back-mutate” one or more framework residues to the corresponding germline sequence. More specifically, an antibody or fragment thereof that has undergone somatic mutation can contain framework residues that differ from the germline sequence from which the antibody is derived. Such residues can be identified by comparing the antibody or fragment framework sequences to the germline sequences from which the antibody or fragment thereof is derived. Another approach is to revert to the original parental (e.g., rodent) residue at one or more positions of the engineered (e.g., humanized) antibody, e.g., to restore binding affinity that may have been lost in the process of replacing the framework residues. See, e.g., U.S. Patent No. 5,693,762, U.S. Patent No. 5,585,089 and U.S. Patent No. 5,530,101.

[0221] Another type of modification involves mutating one or more residues within the framework region, or even within one or more CDR regions, to remove T cell epitopes to thereby reduce the potential immunogenicity of the antibody. This approach is also referred to as “deimmunization” and is described in further detail in U.S. Patent No. 7,125,689.

[0222] In some embodiments, engineering may include changing certain amino acids containing exposed side-chains to another amino acid residue in order to provide for greater chemical stability of the final antibody, so as to avoid deamidation or isomerization. The deamidation of asparagine may occur on NG, DG, NG, NS, NA, NT, QG or QS sequences and result in the creation of an isoaspartic acid residue that introduces a kink into the polypeptide chain and decreases its stability (isoaspartic acid effect). Isomerization can occur at DG, DS, DA or DT sequences. In certain embodiments, the antibodies do not contain deamidation or asparagine isomerism sites.

[0223] For example, an asparagine (Asn) residue may be changed to Gin or Ala to reduce the potential for formation of isoaspartate at any Asn-Gly sequences, particularly within a CDR.

[0224] A similar problem may occur at an Asp-Gly sequence. See, e.g., Reissner and Aswad (2003) Cell. Mol. Life Sci. 60: 1281. Isoaspartate formation may debilitate or completely abrogate binding of an antibody to its target antigen. See, e.g., Presta (2005) J. Allergy Clin. Immunol. 116:731.

[0225] In one embodiment, the asparagine is changed to glutamine (Gin). It may also be desirable to alter an amino acid adjacent to an asparagine (Asn) or glutamine (Gin) residue to reduce the likelihood of deamidation, which occurs at greater rates when small amino acids75NAI-5006826801V1occur adjacent to asparagine or glutamine. See, Bischoff & Kolbe (1994) J. Chromatog. 662:261. In addition, Met residues (typically solvent exposed Met) in CDRs may be changed to Lys, Leu, Ala, or Phe or other amino acids in order to reduce the possibility that the methionine sulfur would oxidize, which could reduce antigen-binding affinity and also contribute to molecular heterogeneity in the final antibody preparation. Additionally, in order to prevent or minimize potential scissile Asn-Pro peptide bonds, it may be desirable to alter any Asn-Pro combinations found in a CDR to Gin-Pro, Ala-Pro, or Asn-Ala. Antibodies with such substitutions are subsequently screened to ensure that the substitutions do not decrease the affinity or specificity of the antibody for Nectin-4, or other desired biological activity to unacceptable levels. Exemplary variants for stabilizing CDRs are shown in the table below.Antibody Engineering of the Fc Region

[0226] The antibodies (e.g., anti-Nectin-4 antibodies) and antigen-binding fragments thereof disclosed herein can also be engineered to include modifications within the Fc region, typically to alter one or more properties of the antibody, such as serum half-life, complement fixation, Fc receptor binding, and / or effector function (e.g., antigen-dependent cellular cytotoxicity). Furthermore, the antibodies and antigen-binding fragments thereof disclosed herein can be chemically modified (e.g., one or more chemical moieties can be attached to the antibody) or be modified to alter its glycosylation, again to alter one or more properties of the antibody or fragment thereof. Each of these embodiments is described in further detail below. The numbering of residues in the Fc region is that of the EU index of Kabat. The antibodies and antigen-binding fragments thereof disclosed herein also include antibodies and fragments with modified (or blocked) Fc regions to provide altered effector functions. See, e.g., U.S. Pat. No. 5,624,821; and PCT publications numbers W02003 / 086310; W02005 / 120571; and W02006 / 0057702. Such modifications can be used to enhance or suppress various reactions of the immune system, with possible beneficial effects in diagnosis and therapy. Alterations of the Fc region include amino acid changes (substitutions, deletions, and insertions), glycosylation or76NAI-5006826801V1deglycosylation, and adding multiple Fc regions. Changes to the Fc can also alter the half-life of antibodies in therapeutic antibodies, enabling less frequent dosing and thus increased convenience and decreased use of material. See Presta (2005) J. Allergy Clin. Immunol. 116:731 at 734-35.

[0227] In one embodiment, the antibody or antigen-binding fragment thereof described herein is an IgG4 isotype antibody or fragment thereof comprising a Serine to Proline mutation at a position corresponding to position 228 (S228P; EU index) in the hinge region of the heavy chain constant region. This mutation has been reported to abolish the heterogeneity of inter-heavy chain disulfide bridges in the hinge region (Angal et al. supra; position 241 is based on the Kabat numbering system).

[0228] In one embodiment of the invention, the hinge region of CHI is modified such that the number of cysteine residues in the hinge region is increased or decreased. This approach is described further in U.S. Patent No. 5,677,425. The number of cysteine residues in the hinge region of CHI is altered, for example, to facilitate assembly of the light and heavy chains or to increase or decrease the stability of the antibody.

[0229] In another embodiment, the Fc hinge region of an antibody or antigen-binding fragment thereof is mutated to decrease the biological half-life of the antibody or fragment thereof. More specifically, one or more amino acid mutations are introduced into the CH2-CH3 domain interface region of the Fc- hinge fragment such that the antibody or fragment thereof has impaired Staphylococcal protein A (SpA) binding relative to native Fc-hinge domain SpA binding. This approach is described in further detail in U.S. Patent No. 6,165,745.

[0230] In another embodiment, the antibody or antigen-binding fragment thereof is modified to increase its biological half-life. Various approaches are possible. For example, one or more of the following mutations can be introduced: T252L, T254S, T256F, as described in U.S. Patent No. 6,277,375. Alternatively, to increase the biological half-life, the antibody can be altered within the CHI or CL region to contain a salvage receptor binding epitope taken from two loops of a CH2 domain of an Fc region of an IgG, as described in U.S. Patent Nos. 5,869,046 and 6,121,022.

[0231] In yet other embodiments, the Fc region is altered by replacing at least one amino acid residue with a different amino acid residue to alter the effector function(s) of the antibody or antigen-binding fragment thereof. For example, one or more amino acids selected from amino acid residues 234, 235, 236, 237, 297, 318, 320 and 322 can be replaced with a different amino acid residue such that the antibody has an altered affinity for an effector ligand and retains the antigen-binding ability of the parent antibody. The effector ligand to which affinity is altered77NAI-5006826801V1can be, for example, an Fc receptor or the Cl component of complement. This approach is described in further detail in U.S. Patent Nos. 5,624,821 and 5,648,260.

[0232] In another example, one or more amino acids selected from amino acid residues 329, 331 and 322 can be replaced with a different amino acid residue such that the antibody has altered Clq binding and / or reduced or abolished complement dependent cytotoxicity (CDC). This approach is described in further detail in U.S. Patent No. 6,194,551.

[0233] In another example, one or more amino acid residues within amino acid positions 231 and 239 are altered to thereby alter the ability of the antibody to fix complement. This approach is described further in PCT Publication WO 94 / 29351.

[0234] In some embodiments, the Fc region of an antibody is modified to increase or reduce the ability of the antibody or antigen-binding fragment thereof to mediate effector function and / or to increase / decrease their binding to the Fc gamma receptors (FcyRs).

[0235] For example, in some embodiments, the Fc region is modified to decrease the ability of the antibody or antigen-binding fragment thereof to mediate antibody dependent cellular cytotoxicity (ADCC) and / or to decrease the affinity of the antibody or fragment thereof for an Fey receptor by modifying one or more amino acids at the following positions: 238, 239, 243, 248, 249, 252, 254, 255, 256, 258, 264, 265, 267, 268, 269, 270, 272, 276, 278, 280, 283, 285,286, 289, 290, 292, 293, 294, 295, 296, 298, 301, 303, 305, 307, 309, 312, 315, 320, 322, 324,326, 327, 329, 330, 331, 333, 334, 335, 337, 338, 340, 360, 373, 376, 378, 382, 388, 389, 398,414, 416, 419, 430, 434, 435, 437, 438 or 439. This approach is described further in PCTPublication WO 00 / 42072. Moreover, the binding sites on human IgGl for FcyRl, FcyRII, FcyRIII and FcRn have been mapped and variants with improved binding have been described (see Shields et al. (2001) J. Biol. Chem. 276:6591-6604).

[0236] In one embodiment, the Fc region is modified to decrease the ability of the antibody to mediate effector function and / or to increase anti-inflammatory properties by modifying residues 243 and 264. In one embodiment, the Fc region of the antibody or fragment thereof is modified by changing the residues at positions 243 and 264 to alanine. In one embodiment, the Fc region is modified to decrease the ability of the antibody or fragment thereof to mediate effector function and / or to increase anti-inflammatory properties by modifying residues 243, 264, 267 and 328.

[0237] The interaction between the constant region of an antigen binding protein and various Fc receptors (FcR) including Fc gammaRI (CD64), Fc gammaRII (CD32) and Fc gammaRIII (CD 16) is believed to mediate the effector functions, such as ADCC and CDC, of the antigen78NAI-5006826801V1binding protein. The Fc receptor is also important for antibody cross-linking, which can be important for anti-tumor immunity.

[0238] Effector function can be measured in a number of ways including for example via binding of the Fc gammaRIII to Natural Killer cells or via Fc gammaRI to monocytes / macrophages to measure for ADCC effector function. For example, an antigen binding protein of the present invention can be assessed for ADCC effector function in a Natural Killer cell assay. Examples of such assays can be found in Shields et al., 2001 J. Biol. Chem., Vol. 276, p 6591-6604; Chappel et al., 1993 J. Biol. Chem., Vol 268, p 25124-25131; Lazar et al., 2006 PNAS, 103; 4005-4010.

[0239] Human IgGl constant regions containing specific mutations or altered glycosylation on residue Asn297 have been shown to reduce binding to Fc receptors. In other cases, mutations have also been shown to enhance ADCC and CDC (Lazar et al., PNAS 2006, 103; 4005-4010; Shields et al. J Biol Chem 2001, 276; 6591-6604; Nechansky et al., Mol Immunol, 2007, 44; 1815-1817).

[0240] In one embodiment of the present disclosure, such mutations are in one or more of positions selected from 239, 332 and 330 (IgGl), or the equivalent positions in other IgG isotypes. Examples of suitable mutations are S239D and I332E and A330L. In one embodiment, the antigen binding protein of the invention herein described is mutated at positions 239 and 332, for example S239D and I332E or in a further embodiment it is mutated at three or more positions selected from 239 and 332 and 330, for example S239D and I332E and A330L (EU index numbering).

[0241] In an alternative embodiment, provided herein is an antibody or antigen binding fragment thereof comprising a heavy chain constant region with an altered glycosylation profile such that the antigen binding protein has enhanced effector function. For example, wherein the antibody has enhanced ADCC or enhanced CDC or wherein it has both enhanced ADCC and CDC effector function. Examples of suitable methodologies to produce antigen binding proteins with an altered glycosylation profile are described in PCT Publication Nos. W02003011878 and W02006014679 and European Patent Application No. EP1229125.

[0242] In a further aspect, the present disclosure provides “non-fucosylated” or “afucosylated” antibodies. Non-fucosylated antibodies harbor a tri-mannosyl core structure of complex-type N- glycans of Fc without fucose residue. These glycoengineered antibodies that lack core fucose residue from the Fc N-glycans may exhibit stronger ADCC than fucosylated equivalents due to enhancement of Fc gammaRIIIa binding capacity.79NAI-5006826801V1

[0243] Methods for producing an antibody or antigen binding fragment thereof may include the steps of: a) culturing a recombinant host cell comprising an expression vector comprising an isolated polynucleotide as described herein, wherein the recombinant host cell does not comprise an alpha- 1,6-fucosyltransf erase; and b) recovering the antigen binding protein. The recombinant host cell may not normally contain a gene encoding an alpha- 1,6-fucosyltransferase (for example yeast host cells such as Pichia sp.) or may have been genetically modified to inactivate an alpha- 1,6-fucosyltransferase. Recombinant host cells which have been genetically modified to inactivate the FUT8 gene encoding an alpha- 1,6-fucosyltransferase are available. See, e.g., the POTELLIGENT™ technology system available from BioWa, Inc. (Princeton, N.J.) in which CHOK1SV cells lacking a functional copy of the FUT8 gene produce monoclonal antibodies having enhanced ADCC activity that is increased relative to an identical monoclonal antibody produced in a cell with a functional FUT8 gene. Aspects of the POTELLIGENT™ technology system are described in US7214775, US6946292, W00061739 and WO0231240. Those of ordinary skill in the art will also recognize other appropriate systems.

[0244] It will be apparent to those skilled in the art that such modifications may not only be used alone but may be used in combination with each other in order to further enhance or decrease effector function.Production of Antibodies with Modified Glycosylation

[0245] In still another embodiment, the antibodies (e.g., anti-Nectin-4 antibodies) or antigenbinding fragments comprise a particular glycosylation pattern. For example, an afucosylated or an aglycosylated antibody or fragment thereof can be made (i.e., the antibody lacks fucose or glycosylation, respectively). The glycosylation pattern of an antibody or fragment thereof may be altered to, for example, increase the affinity or avidity of the antibody or fragment thereof for an antigen. Such modifications can be accomplished by, for example, altering one or more of the glycosylation sites within the antibody or fragment thereof sequence. For example, one or more amino acid substitutions can be made that result in removal of one or more of the variable region framework glycosylation sites to thereby eliminate glycosylation at that site. Such aglycosylation may increase the affinity or avidity of the antibody or fragment thereof for antigen. See, e.g., U.S. Patent Nos. 5,714,350 and 6,350,861.

[0246] Antibodies and antigen-binding fragments may further include those produced in lower eukaryote host cells, in particular fungal host cells such as yeast and filamentous fungi that have been genetically engineered to produce glycoproteins that have mammalian- or human-like glycosylation patterns (See for example, Choi et al., (2003) Proc. Natl. Acad. Sci. 100: 5022- 80NAI-5006826801V15027; Hamilton et al., (2003) Science 301 : 1244-1246; Hamilton et al., (2006) Science 313: 1441-1443; Nett et al., Yeast 28(3):237-52 (2011); Hamilton et al., Curr. Opin. Biotechnol. Oct;18(5):387-92 (2007)). A particular advantage of these genetically modified host cells over currently used mammalian cell lines is the ability to control the glycosylation profile of glycoproteins that are produced in the cells such that compositions of glycoproteins can be produced wherein a particular N-glycan structure predominates (see, e.g., U.S. Patent No. 7,029,872 and U.S. Patent No. 7,449,308). These genetically modified host cells have been used to produce antibodies that have predominantly particular N-glycan structures (See for example, Li et al., (2006) Nat. Biotechnol. 24: 210-215).

[0247] In particular embodiments, the antibodies and antigen-binding fragments thereof further include those produced in lower eukaryotic host cells and which comprise fucosylated and non-fucosylated hybrid and complex N-glycans, including bisected and multi antennary species, including but not limited to N-glycans such as GlcNAc(i-4)Man3GlcNAc2; Gal(i-4)GlcNAc(i-4)Man3GlcNAc2; or NANA(i-4)Gal(i-4)GlcNAc(i-4)Man3GlcNAc2.

[0248] In particular embodiments, the antibodies and antigen-binding fragments thereof may comprise antibodies or fragments having at least one hybrid N-glycan selected from the group consisting of GlcNAcMansGlcNAc2; GalGlcNAcMansGlcNAc2; and NANAGalGlcNAcMansGlcNAc2. In particular aspects, the hybrid N-glycan is the predominant N-glycan species in the composition.

[0249] In particular embodiments, the antibodies and antigen-binding fragments thereof comprise antibodies and fragments having at least one complex N-glycan selected from the group consisting of GlcNAcMan3GlcNAc2; GalGlcNAcMan3GlcNAc2;NANAGalGlcNAcMan3GlcNAc2; GlcNAc2Man3GlcNAc2; GalGlcNAc2Man3GlcNAc2; Gal2GlcNAc2Man3GlcNAc2; NANAGal2GlcNAc2Man3GlcNAc2; and NANA2Gal2GlcNAc2Man3GlcNAc2. In particular aspects, the complex N-glycan are the predominant N-glycan species in the composition. In further aspects, the complex N-glycan is a particular N-glycan species that comprises about 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%, 97%, 98%, 99%, or 100% of the complex N- glycans in the composition. In one embodiment, the antibody and antigen binding fragments thereof provided herein comprise complex N- glycans, wherein at least 50%, 60%, 70%, 80%, 90%, 95%, 97%, 98%, 99%, or 100% of the complex N-glycans comprise the structure NANA2Gal2GlcNAc2Man3GlcNAc2, wherein such structure is afucosylated. Such structures can be produced, e.g., in engineered Pichia pastoris host cells.

[0250] In some embodiments, the N-glycan is fucosylated. In general, the fucose is in an81NAI-5006826801V1al,3-linkage with the GlcNAc at the reducing end of the N-glycan, an al,6-linkage with the GlcNAc at the reducing end of the N-glycan, an al,2-linkage with the Gal at the non- reducing end of the N-glycan, an al,3-linkage with the GlcNac at the non-reducing end of the N-glycan, or an al,4-linkage with a GlcNAc at the non-reducing end of the N-glycan.

[0251] Therefore, in some aspects of the above the glycoprotein compositions, the glycoform is in an al,3-linkage or al,6-linkage fucose to produce a glycoform selected from the group consisting of MansGlcNAc2(Fuc), GlcNAcMansGlcNAc2(Fuc), Man3GlcNAc2(Fuc), GlcNAcMan3GlcNAc2(Fuc), GlcNAc2Man3GlcNAc2(Fuc), GalGlcNAc2Man3GlcNAc2(Fuc), Gal2GlcNAc2Man3GlcNAc2(Fuc), NANAGal2GlcNAc2Man3GlcNAc2(Fuc), and NANA2Gal2GlcNAc2Man3GlcNAc2(Fuc); in an al,3-linkage or al,4-linkage fucose to produce a glycoform selected from thegroup consisting of GlcNAc(Fuc)MansGlcNAc2, GlcNAc(Fuc)Man3GlcNAc2, GlcNAc2(Fuci- 2)Man3GlcNAc2, GalGlcNAc2(Fuci-2)Man3GlcNAc2, Gal2GlcNAc2(Fuci-2)Man3GlcNAc2, NANAGal2GlcNAc2(Fuci-2)Man3GlcNAc2, and NANA2Gal2GlcNAc2(Fuci-2)Man3GlcNAc2; or in an al,2-linkage fucose to produce a glycoform selected from the group consisting of Gal(Fuc)GlcNAc2Man3GlcNAc2, Gal2(Fuci-2)GlcNAc2Man3GlcNAc2, NANAGal2(Fuci-2)GlcNAc2Man3GlcNAc2, and NANA2Gal2(Fuci-2)GlcNAc2Man3GlcNAc2.

[0252] In further examples, the antibodies (e.g., humanized antibodies) or antigen -binding fragments thereof may comprise high mannose N-glycans, including but not limited to, ManxGlcNAc2, Man?GlcNAc2, ManeGlcNAc2, MansGlcNAc2, Man4GlcNAc2, or N-glycans that consist of the Man3GlcNAc2N-glycan structure.

[0253] In further aspects of the above, the complex N-glycans may further include fucosylated and non-fucosylated bisected and multi antennary species.Antibody Physical Properties

[0254] The antibodies and antigen-binding fragments thereof disclosed herein may further contain one or more glycosylation sites in either the light or heavy chain immunoglobulin variable region. Such glycosylation sites may result in increased immunogenicity of the antibody or fragment thereof or an alteration of the pK of the antibody due to altered antigenbinding (Marshall et al. (1972) Annu Rev Biochem 41 :673-702; Gala and Morrison (2004) J Immunol 172:5489-94; Wallick et al. (1988) J Exp Med 168: 1099-109; Spiro (2002) Glycobiology 12:43R-56R; Parekh et al. (1985) Nature 316:452-7; Mimura et al. (2000) Mol Immunol 37:697-706). Glycosylation has been known to occur at motifs containing an N-X-S / T sequence.82NAI-5006826801V1

[0255] Each antibody or antigen-binding fragment thereof has a unique isoelectric point (pl), which generally falls in the pH range between 6 and 9.5. The pl for an IgGl antibody typically falls within the pH range of 7-9.5 and the pl for an IgG4 antibody typically falls within the pH range of 6-8.

[0256] Each antibody or antigen-binding fragment thereof has a characteristic melting temperature, with a higher melting temperature indicating greater overall stability in vivo (Krishnamurthy R and Manning MC 2002 Curr Pharm Biotechnol 3:361-71). In general, the TMi (the temperature of initial unfolding) may be greater than 60 °C, greater than 65 °C, or greater than 70 °C. The melting point of an antibody or fragment thereof can be measured using differential scanning calorimetry (Chen et al., 2003 Pharm Res 20: 1952-60; Ghirlando et al., 1999 Immunol Lett 68:47-52) or circular dichroism (Murray et al., 2002 J. Chromatogr Sci 40:343-9).

[0257] In a further example, the antibodies and antigen-binding fragments thereof do not degrade rapidly. Degradation of an antibody or fragment thereof can be measured using capillary electrophoresis (CE) and MALDI-MS (Alexander AJ and Hughes DE (1995) Anal Chem 67:3626-32).

[0258] In a further example, the antibodies and antigen-binding fragments thereof have minimal aggregation effects, which can lead to the triggering of an unwanted immune response and / or altered or unfavorable pharmacokinetic properties. Generally, antibodies and fragments are acceptable with aggregation of 25% or less, 20% or less, 15% or less, 10% or less, or 5% or less. Aggregation can be measured by several techniques, including size-exclusion column (SEC), high performance liquid chromatography (HPLC), and light scattering.Antibody Conjugates

[0259] The antibodies (e.g., anti-Nectin-4 antibodies) and antigen-binding fragments thereof disclosed herein may be conjugated to a chemical moiety. The chemical moiety may be, inter alia, a polymer, a radionucleotide, a therapeutic agent, or a cytotoxic factor. In some embodiments, the chemical moiety is a polymer which increases the half-life of the antibody or fragment thereof in the body of a subject. Suitable polymers include, but are not limited to, hydrophilic polymers which include but are not limited to polyethylene glycol (PEG) (e.g., PEG with a molecular weight of 2kDa, 5 kDa, 10 kDa, 12kDa, 20 kDa, 30kDa or 40kDa), dextran and monomethoxypolyethylene glycol (mPEG). Lee, et al., (1999) (Bioconj. Chem. 10:973-981) discloses PEG conjugated single-chain antibodies. Wen, et al., (2001) (Bioconj. Chem. 12:545- 553) disclose conjugating antibodies with PEG which is attached to a radiometal chelator (diethylenetriaminpentaacetic acid (DTP A)).83NAI-5006826801V1

[0260] The antibodies and antigen-binding fragments disclosed herein may be PEGylated, for example to increase its biological (e.g., serum) half-life. To PEGylate an antibody or fragment thereof, the antibody or fragment thereof, typically is reacted with a reactive form of PEG, such as a reactive ester or aldehyde derivative of PEG, under conditions in which one or more PEG groups become attached to the antibody or antibody fragment thereof. In some embodiments, the PEGylation is carried out via an acylation reaction or an alkylation reaction with a reactive PEG molecule (or an analogous reactive water-soluble polymer). In certain embodiments, the antibody or fragment thereof to be PEGylated is an aglycosylated antibody or fragment thereof. Methods for PEGylating proteins are known in the art and can be applied to the antibodies of the invention. See, e.g., EP 0 154 316 and EP 0 401 384.

[0261] The antibodies and antigen-binding fragments thereof disclosed herein may be conjugated with labels such as "Tc,90Y,n iIn,32P,14C,1251,3H,131I,nC,15O,13N,18F,35S,51Cr,57TO,226Ra,60Co,59Fe,57Se,152Eu,67CU,217Ci,211At,212Pb,47Sc,109Pd,234Th,40K,157Gd,55Mn,52Tr, and / or56Fe.

[0262] The antibodies and antigen-binding fragments disclosed herein may be conjugated with fluorescent or chemilluminescent labels, including fluorophores such as rare earth chelates, fluorescein and its derivatives, rhodamine and its derivatives, isothiocyanate, phycoerythrin, phycocyanin, allophycocyanin, o-phthaladehyde, fluorescamine,152Eu, dansyl, umbelliferone, luciferin, luminal label, isoluminal label, an aromatic acridinium ester label, an imidazole label, an acridimium salt label, an oxalate ester label, an aequorin label, 2,3-dihydrophthalazinediones, biotin / avidin, spin labels, and stable free radicals.

[0263] The antibodies and antigen-binding fragments thereof also be conjugated to a cytotoxic factor such as diptheria toxin, Pseudomonas aeruginosa exotoxin A chain, ricin A chain, abrin A chain, modeccin A chain, alpha-sarcin, Aleurites fordii proteins and compounds (e.g., fatty acids), dianthin proteins, Phytoiacca americana proteins PAPI, PAPII, and PAP-S, Momordica charantia inhibitor, curcin, crotin, Saponaria officinalis inhibitor, mitogellin, restrictocin, phenomycin, and enomycin.

[0264] The antibodies and antigen-binding fragments thereof also be conjugated to a therapeutic agent such as a molecule with anti-tumor biological activity. A molecule with antitumor biological activity includes, but is not limited to, a cytotoxic agent, chemotherapeutic agent, radioisotope, immune checkpoint inhibitor, antibody targeting tumor-specific antigen, and other anti-tumor drugs. In some embodiments, the therapeutic agent is a radioisotope. In some embodiment, the therapeutic agent is a cytotoxic agent.84NAI-5006826801V1

[0265] In a specific embodiment, the therapeutic agent is,

[0266] The therapeutic agent can be conjugated to the antibody or antigen-binding fragment of the present invention through a linker using any technique known in the art. The linker may comprise a reactive group for covalent conjugation, such as amine, hydroxylamine, maleimide, carboxyl, phenyl, thiol, sulfhydryl, or hydroxyl group. The linker can be cleavable or non- cleavable. The cleavable linker is, for example, an enzymatically cleavable linker (e.g., peptide containing protease cleavage site), pH sensitive linker (e.g., hydrazone-type linker), or reducible linker (e.g., disulfide bond).

[0267] In one embodiment, the linker comprises a reactive group selected from the group consisting of amine, hydroxylamine, maleimide, carboxyl, phenyl, thiol, sulfhydryl, and hydroxyl groups. In one embodiment, the linker is a chemical bond. In one embodiment, the linker comprises an amino acid or a peptide consisting of 2-10 amino acids. The amino acid can be a natural or non-natural amino acid.

[0268] In another embodiment, the linker is selected from the group consisting of Vai, Cit,Phe, Lys, D-Val, Leu, Gly, Ala, Asn,85NAI-5006826801V1wherein one of position 1 and position 2 represents a position at which the linker is attached to the antibody or antigen-binding fragment thereof of the present invention, and the other is a position at which the linker is attached to the therapeutic agent.

[0269] In some embodiments, the therapeutic agent and the linker are conjugated to form an intermediate prior to conjugation to the antibody or antigen-binding fragment of the present invention. In some embodiments, the intermediate is linked by forming a thioether bond with a sulfhydryl group on the antibody or antigen-binding fragment of the present invention. The structure and preparation method of such intermediate and the method of using the same to prepare antibody conjugate are described, for example, in the international patent application publication WO2019 / 114666, and the relevant contents of which are incorporated herein by reference in their entirety.

[0270] In one embodiment, the antibody conjugate has the structure of Formula (I), A-(L-D)y(i); wherein, A is the antibody or antigen-binding fragment thereof of the present invention;L is a linker; preferably, L comprises an amino acid or a peptide consisting of 2-10 amino acids; further preferably, L is selected from Vai, Cit, Phe, Lys, D-Val, Leu, Gly, Ala, Asn,86NAI-5006826801V1position 1 and position 2 represents a position where L and A are connected, and the other represents a position where L and D are connected;D is a therapeutic agent; preferably, D is selected from the group consisting of cytotoxic agents, therapeutic antibodies, radioisotopes, oligonucleotides and analogs thereof, biologically87NAI-5006826801V1active peptides, protein toxins and enzymes; more preferably, D is an anti-tumor biologicallyand y represents the number of (L-D) moi eties linked to A, which is an integer from 1 to 10.

[0271] In a specific embodiment, the antibody conjugate is formed by connecting the (L-D) moiety of Formula (1) with a sulfhydryl group of A, for example, the sulfhydryl group of the side chain of a cysteine residue of A, through a thioether bond,88NAI-5006826801V1Formula (1), wherein the antibody conjugate of the present invention has the structure of formula (2):wherein A is the antibody or antigen-binding fragment of the present invention, and y represents the number of (L-D) moi eties linked to A by forming a thioether bond with the sulfhydryl group of A, which is an integer from 1 to 10. In a specific embodiment, A is an anti -human Nectin-4 humanized monoclonal antibody of the present invention.

[0272] In yet another aspect, the present invention provides a composition, which comprises the antibody conjugate of the present invention. In one embodiment, the molar ratio (DAR value) of the therapeutic agent to the anti-Nectin-4 antibody or antigen-binding fragment thereof of the present invention in the composition is a decimal or integer between 1 and 10, such as a decimal or integer between 1 and 8, such as 1.0, 1.5, 2.0, 2.5, 3.0, 3.1, 3.2, 3.3, 3.4, 3.5, 3.6, 3.7, 3.79, 3.8, 3.9, 4.0, 4.1, 4.2, 4.3, 4.4, 4.5, 4.6, 4.7, 4.8, 4.9, 5.0, 5.1, 5.2, 5.3, 5.4, 5.5, 5.6, 5.7, 5.8, 5.9, 6.0, 6.1, 6.2, 6.3, 6.4, 6.5, 6.6, 6.7, 6.8, 6.9, 6.95, 7.0, 7.03, 7.1, 7.12, 7.2, 7.3, 7.4, 7.5, 7.6, 7.7, 7.8, 7.9 or 8.0.

[0273] Any method known in the art for conjugating the antibodies and antigen-binding fragments thereof of the invention to the various moieties may be employed, including those methods described by Hunter, et al., (1962) Nature 144:945; David, et al., (1974) Biochemistry 13: 1014; Pain, et al., (1981) J. Immunol. Meth. 40:219; and Nygren, J., (1982) Histochem. and Cytochem. 30:407.89NAI-5006826801V1Pharmaceutical Compositions and Administration

[0274] To prepare pharmaceutical or sterile compositions of the compositions disclosed herein (e.g., anti-Nectin-4 antibodies and antigen-binding fragments thereof, or chemotherapeutic agents), the composition may be admixed with a pharmaceutically acceptable carrier or excipient. See, e.g., Remington's Pharmaceutical Sciences and U.S. Pharmacopeia: National Formulary, Mack Publishing Company, Easton, PA (1984).

[0275] Formulations of therapeutic and diagnostic agents may be prepared by mixing with acceptable carriers, excipients, or stabilizers in the form of, e.g., lyophilized powders, slurries, aqueous solutions or suspensions. (See, e.g., Hardman, et al. (2001) Goodman and Gilman’s The Pharmacological Basis of Therapeutics, McGraw-Hill, New York, NY; Gennaro (2000) Remington: The Science and Practice of Pharmacy, Lippincott, Williams, and Wilkins, New York, NY; Avis, et al. (eds.) (1993) Pharmaceutical Dosage Forms: Parenteral Medications, Marcel Dekker, NY; Lieberman, et al. (eds.) (1990) Pharmaceutical Dosage Forms: Tablets, Marcel Dekker, NY; Lieberman, et al. (eds.) (1990) Pharmaceutical Dosage Forms: Disperse Systems, Marcel Dekker, NY; Weiner and Kotkoskie (2000) Excipient Toxicity and Safety, Marcel Dekker, Inc., New York, NY)

[0276] Toxicity and therapeutic efficacy of the therapeutic agents disclosed herein, administered alone or in combination with another therapeutic agent, can be determined by standard pharmaceutical procedures in cell cultures or experimental animals, e.g., for determining the LD50 (the dose lethal to 50% of the population) and the ED50 (the dose therapeutically effective in 50% of the population). The dose ratio between toxic and therapeutic effects is the therapeutic index (LD50 / ED50). The data obtained from these cell culture assays and animal studies can be used in formulating a range of dosage for use in human. The dosage of such compounds lies preferably within a range of circulating concentrations that include the ED50 with little or no toxicity. The dosage may vary within this range depending upon the dosage form employed and the route of administration.

[0277] Any suitable route of administration may be used. Routes of administration (e.g., for an anti-Nectin-4 antibody, antigen binding fragment thereof, or antibody conjugate) include oral, rectal, transmucosal, intestinal, parenteral; intramuscular, subcutaneous, intradermal, intramedullary, intrathecal, direct intraventricular, intravenous, intraperitoneal, intranasal, intratumor, intraocular, inhalation, insufflation, topical, cutaneous, transdermal, or intra-arterial. For example, the antibody or antigen binding fragment thereof may be administered intravenously or subcutaneously. In one example, the anti-Nectin-4 antibody, antigen binding90NAI-5006826801V1fragment thereof, or antibody conjugate is administered intravenously by infusion. The administration may be systemic or local.

[0278] A vessel (e.g., a plastic or glass vial, a hollow bore needle, or a syringe cylinder) may be used that contains any of the therapeutic agents disclosed herein (e.g., an anti-Nectin-4 antibody, antigen binding fragment thereof, or antibody conjugate, and / or one or more chemotherapeutic agents) or a pharmaceutical composition thereof.

[0279] The present disclosure also provides an injection device comprising any of the therapeutic agents disclosed herein (e.g., an anti-Nectin-4 antibody, antigen binding fragment thereof, or antibody conjugate, and / or one or more chemotherapeutic agents) or a pharmaceutical composition thereof. An injection device is a device that introduces a substance into the body of a patient via a parenteral route, e.g., intramuscular, subcutaneous or intravenous. For example, an injection device may be a syringe (e.g., pre-filled with the pharmaceutical composition, such as an auto-injector) which, for example, includes a cylinder or barrel for holding fluid to be injected (e.g., antibody or fragment thereof or a pharmaceutical composition thereof), a needle for piecing skin and / or blood vessels for injection of the fluid; and a plunger for pushing the fluid out of the cylinder and through the needle bore. In an embodiment, an injection device that comprises an antibody, antigen-binding fragment thereof, or antibody conjugate of the present disclosure or a pharmaceutical composition thereof is an intravenous (IV) injection device. Such a device includes the antibody or fragment thereof or a pharmaceutical composition thereof in a cannula or trocar / needle which may be attached to a tube which may be attached to a bag or reservoir for holding fluid (e.g., saline; or lactated ringer solution comprising NaCl, sodium lactate, KC1, CaCh and optionally including glucose) introduced into the body of the patient through the cannula or trocar / needle. The antibody or fragment thereof or a pharmaceutical composition thereof may be introduced into the device once the trocar and cannula are inserted into the vein of a subject and the trocar is removed from the inserted cannula. The IV device may, for example, be inserted into a peripheral vein (e.g., in the hand or arm); the superior vena cava or inferior vena cava, or within the right atrium of the heart (e.g., a central IV); or into a subclavian, internal jugular, or a femoral vein and, for example, advanced toward the heart until it reaches the superior vena cava or right atrium (e.g., a central venous line). In an embodiment, an injection device is an autoinjector; a jet injector or an external infusion pump. A jet injector uses a high-pressure narrow jet of liquid which penetrate the epidermis to introduce the antibody or fragment thereof or a pharmaceutical composition thereof to a patient’s body. External infusion pumps are medical devices that deliver the antibody or fragment thereof or a pharmaceutical composition thereof into a patient’s91NAI-5006826801V1body in controlled amounts. External infusion pumps may be powered electrically or mechanically. Different pumps operate in different ways, for example, a syringe pump holds fluid in the reservoir of a syringe, and a moveable piston controls fluid delivery, an elastomeric pump holds fluid in a stretchable balloon reservoir, and pressure from the elastic walls of the balloon drives fluid delivery. In a peristaltic pump, a set of rollers pinches down on a length of flexible tubing, pushing fluid forward. In a multi-channel pump, fluids can be delivered from multiple reservoirs at multiple rates.

[0280] The pharmaceutical compositions disclosed herein may also be administered with a needleless hypodermic injection device; such as the devices disclosed in U.S. Patent Nos. 6,620,135; 6,096,002; 5,399,163; 5,383,851; 5,312,335; 5,064,413; 4,941,880; 4,790,824 or 4,596,556. Such needleless devices comprising the pharmaceutical composition are also part of the present disclosure. The pharmaceutical compositions disclosed herein may also be administered by infusion. Examples of well-known implants and modules for administering the pharmaceutical compositions include those disclosed in: U.S. Patent No. 4,487,603, which discloses an implantable micro-infusion pump for dispensing medication at a controlled rate; U.S. Patent No. 4,447,233, which discloses a medication infusion pump for delivering medication at a precise infusion rate; U.S. Patent No. 4,447,224, which discloses a variable flow implantable infusion apparatus for continuous drug delivery; U.S. Patent. No. 4,439,196, which discloses an osmotic drug delivery system having multi-chamber compartments. Many other such implants, delivery systems, and modules are well known to those skilled in the art and those comprising the pharmaceutical compositions of the present invention are within the scope of the present invention.

[0281] Alternately, one may administer the therapeutic agent (e.g., an anti-Nectin-4 antibody, antigen binding fragment thereof, or antibody conjugate, and / or one or more chemotherapeutic agents) in a local rather than systemic manner, for example, via injection of the antibody or fragment thereof directly into a tumor, e.g., a Nectin-4+ tumor. Furthermore, one may administer the antibody or fragment thereof in a targeted drug delivery system, for example, in a liposome coated with a tissue-specific antibody, targeting, for example, a tumor e.g., a Nectin- 4+ tumor, e.g., characterized by immunopathology. The liposomes will be targeted to and taken up selectively by the afflicted tissue. Such methods and liposomes are part of the present disclosure.

[0282] With respect to antibodies, antigen-binding fragments thereof, or antibody conjugates, the administration regimen may depend on several factors, including the serum or tissue turnover rate of the therapeutic antibody, antigen-binding fragment thereof, or antibody92NAI-5006826801V1conjugate, the level of symptoms, the immunogenicity of the therapeutic antibody, and the accessibility of the target cells in the biological matrix. Preferably, the administration regimen delivers sufficient therapeutic antibody, antigen-binding fragment thereof, or antibody conjugate to effect improvement in the target disease state, while simultaneously minimizing undesired side effects. Accordingly, the amount of biologic delivered depends in part on the particular therapeutic antibody and the severity of the condition being treated. Guidance in selecting appropriate doses of therapeutic antibodies or fragments is available. (See, e.g., Wawrzynczak (1996) Antibody Therapy, Bios Scientific Pub. Ltd, Oxfordshire, UK; Kresina (ed.) (1991) Monoclonal Antibodies, Cytokines and Arthritis, Marcel Dekker, New York, NY; Bach (ed.) (1993) Monoclonal Antibodies and Peptide Therapy in Autoimmune Diseases, Marcel Dekker, New York, NY; Baert, et al. (2003) New Engl. J. Med. 348:601-608; Milgrom et al. (1999) New Engl. J. Med. 341 : 1966-1973; Slamon et al. (2001) New Engl. J. Med. 344:783-792;Beniaminovitz et al. (2000) New Engl. J. Med. 342:613-619; Ghosh et al. (2003) New Engl. J. Med. 348:24-32; Lipsky et al. (2000) New Engl. J. Med. 343: 1594-1602) In various embodiments, the patient is provided a short, repeated exposure of the antibody or antigen binding fragment thereof rather than continuous exposure. Alternatively, the patient is provided a continuous exposure.

[0283] Antibodies, antigen binding fragments, and antibody conjugates thereof disclosed herein may be provided by continuous infusion, or by doses administered, e.g., daily, 1-7 times per week, weekly, bi-weekly, every three weeks, monthly, every five weeks, every six weeks, every seven weeks, bimonthly, quarterly, semiannually, or annually. Doses may be provided, e.g., intravenously, subcutaneously, topically, orally, nasally, rectally, intramuscular, intracerebrally, intraspinally, or by inhalation, (see, e.g., Yang, et al., 2003 New Engl. J. Med. 349:427-434; Herold, et al., 2002 New Engl. J. Med. 346: 1692-1698; Liu, et al., 1999 J. Neurol. Neurosurg. Psych. 67:451-456; Portielji, et al., 2003 Cancer Immunol. Immunother. 52: 151- 144). Doses may also be provided to achieve a pre-determined target concentration of an anti- Nectin-4 antibody, antigen binding fragment thereof, or antibody conjugate in the subject’s serum.

[0284] Also provided herein is a pharmaceutical composition for treating a cancer in a patient, the pharmaceutical composition comprising an anti-Nectin-4 antibody, antigen binding fragment thereof, or antibody conjugate, wherein the therapeutic is for use in combination with another therapeutic agent.93NAI-5006826801V1Kits

[0285] Further provided herein are kits comprising one or more components that include, but are not limited to an anti-Nectin-4 antibody, antigen binding fragment thereof, or antibody conjugate, as discussed herein alone or in association with one or more additional components including, but not limited to a pharmaceutically acceptable carrier and / or one or more additional therapeutic agents, e.g., one or more chemotherapeutic agents. The anti-Nectin-4 antibody, antigen binding fragment thereof, or antibody conjugate and / or the one or more additional therapeutic agents can be formulated as a pure composition or in combination with a pharmaceutically acceptable carrier, in a pharmaceutical composition.

[0286] In one embodiment, the kit includes an anti-Nectin-4 antibody, antigen binding fragment thereof, or antibody conjugate or a pharmaceutical composition thereof in one container (e.g., in a sterile glass or plastic vial) and / or one or more additional therapeutic agents (e.g., one or more chemotherapeutic agents) or a pharmaceutical composition thereof in another container (e.g., in a sterile glass or plastic vial).

[0287] In another embodiment, the kit comprises an anti-Nectin-4 antibody, antigen binding fragment thereof, or antibody conjugate of the invention along with a pharmaceutically acceptable carrier, optionally in combination with one or more therapeutic agents (e.g., one or more chemotherapeutic agents) formulated together, optionally, in a pharmaceutical composition, in a single, common container.

[0288] If the kit includes a pharmaceutical composition for parenteral administration to a subject, the kit can include a device for performing such administration. For example, the kit can include one or more hypodermic needles or other injection devices as discussed herein.

[0289] The kit can include a package insert including information concerning the pharmaceutical compositions and dosage forms in the kit. Generally, such information aids patients and physicians in using the enclosed pharmaceutical compositions and dosage forms effectively and safely. For example, the following information regarding a therapeutic agent of the disclosure may be supplied in the insert: pharmacokinetics, pharmacodynamics, clinical studies, efficacy parameters, indications and usage, contraindications, warnings, precautions, adverse reactions, overdosage, proper dosage and administration, how supplied, proper storage conditions, references, manufacturer / distributor information and patent information.

[0290] Containers for use in such kits may typically comprise at least one vial, test tube, flask, bottle, syringe or other suitable container, into which one or more of the detection and / or therapeutic composition(s) may be placed, and preferably suitably aliquoted. Where one or more additional therapeutic agents is also provided, the kit may also contain one or more additional94NAI-5006826801V1distinct containers into which the one or more additional therapeutic agents may be placed. Alternatively, a plurality of compounds may be prepared in a single pharmaceutical composition, and may be packaged in a single container means, such as a vial, flask, syringe, bottle, or other suitable single container. The kits disclosed herein will also typically include a means for containing the vial(s) in close confinement for commercial sale, such as, e.g., injection or blow-molded plastic containers into which the desired vial(s) are retained.

[0291] Any of the kits provided herein may further include a diagnostic anti-Nectin-4 antibody. Any suitable diagnostic anti-Nectin-4 antibody may be included.

[0292] Any of the kits disclosed herein may further include one or more reagents for a diagnostic assay, e.g., an IHC assay for detection of Nectin-4. The reagent may be a detection reagent.

[0293] Any of the kits disclosed herein may further include a radiolabel, chromogenic, fluorogenic, or other type of detectable label or detecting means, which optionally may be conjugated to the diagnostic anti-Nectin-4 antibody.

[0294] Where a radiolabel, chromogenic, fluorogenic, or other type of detectable label or detecting means is included within the kit, the labeling agent may be provided either in the same container as the detection or therapeutic composition itself or may alternatively be placed in a second distinct container means into which this second composition may be placed and suitably aliquoted. Alternatively, the detection reagent and the label may be prepared in a single container means, and in most cases, the kit will also typically include a means for containing the vial(s) in close confinement for commercial sale and / or convenient packaging and delivery.

[0295] A device or apparatus for carrying out the detection or monitoring methods described herein is also provided. Such an apparatus may include a chamber or tube into which sample can be input, a fluid handling system optionally including valves or pumps to direct flow of the sample through the device, optionally filters to separate plasma or serum from blood, mixing chambers for the addition of capture agents or detection reagents, and optionally a detection device for detecting the amount of detectable label bound to the capture agent immunocomplex. The flow of sample may be passive (e.g., by capillary, hydrostatic, or other forces that do not require further manipulation of the device once sample is applied) or active (e.g., by application of force generated via mechanical pumps, electroosmotic pumps, centrifugal force, or increased air pressure), or by a combination of active and passive forces.

[0296] In further embodiments, also provided is a processor, a computer readable memory, and a routine stored on the computer readable memory and adapted to be executed on the processor to perform any of the methods described herein. Examples of suitable computing95NAI-5006826801V1systems, environments, and / or configurations include personal computers, server computers, hand-held or laptop devices, multiprocessor systems, microprocessor-based systems, set top boxes, programmable consumer electronics, network PCs, minicomputers, mainframe computers, distributed computing environments that include any of the above systems or devices, or any other systems known in the art.EXAMPLES

[0297] The following examples are meant to be illustrative and should not be construed as further limiting. The contents of the figures and all references, patents, and published patent applications cited throughout this application are expressly incorporated herein by reference.Example 1: Generation of anti-human Nectin-4 ADCs

[0298] The generation and characterization of anti-human Nectin-4 antibodies and ADC molecules is described in WO2022 / 057651A1, the relevant content of which is incorporated herein by reference in its entirety. The amino acid sequence composition of anti-human Nectin-4 antibodies is shown in Table 3 below.Table 3. Anti-human Nectin-4 antibodies96NAI-5006826801V197NAI-5006826801V198NAI-5006826801V1

[0299] Anti-human Nectin-4 ADCs 31HZ-TL001, 56HZ-TL001, and 74HZ-TL001 were prepared by coupling anti-human Nectin-4 humanized monoclonal antibodies comprising VH and VL amino acid sequences set forth in SEQ ID NOs 55-60 with a TL001 compound comprising the structure shown in Formula (1):Formula (1).

[0300] The anti-human Nectin-4 reference ADC Enfortumab-TLOOl was prepared by coupling the anti-human Nectin-4 antibody Enfortumab, described in WO2012 / 047724, with the TL001 compound comprising the structure shown in Formula (1). The preparation method of TL001 is described in WO2019 / 114666, the relevant content of which is incorporated herein by reference in its entirety.

[0301] The preparation method of anti -human Nectin-4 antibody conjugates, / .<?., anti-human Nectin-4 ADCs, was as follows.

[0302] (1) Coupling: 30 mg of anti-human Nectin-4 humanized monoclonal antibody was diluted with a diluent (20 mM PB+105 mM NaCl, pH 7.7) and a 5 mM edetate sodium solution was added and mixed well. A TCEP solution was added (the molar ratio of TCEP to the antibody was 4.4: 1), mixed well, and incubated at room temperature for 30 minutes. TL001 (the molar ratio of TL001 to the antibody was 9: 1) was dissolved in dimethyl sulfoxide and added to the above antibody solution, mixed well, and incubated at room temperature for 2 hours to obtain the coupled samples, which were named as 31HZ-TL001, 56HZ-TL001, 74HZ-TL001, and Enfortumab-TLOOl, respectively. The anti -human Nectin-4 ADCs were subjected to buffer exchange and then stored in aliquots at 4°C.

[0303] (2) Detection: Molecular weight of anti-human Nectin-4 ADCs was analyzed by LC- MS under the following conditions.Chromatography conditionsLiquid chromatography column: Thermo MAbPac RP 3.0x 100 mm;Mobile phase A: 0.1% FA / 98% H2O / 2% ACN;Mobile phase B: 0.1% FA / 2% H2O / 98% ACN;99NAI-5006826801V1Flow rate: 0.25 mL / min; sample chamber temperature: 8°C; column temperature: 60°C; sample size: 1 pL;Switching valve: 0-3 min to waste, 3-22 min to MS, 22-30 min to waste.Mass spectrometry conditionsMass spectrometer model: AB Sciex Triple TOF 5600+;Parameters: GS1 35; GS2 35; CUR 30; TEM 350; ISVF 5500; DP 200; CE 10; m / z 600-4000; Time bins to sum 40.

[0304] The theoretical molecular weight and the measured molecular weight of the light chain and heavy chain of the coupled anti -human 31HZ-TL001 (the heavy chain was calculated by the main glycoform G0F) are shown in the following table:

[0305] The light chain of 31HZ-TL001 coupled with 0 to 1 toxin (the proportions of LC and DARI were 0.1% and 99.9%, respectively), and the heavy chain coupled with 0 to 4 toxins (the proportions of HC, DARI, DAR2, DAR3, and DAR4 were 0.0%, 2.0%, 10.2%, 86.1%, and 1.8%, respectively), the drug-antibody ratio (DAR) of toxin to antibody was therefore calculated to be 7.75, according to the calculation formula: DAR = light chain DARI x 2 + heavy chain (DARI x 1 + DAR.2 x 2 + DAR3 x 3 + DAR4 x 4) x 2.

[0306] By using the same method, the detected DAR values of 56HZ-TL001, 74HZ-TL001, and Enfortumab-TLOOl were 7.71, 7.98, and 8.01, respectively.

[0307] According to the above analysis results of mass spectrometry, the structure of antihuman Nectin-4 ADC should be determined as:100NAI-5006826801V1

[0308] wherein A represents an anti-human Nectin-4 humanized monoclonal antibody; and y refers to the number of TL001 linked to A by forming thioether bond with the thiol group of A, and is an integer of 1 to 10.

[0309] Enfortumab-VC-MMAE was prepared by coupling VC-MMAE with Enfortumab. The preparation and detection methods were as described above, and the molar ratio of antibody to VC-MMAE was 1 :5. The DAR value of this batch of Enfortumab-VC-MMAE was 5.09.Example 2: Detection of killing activity of anti-human Nectin-4 ADCs on a tumor cell line with high expression of human Nectin-4

[0310] The A549-Nectin-4 stable cell line was selected to detect the killing activity of 31HZ- TL001, 74HZ-TL001, and Enfortumab-TLOOl on a cell line with high expression of human Nectin-4. The specific experimental steps were as follows. A549-Nectin-4 cells were digested, collected by centrifugation, resuspended to 100,000 / mL with DMEM+4% FBS medium, and plated at 100 pL / well cells into a 96-well plate. ADC molecules were diluted by gradient with DMEM basal medium, starting from a final concentration of 150 pg / mL (4-fold dilution, 11 concentration points) and added to corresponding wells (100 pL per well, 2% final serum concentration). Incubation was performed in a 37°C, 5% CO2 incubator for 72 hours. CCK8 (Rhinogen) was added, 20 pL / well, and incubated for 0.5-2.5 hours in a 37°C, 5% CO2 incubator. OD450 nm readings were obtained on a microplate reader (MD) every half hour and imported into Graphpad Prism for curve fitting.

[0311] The experimental results are shown in FIGs. 1A-1B, indicating that 31HZ-TL001, 74HZ-TL001, and Enfortumab-TLOOl all could effectively kill A549-Nectin-4 cells, with EC50 values of 1390 ng / mL, 1364 ng / mL, and 2049 ng / mL, respectively. The killing activities of 31HZ-TL001 and 74HZ-TL001 on tumor cell lines with high expression of human Nectin-4 were better than that of Enfortumab-TLOOl. Meanwhile, the killing activity EC50 values of 31HZ-TL001, 74HZ-TL001, and Enfortumab-TLOOl on A549 cells without overexpressing human Nectin-4 were 5982 ng / mL, 6251 ng / mL, and 5604 ng / mL, respectively. Overall, in terms of killing activity on A549-Nectin-4 cells and A549 cells, 31HZ-TL001, 74HZ-TL001, and Enfortumab-TLOOl are Nectin-4 specific.101NAI-5006826801V1

[0312] The killing activities of 31HZ-TL001 and 56HZ-TL001 on T24-Nectin-4 cell line with high expression of human Nectin-4 were detected by the same experimental method described above. The experimental results were shown in FIGs. 1C-1D, which showed that both 31HZ- TL001 and 56HZ-TL001 could effectively kill T24-Nectin-4 cells, with EC50 values of 1277 ng / mL and 1568 ng / mL, respectively. This indicated that the killing activities of the two ADCs were substantially equivalent. The killing activity EC50 values of 31HZ-TL001 and 56HZ- TL001 on T24 cells without overexpressing human Nectin-4 were 2884 ng / mL and 3665 ng / mL, respectively, and these killing activities were significantly weaker than those on T24-Nectin-4 cells, indicating that the killing effect of both was Nectin-4 specific.Example 3: Detection of killing activity of anti-human Nectin-4 ADCs on tumor cell lines endogenously expressing human Nectin-4

[0313] The T47D tumor cell line was selected to detect the killing activity of 31HZ-TL001, 74HZ-TL001, and Enfortumab-TLOOl on tumor cell line endogenously expressing human Nectin-4. The specific experimental steps were as follows. T47D cells were digested, collected by centrifugation, resuspended to 100,000 / mL in 1640+4% FBS medium, and plated at 100 pL / well cells into a 96-well plate. ADC molecules were diluted by gradient with 1640 basal medium, starting at a final concentration of 150 pg / mL (4-fold dilution, 11 concentration points) and added to the corresponding wells (100 pL per well, 2% final serum concentration).Incubation was performed in a 37°C, 5% CO2 incubator for 72 hours. CCK8 (Rhinogen) was added, 20 pL / well, and incubated for 0.5-2.5 hours in a 37°C, 5% CO2 incubator. OD450 nm readings were collected on a microplate reader (MD) every half hour and imported into Graphpad Prism for curve fitting.

[0314] The experimental results are shown in FIG. 2A, indicating that 31HZ-TL001, 74HZ- TL001 and Enfortumab-TLOOl could effectively kill T47D cells, with EC50 values of 206.1 ng / mL, 611.3 ng / mL and 229.2 ng / mL, respectively. The results showed that the killing activity of 31HZ-TL001 on T47D cells was comparable to that of Enfortumab-TLOOl.

[0315] The killing activity of 31HZ-TL001, 74HZ-TL001, and Enfortumab-TLOOl on the tumor cell line MDA-MB-468 endogenously expressing human Nectin-4 was detected by the same experimental method described above. The experimental results were shown in FIG. 2B, which showed that 31HZ-TL001, 74HZ-TL001, and Enfortumab-TLOOl could effectively kill MDA-MB-468 cells, and their EC50 values were 1030 ng / mL, 2277 ng / mL, and 1190 ng / mL, respectively, indicating that the killing activity of 31HZ-TL001 on MDA-MB-468 cells was comparable to that of Enfortumab-TLOOl.102NAI-5006826801V1

[0316] The killing activity of 31HZ-TL001 and 56HZ-TL001 on the tumor cell line NCI-N87 endogenously expressing human Nectin-4 was detected by the same experimental method as described above. The experimental results were shown in FIG. 2C, which showed that both 31HZ-TL001 and 56HZ-TL001 could effectively kill NCI-N87 cells, and their ECso values were 9376 ng / mL and 10935 ng / mL, respectively, indicating that the killing activities of the two were substantially equivalent.

[0317] In conclusion, the killing activities of 31HZ-TL001 and 56HZ-TL001 on tumor cell lines endogenously expressing human Nectin-4 were substantially equivalent to those of Enfortumab-TLOO 1.Example 4: In vivo pharmacodynamics of anti-human Nectin-4 ADCs in a CDX model

[0318] The anti-tumor effect of ADC molecules was evaluated by constructing a CDX model of the human non-small cell lung cancer cell line NCI-H322M. The specific steps were as follows. NCI-H322M cells were cultured with RPMI1640 medium containing 10% fetal bovine serum (FBS) at 37°C and 5% CO2. The cells were collected in exponential growth phase, resuspended in PBS, and inoculated subcutaneously into female NOD / SCID mice (Biocytogen Jiangsu Gene Biotechnology Co., Ltd.) at l * 107 / mouse (suspended in 0.1 mL of PBS) to establish a subcutaneous xenograft tumor model. When the average tumor volume reached 70- 100 mm3, the mice were randomized according to tumor volume with 5 mice in each group. The day of grouping was recorded as Day 0 and the groups included a vehicle group (normal saline; negative control), 31HZ-TL001 groups (3 mg / kg and 10 mg / kg), 56HZ-TL001 groups (3 mg / kg and 10 mg / kg), an Enfortumab-TLOO 1 group (10 mg / kg), and an Enfortumab-VC-MMAE group (10 mg / kg). All samples were administered via tail vein injection twice a week for a total of 6 doses.

[0319] Tumor diameters were measured by a vernier caliper twice a week after administration, and tumor volume was calculated according to the following formula: V = 0.5 a x b2, wherein a and b represent the long and short diameters of the tumor, respectively. Animal deaths were observed and recorded daily.

[0320] The following formula was used to calculate the tumor growth inhibition rate TGI (%), which was used to evaluate the tumor inhibition efficacy:

[0321] TGI (%) [1 ■ (Vr-end “ VT-start) / (V C-end ~ V C-start)] * 100%

[0322] wherein

[0323] V T-end: mean tumor volume of the treatment group at the end of the experiment;

[0324] V T-start: mean tumor volume of the treatment group at the start of the administration;

[0325] V c-ena: mean tumor volume of the negative control group at the end of the experiment;103NAI-5006826801V1

[0326] V c-start: mean tumor volume of the negative control group at the start of the administration.

[0327] The experimental results are shown in Table 4 and FIGs. 3 and 4, which demonstrate that 31HZ-TL001, 56HZ-TL001, Enfortumab-TLOOl, and Enfortumab-VC-MMAE had significant inhibitory effects on the tumor growth in the NCI-H322M xenograft tumor model in a dose-dependent manner. After 6 doses (Day 21), compared to the negative control group, the TGI values of 31HZ-TL001, 56HZ-TL001, Enfortumab-TLOOl, and Enfortumab-VC-MMAE at the 10 mg / kg dose level were 155.79%, 163.56%, 156.31% and 173.79%, respectively. There was no significant body weight loss in all treatment groups during the observation period, indicating the ADCs were well-tolerated by the animals.Table 4. NCI-H322M + NOD / SCID mouse CDX modelDay 21Tumor volume TumorGroup Dose3TGI(mm) regression P value(mg / kg) (%)Vehicle 10 347.06±25.84 / (0 / 0) / 31HZ-TL001 3 119.96±32.99 90.61 (1 / 0) < 0.00131HZ-TL001 10 42.92±3.00 155.79 (5 / 0) < 0.00156HZ-TL001 3 100.12±25.64 98.33 (4 / 0) < 0.00156HZ-TL001 10 35.53±2.12 163.56 (5 / 0) < 0.001Enfortumab-TLOOl 10 42.62±5.38 156.31 (5 / 0) < 0.001Enfortumab-VC-MMAE 10 25.47±4.70 173.79 (5 / 0) < 0.001Note: TGI: tumor growth inhibition rate; PR: partial regression of tumor; CR: complete regression of tumor. P value was a result compared with the vehicle group.

[0328] The above results indicated that, in view of the TGI values of 31HZ-TL001 and 56HZ- TL001 at the dose levels of 3 mg / kg and 10 mg / kg, both 31HZ-TL001 and 56HZ-TL001 could effectively inhibit tumor growth in a dose-dependent manner. At the dose level of 10 mg / kg, both 31HZ-TL001 and 56HZ-TL001 could partially regress tumors of all the 5 mice in the group, and the efficacy was basically equivalent to those of Enfortumab-TLOOl and Enfortumab-VC-MMAE. In addition, 31HZ-TL001 and 56HZ-TL001 were well-tolerated by tumor-bearing mice.104NAI-5006826801V1Example 5: In vivo pharmacodynamic of anti-human Nectin-4 ADC in PDX model

[0329] Tumor tissue was collected from HuPrime® bladder cancer xenograft model BL9200 (Crown Bioscience (Taicang) Co., Ltd., with high expression of Nectin-4, derived from a 63- year-old male patient) tumor-bearing mice, cut into pieces with a diameter of 3x3x3 mm, and inoculated subcutaneously at right anterior scapula of NOD / SCID mice (Jiangsu Jicui Yaokang Biotechnology Co., Ltd.). When the average tumor volume of tumor-bearing mice reached about 150-250 mm3, the mice were randomly divided into groups according to tumor size (7 mice per group) and the day of grouping was defined as Day 0. There were 4 groups: a human IgGl isotype control antibody group (IgGl 3 mg / kg, negative control); a human IgGl-TLOOl group (abbreviated as IgGl-TLOOl, 3 mg / kg); and a 1 mg / kg group and a 3 mg / kg group for 56HZ- TL001. All samples were administered via tail vein injection twice a week for a total of 6 doses. After administration, the tumor volume and body weight of the mice were observed and periodically measured according to the methods described in Example 4.

[0330] The experimental results are shown in Table 5 and FIGs. 5 and 6. After 6 doses (Day 20), compared to the 3 mg / kg human IgGl (hlgGl) negative control group, 56HZ-TL001 at 3 mg / kg showed an inhibitory TGI of 105.55% (P < 0.05) for tumor growth, and partial regression of tumors was observed in 4 mice of this group. 56HZ-TL001 at 3mg / kg showed significantly improved inhibitory effect on tumor growth than the isotype control ADC (hlgGl -TL001 at 3 mg / kg) (P < 0.001) at the same dose. One mouse died in the hlgGl group, and no death was observed in other groups with normal activities. During the administration period, the body weight of the mice in each group decreased slightly, among which the body weight of the IgGl group decreased the most due to the lack of effective treatment, indicating that the tumorbearing mice were well-tolerated to the ADCs.

[0331] The above results showed that 56HZ-TL001 showed significant inhibitory effect on the tumor growth of the BL9200 bladder cancer PDX xenograft model. In addition, the tumorbearing mice were well-tolerated to 56HZ-TL001.Table 5. HuPrime® bladder cancer + NOD / SCID mouse PDX modelDay 20Tumor volume Tumor P value P valueGroup Dose3TGI(mm) regression (relative (relative to105NAI-5006826801V1hlgGl-3 926.31±163.85 37.47 0 / 0 > 0.05 / TL00156HZ-3 101.71±16.78 105.55 4 / 0 < 0.001 < 0.001TL001Note: TGI: tumor growth inhibition rate; PR: partial regression of tumor; CR: complete regression of tumor; I', not applicable.Example 6: Phase 1 / 2 open-label study to evaluate the safety and efficacy of 56HZ-TL001 in participants with advanced solid tumors

[0332] This Example describes a cohort-expansion of a first-in-human, open-label, phase la study of a humanized anti-human NECTIN-4 antibody drug conjugate.Obiectives / Hypotheses and Endpoints

[0333] The study included male and female participants with advanced or metastatic solid tumors who were at least 18 years of age. Table 6 summarizes objectives and endpoints of the study.Table 6. Objectives and Endpoints106NAI-5006826801V1Overall Design

[0334] This was an open-label, multicenter, Phase 1 / 2 study of 56HZ-TL001 monotherapy in participants with histologically or cytologically confirmed diagnosis of select advanced solid tumors. The objectives of the study were to evaluate the safety, efficacy, and preliminary PK of 56HZ-TL001 monotherapy. A total of 180 participants in this study were allocated / randomized into the following cohorts:• Cohort A: Approximately 60 participants with 2L to 4L UC randomly assigned (1 : 1) to receive 56HZ-TL001 IV at 4 mg / kg q2w (Arm 1) or 5 mg / kg q2w (Arm 2). Participants were stratified by liver metastases (yes vs. no) and prior EV treatment (yes vs. no).• Cohort B: Approximately 30 participants with 2L to 4L HNSCC allocated to receive 56HZ-TL001 IV 5 mg / kg q2w (Arm 2)• Cohort C: Approximately 30 participants with 2L+ cervical cancer allocated to receive 56HZ-TL001 IV 5 mg / kg q2w (Arm 2)• Cohort D: Approximately 30 participants with 2L to 4L endometrial carcinoma allocated to receive 56HZ-TL001 IV 5 mg / kg q2w (Arm 2)• Cohort E: Approximately 30 participants with 2L or 3L TNBC allocated to receive 56HZ- TL001 IV 5 mg / kg q2w (Arm 2).

[0335] Participants received 56HZ-TL001 at the assigned dose until one of the criteria for discontinuation of study intervention was met (e.g., confirmed pregnancy, radiographic disease progression, treatment-related AEs or SAEs, treatment-unrelated medical conditions, etc.) or the 107NAI-5006826801V1participant withdrew from the study. In some instances, a cohort may have been terminated if the results of a nonbinding futility analysis did not cross the futility bar. There is no protocol- defined limit on the number of treatment cycles for 56HZ-TL001.

[0336] Participants who discontinued from treatment for reasons other than radiographic disease progression underwent posttreatment follow-up imaging to assess disease status until any of the conditions for discontinuation of imaging were met.

[0337] Preliminary efficacy was evaluated using OR assessed by the investigator as a secondary endpoint. Additional efficacy endpoints include DOR, PFS, and OS. After disease progression per RECIST 1.1 by investigator and / or initiation of a subsequent anticancer treatment, all participants were followed for vital status (e.g., by phone contact or clinic visit) until death, withdrawal of consent, loss to follow up, or until the study was concluded or terminated early, whichever occurred first.

[0338] AEs and SAEs were reported for participants and were graded for severity according to the guidelines outlined in the NCI CTCAE, Version 5.0.

[0339] Table 7 summarizes the overall design of the study. See also FIG. 7.Table 7. Summary of Study Design

[0340] A cohort may have been terminated if the results of a nonbinding futility analysis did not cross the futility bar.Inclusion and Exclusion Criteria

[0341] Inclusion Criteria: (1) Is an individual of any sex / gender, at least 18 years of age at the time of providing the informed consent or assent, as applicable; (2) has a confirmed advanced (unresectable and / or metastatic) solid tumor in one of the disease cohorts identified in Table 8108NAI-5006826801V1below; and (3) has received, or been intolerant to, all available treatments known to confer clinical benefit.Table 8. Disease Cohorts

[0342] Additional inclusion categories:• Measurable disease by RECIST 1.1 as assessed by the local site investigator / radiology. Lesions situated in a previously irradiated area are considered measurable if progression has been shown in such lesions.• Archival tumor tissue sample or newly obtained biopsy of a tumor lesion not previously irradiated has been provided.• Participants who have AEs due to previous anticancer therapies must have recovered to < Grade 1 or baseline. Participants with endocrine-related AEs who are adequately treated with hormone replacement or participants who have <Grade 2 neuropathy are eligible.• HIV-infected participants must have well controlled HIV on ART (antiretroviral therapy), defined as:- Participants on ART must have a CD4+ T-cell count >350 cells / mm3at the time of screening.- Participants on ART must have achieved and maintained virologic suppression defined as confirmed HIV RNA level below 50 or the LLOQ (below the limit of detection) using the locally available assay at the time of screening and for at least 12 weeks before screening.- It is advised that participants must not have had any AIDS-defining opportunistic infections within the past 12 months.- Participants on ART must have been on a stable regimen, without changes in drugs or dose modification, for at least 4 weeks before study entry (Day 1) and agree to continue ART throughout the study.- The combination ART regimen must not contain any antiretroviral medications that interact with CYP3A4 inhibitors / inducers / substrates109NAI-5006826801V1(https: / / www.fda.gov / drugs / drug-interactions-labeling / drug-development-and-drug- interactions-table-substrates-inhibitors-and-inducers).• Participants who are HBsAg positive are eligible if they have received HBV antiviral therapy for at least 4 weeks, and have undetectable HBV viral load prior to randomization. Note: Participants should remain on antiviral therapy throughout study intervention and follow local guidelines for HBV antiviral therapy post completion of study intervention. Hepatitis B screening tests are not required unless: known history of HBV infection; or as mandated by local health authority.• Participants with history of HCV infection are eligible if HCV viral load is undetectable at screening. Note: Participants must have completed curative antiviral therapy at least 4 weeks prior to randomization. Hepatitis C screening tests are not required unless: known history of HCV infection; or as mandated by local health authority.• An ECOG performance status of 0 to 1 assessed within 3 days before intervention all ocati on / randomizati on .• Adequate organ function as defined in the following table (Table 9). Specimens must be collected within 7 days before the start of study intervention.Table 9. Adequate Organ Function Laboratory Values110NAI-5006826801V1

[0343] Urothelial Carcinoma-Specific Inclusion Criteria (Cohort A)• Has pathologically or cytologically documented, locally advanced / unresectable or metastatic urothelial carcinoma of the bladder, renal pelvis, ureter, or urethra). Participants with histological variants are allowed if urothelial histology is predominant. Small cell / neuroendocrine tumors are not allowed, even if mixed histology.- Required treatments can be given in combination or sequentially.- Prior cisplatin-based therapy or PD-1 / PD-L1 therapy given for the treatment of MIBC is counted as 1 line of therapy.- To meet inclusion criteria, requirement of prior anti-PD-l / PD-Ll therapy use in theNMIBC (Non-Muscle-Invasive Bladder Cancer), MIBC (Muscle-Invasive Bladder Cancer), or metastatic setting will suffice.- Prior platinum-based therapy followed by PD-1 / PD-L1 maintenance therapy is counted as 1 line of therapy.- Prior FGFR-inhibitor treatment is allowed for eligible participants.- The same regimen administered twice in different disease settings will be counted as 1 line of prior therapy.- Has relapsed or progressed after treatment with at least 1 prior line of therapy(maximum of 4) that includes anti-PD-l / PD-Ll therapy. At least 1 line of therapy must also contain one of the following modalities: chemotherapy or EV (a Nectin-4- targeted drug conjugate).

[0344] HNSCC-Specific Inclusion Criteria (Cohort B)• Has histologically confirmed advanced or metastatic HNSCC of the oral cavity, oropharynx, hypopharynx, and / or larynx that is considered incurable by local therapies.• Note: Population should exclude nasopharynx, nasal cavity and paranasal sinuses, and unknown primary patients.• Disease progression after having received at least 1 but no more than 3 prior lines of systemic therapy in the unresectable recurrent or metastatic setting.• Must have had disease progression on anti-PD-l / Ll therapy (either as monotherapy or in combination with chemotherapy or other therapies.• Must have had disease progression on platinum-based therapy (either in the recurrent or metastatic setting or in the curative-intent locally advanced setting)• Has documentation of results from HPV status testing for oropharyngeal cancer. If HPV status has previously been tested using this procedure, no retesting is required. Note: Oral111NAI-5006826801V1cavity, hypopharynx, and larynx cancer are not required to undergo HPV testing as by convention these tumor locations are assumed to be HPV negative.

[0345] Cervical Cancer-Specific Inclusion Criteria (Cohort C)• Has histologically confirmed unresectable or metastatic squamous cell carcinoma, adenosquamous carcinoma, or adenocarcinoma of the cervix with disease progression after having received >1 prior line of therapy that includes systemic therapy in the advanced or metastatic setting. (Must be used platinum doublet chemotherapy with or without bevacizumab). Note: Participants may have also received prior chemoradiotherapy in the locally advanced cervical cancer setting; this is not considered a line of treatment. Note: Participants may receive prior anti-PD-l / PD-Ll treatment and / or tisotumab vedotin if those are SoC in the country.

[0346] Endometrial Carcinoma-specific Inclusion Criteria (Cohort D)• Has histologically confirmed diagnosis of endometrial carcinoma or carcinosarcoma.• Has radiographically evaluable disease (measurable or non-measurable) per RECIST 1.1• Has received prior systemic, platinum-based chemotherapy and anti-PD-l / anti-PD-Ll therapy, either separately or in combination, for EC or carcinosarcoma.• Has received up to 3 prior lines of therapy. Adjuvant ± neoadjuvant therapy is considered 1 line of therapy. The same regimen used in both the metastatic and adjuvant settings (e.g., platinum + taxanes) would be considered 2 lines of therapy.

[0347] TNBC-specific Inclusion Criteria (Cohort E)• Has TNBC as defined by the most recent ASCO / CAP guidelines.• Has received either 1 or 2 prior systemic treatments for metastatic breast cancer and have documented disease progression on or after their most recent therapy.• Has been previously treated with an anthracycline and / or taxane in the (neo)adjuvant or metastatic setting.

[0348] Exclusion Criteria• Medical Conditions- Has active inflammatory bowel disease requiring immunosuppressive medication or previous history of inflammatory bowel disease (e.g., Crohn’s disease, ulcerative colitis, or chronic diarrhea).- Has uncontrolled significant cardiovascular disease or cerebrovascular disease, including New York Heart Association Class III or IV congestive heart failure, unstable angina, uncontrolled symptomatic arrhythmia, prolongation of QTcF interval112NAI-5006826801V1to >480 ms, and / or other serious cardiovascular and cerebrovascular diseases within the 6 months preceding study intervention.- Has history of documented severe dry eye syndrome, severe Meibomian gland disease and / or blepharitis, or corneal disease that prevents / delays corneal healing.- Has pleural effusion, ascites, and / or pericardial effusion that are symptomatic or require repeated drainage, except for cases where small amounts of effusion detected by imaging.- HIV-infected participants with a history of Kaposi’s sarcoma and / or MulticentricCastleman’s Disease.• Prior / Concomitant Therapy- All cohorts except Cohort A: Received prior treatment with a Nectin-4-targeted ADC.For Cohort A, prior treatment with EV is allowed.- Received prior treatment with a topoisomerase I inhibitor, including ADCs.- Received strong CYP3 A4 inhibitors, inducers, or substrates within 2 weeks before the first dose of study intervention or within 5 half-lives of drug elimination, whichever is longer.- Note: A list of strong inhibitors or inducers of CYP3 A4 can be found at the following website: https: / / www.fda.gov / drugs / drug-interactions-labeling / drug-development- and-drug-interactions-table-substrates-inhibitors-and-inducers.- Received prior systemic anticancer therapy including investigational agents within4 weeks before the start of study intervention.- Received prior radiotherapy within 2 weeks of start of study intervention, or has radiation-related toxicities, requiring corticosteroids. Note: Two weeks or fewer of palliative radiotherapy for non-CNS disease is permitted. The last palliative radiotherapy treatment must have been performed at least 7 days before the first dose of study intervention.- Received a live or live-attenuated vaccine within 30 days before the first dose of study intervention. Administration of killed vaccines is allowed, as is COVID-19 vaccines.• Prior / Concurrent Clinical Study Experience- Has received an investigational agent or has used an investigational device within4 weeks prior to study intervention administration.• Diagnostic Assessments- Known additional malignancy that is progressing or has required active treatment within the past 2 years. Note: Participants with basal cell carcinoma of the skin,113NAI-5006826801V1squamous cell carcinoma of the skin, or carcinoma in situ, excluding carcinoma in situ of the bladder, that have undergone potentially curative therapy are not excluded. Participants with low-risk early-stage prostate cancer (Tl-T2a, Gleason score <6, and PSA <10 ng / mL) either treated with definitive intent or untreated in active surveillance with stable disease are not excluded.- Known active CNS metastases and / or carcinomatous meningitis. Participants with previously treated brain metastases may participate provided they are radiologically stable (i.e., without evidence of progression) for at least 4 weeks as confirmed by repeat imaging performed during the study screening, are clinically stable and have not required steroid treatment for at least 14 days before the first dose of study intervention.- Active infection requiring systemic therapy other than those expressly permitted.- History of (noninfectious) pneumonitis / interstitial lung disease that required steroids or has current pneumonitis / interstitial lung disease.- Concurrent active Hepatitis B (defined as HBsAg positive and / or detectable HBVDNA) and Hepatitis C virus (defined as anti-HCV Ab positive and detectable HCV RNA) infection. Note: Hepatitis B and C screening tests are not required unless: known history of HBV and HCV infection; or as mandated by local health authority.- History or current evidence of any condition, therapy, laboratory abnormality, or other circumstance that might confound the results of the study or interfere with the participant's ability to cooperate with the requirements of the study, such that it is not in the best interest of the participant to participate, in the opinion of the treating investigator.- A known severe hypersensitivity reaction to 56HZ-TL001 and / or any of its excipients.- Participants who have not adequately recovered from major surgery or have ongoing surgical complications.- Participants who are incapacitated are not eligible for this study.

[0349] Endometrial Carcinoma-Specific Exclusion CriteriaThere are no tumor-specific exclusion criteria for UC, HNSCC, cervical cancer, or TNBC. However, the following EC subjects meeting the following criteria are excluded from consideration.Has endometrial carcinoma or carcinosarcoma recurrence >12 months after completing platinum-based therapy administered in the curative-intent setting without additional platinum therapy given in recurrent setting. If immunotherapy -based treatment is114NAI-5006826801V1administered in recurrent setting, then platinum rechallenge is NOT required, regardless of the duration of platinum free interval. For Stage IVB disease, treatment that includes gynecological surgery followed by platinum-based regimen is NOT considered curativeintent by protocol and does not require platinum rechallenge in recurrent setting regardless of the duration of platinum free interval.• Has neuroendocrine tumors or endometrial sarcoma, including stromal sarcoma, leiomyosarcoma, adenosarcoma, or other types of pure sarcomas.Assessments

[0350] Tumor Imaging and Assessment of Disease

[0351] In addition to survival, efficacy was assessed based on evaluation of scan changes in tumor burden over time, until the participant was discontinued from the study or goes into survival follow-up. Tumor scans by CT were strongly preferred. For the abdomen and pelvis, contrast-enhanced MRI was optionally used when CT with iodinated contrast was contraindicated, or when mandated by local practice. The same scan technique was preferably used in a participant throughout the study to optimize the reproducibility of the assessment of existing and new tumor burden and improve the accuracy of the response assessment based on scans. Note: For the purposes of assessing tumor scans, the term “investigator” refers to the local investigator at the site and / or the radiological reviewer at the site or at an offsite facility.

[0352] If brain scans were performed, magnetic resonance imaging was preferred; however, CT imaging was acceptable, if MRI was medically contraindicated.

[0353] Bone scans were optionally performed to evaluate bone metastases. Any supplemental scans performed to support a positive or negative bone scan, such as plain X-rays acquired for correlation, were preferably submitted and / or retained.

[0354] Initial Tumor Scans

[0355] Initial tumor scans at screening were performed within 28 days prior to the date of randomization / allocation. Any scans obtained after Cycle 1 Day 1 cannot be included in the screening assessment. Sites must review screening scans to confirm the participant has measurable disease per RECIST 1.1.

[0356] If brain scans were required to document the stability of existing metastases, the brain scan were preferably acquired during screening.

[0357] Bone scans were required at screening for participants with a history of bone metastases and / or for those participants with indicative clinical signs / symptoms such as bone pain or elevated alkaline phosphatase levels. Bone scan refers to imaging methods used to assess bone metastasis.115NAI-5006826801V1

[0358] Tumor Scans During the Study

[0359] The first on-study scan was performed at 8 weeks (56 days ±7 days) from the date of allocation / randomization. Subsequent tumor scans were preferably performed every 8 weeks (±7 days) through Week 56, then every 12 weeks (±7 days) or more frequently if clinically indicated. After 24 months, participants who remain on treatment had scans performed every 6 months. Scan timing followed calendar days and was not adjusted for delays in cycle starts.

[0360] Scans were performed until disease progression is identified by the investigator or until any of these conditions were met: (1) the start of a new anticancer treatment; (2) pregnancy; (3) death; (4) withdrawal of consent; or (5) the end of the study. \

[0361] Objective response was confirmed by a repeat scan performed at least 4 weeks after the first indication of a response is observed. Participants then returned to the regular scan schedule, starting with the next scheduled time point. Participants who received additional scans for confirmation did not need to undergo the next scheduled scan if it was fewer than 4 weeks later; scans resumed at the subsequent scheduled time point.

[0362] On-study brain and bone scans were performed if clinically indicated or to confirm CR (if other lesions indicate CR and brain or bone lesions existed at baseline).

[0363] End-of-treatment and Follow-up Tumor Scans

[0364] If participants discontinued study intervention, tumor scans were performed at the time of discontinuation (±4-week window) unless previous scans were obtained within 4 weeks of discontinuation. If participants discontinued study intervention due to documented disease progression, this was the final required tumor scan. The scan type and anatomical coverage performed at end-of-treatment adhered to the requirements of the scheduled imaging visits. Additional imaging was acquired as clinically indicated or to confirm CR assessment.

[0365] If participants discontinued study intervention without documented disease progression, every effort was made to monitor disease status by acquiring tumor scans using the same schedule calculated from the date of randomization or allocation. Scans were continued until one of the following conditions were met: (1) disease progression as defined by RECIST 1.1; (2) the start of a new anticancer treatment; (3) pregnancy; (4) death; (5) withdrawal of consent; or (6) the end of the study.

[0366] RECIST 1.1 Assessment of Disease

[0367] RECIST 1.1 was used as the primary measure for assessment of tumor response, date of disease progression, and as a basis for all protocol guidelines related to disease status (e.g., discontinuation of study intervention). If disease progression was established by the investigator, the process continued as follows: (1) investigator judgment determined action; and116NAI-5006826801V1(2) if the participant was clinically stable and study intervention continued, communication with the Sponsor was required and a reconsent addendum was signed.

[0368] Performance Assessments

[0369] Eastern Cooperative Oncology Group Performance Status. The ECOG Performance Status is standardized criteria to measure how cancer impacts level of functioning (performance status) in terms of ability to care for oneself, daily activity, and physical ability (walking, working, etc.) with Grades 0 to 5. The investigator or qualified designee assessed ECOG status (see https: / / ecog-acrin.org / resources / ecog-performance-status) at screening, before the administration of each dose of study intervention, and during the follow-up period.

[0370] Ophthalmologic Examinations. The ophthalmologic examination for each eye included: best corrected distance acuity; slit lamp exam of the lids, cornea, conjunctiva, anterior chamber, iris and lens; fluorescein staining of the cornea; and tear break-up time.

[0371] Adverse Events, Serious Adverse Events, and Other Reportable Safety Events. The investigator and any designees were responsible for detecting, documenting, and reporting events that meet the definition of an adverse event (AE) or serious adverse event (SAE) as well as other reportable safety events. In addition, AEs, SAEs, and other reportable safety events were reported by the participant (or, when appropriate, by a caregiver, surrogate, or the participant’s legally authorized representative).

[0372] Pharmacokinetics

[0373] To further evaluate 56HZ-TL001 immunogenicity and exposure, and to evaluate exposure of the proposed dosing regimen, sample collections for analysis of PK and ADA occurred. Blood samples were obtained to measure PK of serum TAb and ADC. PK assessment included measurements of free payload, TAb, and ADC in serum or plasma. For free payload, plasma was collected. For ADC and TAb, serum was collected. Cmax and minimum concentration (Ctrough) at planned visits and times was summarized.117NAI-5006826801V1

Claims

WHAT IS CLAIMED IS:

1. A method for treating a cancer in a patient, the method comprising administering to the patient a therapeutically effective amount of an anti-human Nectin-4 antibody conjugate, wherein the antibody conjugate comprises an anti-human Nectin-4 antibody, or antigen-binding fragment thereof, and at least one therapeutic agent.

2. The method of claim 1, wherein the anti-human Nectin-4 antibody, or antigen-binding fragment thereof, comprises:(a) the following six complementarity determining regions (CDRs) for heavy and light chains defined according to the Chothia numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 21 or 18;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 22;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 23;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 24;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 25; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 26; or(b) the following six CDRs for heavy and light chains defined according to the Abm numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 27 or 36;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 28;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 23;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 24;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 25; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 26; or(c) the following six CDRs for heavy and light chains defined according to the Kabat numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 29 or 44;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 30;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 23;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 24;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 25; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 26; or118NAI-5006826801V1(d) the following six CDRs for heavy and light chains defined according to the IMGT numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 31 or 54;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 32;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 33;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 34;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 35; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 26.

3. The method of claim 1, wherein the anti-human Nectin-4 antibody, or antigen-binding fragment thereof, comprises:(a) the following six CDRs for heavy and light chains defined according to the Chothia numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 3;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 4;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 5;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 6;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 7; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 8; or(b) the following six CDRs for heavy and light chains defined according to the Abm numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 9;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 10;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 5;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 6;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 7; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 8; or(c) the following six CDRs for heavy and light chains defined according to the Kabat numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 11;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 12 or 63;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 5;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 6;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 7; and119NAI-5006826801V1CDR-L3 comprising the sequence as set forth in SEQ ID NO: 8; or(d) the following six CDRs for heavy and light chains defined according to the IMGT numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 13;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 14;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 15;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 16;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 17; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 8.

4. The method of claim 1, wherein the anti-human Nectin-4 antibody, or antigen-binding fragment thereof, comprises:(a) the following six CDRs for heavy and light chains defined according to the Chothia numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 39;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 40;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 41;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 42;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 43; andCDR-L3 comprising SEQ ID NO: 8 or a variant thereof; or(b) the following six CDRs for heavy and light chains defined according to the Abm numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 45;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 46;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 41;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 42;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 43; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 8; or(c) the following six CDRs for heavy and light chains defined according to the Kabat numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 47;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 48 or 64;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 41;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 42;120NAI-5006826801V1CDR-L2 comprising the sequence as set forth in SEQ ID NO: 43; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 8; or(d) the following six CDRs for heavy and light chains defined according to the IMGT numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 49;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 50;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 51;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 52;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 53; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 8.

5. The method of any one of claims 1 to 4, wherein the anti-human Nectin-4 antibody, or antigen-binding fragment thereof, comprises: a VH having the sequence as set forth in SEQ ID NO: 19 or SEQ ID NO: 57 and a VL having the sequence as set forth in SEQ ID NO: 20 or SEQ ID NO: 58; a VH having the sequence as set forth in SEQ ID NO: 1 or SEQ ID NO: 55 and a VL having the sequence as set forth in SEQ ID NO: 2 or SEQ ID NO: 56; or a VH having the sequence as set forth in SEQ ID NO: 37 or SEQ ID NO: 59 and a VL having the sequence as set forth in SEQ ID NO: 38 or SEQ ID NO: 60.

6. The method of any one of claims 1 to 5, wherein the anti-human Nectin-4 antibody, or antigen-binding fragment thereof, comprises:(1) a VH having the sequence as set forth in SEQ ID NO: 57 and a VL having the sequence as set forth in SEQ ID NO: 58;(2) a VH having the sequence as set forth in SEQ ID NO: 1 and a VL having the sequence as set forth in SEQ ID NO: 2;(3) a VH having the sequence as set forth in SEQ ID NO: 19 and a VL having the sequence as set forth in SEQ ID NO: 20;(4) a VH having the sequence as set forth in SEQ ID NO: 37 and a VL having the sequence as set forth in SEQ ID NO: 38;(5) a VH having the sequence as set forth in SEQ ID NO: 55 and a VL having the sequence as set forth in SEQ ID NO: 56; or(6) a VH having the sequence as set forth in SEQ ID NO: 59 and a VL having the sequence as set forth in SEQ ID NO: 60.121NAI-5006826801V17. The method of any one of claims 1 to 6, wherein the anti-human Nectin-4 antibody, or antigen-binding fragment thereof, further comprises: a human immunoglobulin heavy chain constant region or a variant thereof, and a light chain constant region or a variant thereof, wherein the variant has a substitution, deletion, or addition of one or more amino acids compared to the wild-type sequence from which it is derived, optionally wherein the variant has a substitution, deletion, or addition of up to 5 amino acids compared to the wild-type sequence from which it was derived.

8. The method of any one of claims 1 to 6, wherein the anti -human Nectin-4 antibody, or antigen-binding fragment thereof, further comprises a heavy chain constant region having the sequence as set forth in SEQ ID NO: 61 and a light chain constant region having the sequence as set forth in SEQ ID NO: 62.

9. The method of any one of claims 1 to 8, wherein the at least one therapeutic agent is a cytotoxic agent.

10. The method of claim 9, wherein the antibody conjugate has the structure shown in Formula (I),A-(L-D)y(i); wherein, A represents the antibody or antigen-binding fragment thereof according to any one of claims 1 to 8;L comprises an amino acid or a peptide consisting of 2-10 amino acids; or L is selected fromVai, Cit, Phe, Lys, D-Val, Leu, Gly, Ala, Asn,122NAI-5006826801V1of position 1 and position 2 represents the position where L and A are connected, and the other represents the position where L and D are connected;D is selected from the group consisting of cytotoxic agents, therapeutic antibodies, radioisotopes, oligonucleotides and analogs thereof, biologically active peptides, protein toxins and enzymes; preferably, D is a molecule with anti-tumor biological activity;123NAI-5006826801V1y is an integer of 1 to 10.

11. The method of claim 10, wherein the antibody conjugate is formed by connecting the (L-D) moiety of Formula (1) with a sulfhydryl group of A through a thioether bond,Formula (1), wherein the antibody conjugate has the structure of Formula (2):124NAI-5006826801V1Formula (2), wherein y is an integer from 1 to 10, and A represents the antibody or antigen-binding fragment thereof according to any one of claims 1 to 8.

12. The method of any one of claims 1 to 11, wherein the molar ratio (DAR value) of therapeutic agent to the antibody, or antigen-binding fragment thereof, in the composition is a decimal or integer between 1 and 10, optionally wherein the DAR is a decimal or integer between 1 and 8.

13. The method of any one of claims 1 to 12, wherein the age of the patient is greater than or equal to 18 years of age.

14. The method of any one of claims 1 to 13, wherein the cancer is selected from the group consisting of urothelial carcinoma (UC), head and neck squamous cell carcinoma (HNSCC), cervical cancer (CC), endometrial carcinoma (EC), and triple-negative breast cancer (TNBC).

15. The method of any one of claims 1 to 14, wherein the cancer is selected from the group consisting of cervical cancer (CC) and endometrial carcinoma (EC).

16. The method of claims 14 or 15, wherein the cancer is (i) locally recurrent unresectable and / or (ii) metastatic.

17. The method of any one of claims 14 to 16, wherein the patient was previously treated for the cancer and / or intolerant to, all available treatments known to confer clinical benefit.

18. The method of claim 17, wherein the patient was previously treated with a chemotherapy, an anti-PD-l / anti-PD-Ll therapy, tisotumab vedotin, and / or enfortumab vedotin.125NAI-5006826801V119. The method of any one of claims 1-18, wherein the antibody conjugate is administered to the patient via intravenous infusion.

20. The method of any one of claims 1-19, wherein the antibody conjugate is administered to the patient at a dose of about 4 mg / kg or about 5 mg / kg.

21. The method of any one of claims 1-20, wherein the antibody conjugate is administered to the patient every two weeks (Q2W).

22. A use of an anti-human Nectin-4 antibody conjugate in the manufacture of a medicament for treating a cancer in a patient, wherein the antibody conjugate comprises an anti-human Nectin-4 antibody, or antigen-binding fragment thereof, and at least one therapeutic.

23. The use of claim 22, wherein the anti -human Nectin-4 antibody, or antigen-binding fragment thereof, comprises:(a) the following six complementarity determining regions (CDRs) for heavy and light chains defined according to the Chothia numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 21 or 18;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 22;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 23;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 24;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 25; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 26; or(b) the following six CDRs for heavy and light chains defined according to the Abm numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 27 or 36;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 28;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 23;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 24;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 25; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 26; or(c) the following six CDRs for heavy and light chains defined according to the Kabat numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 29 or 44;126NAI-5006826801V1CDR-H2 comprising the sequence as set forth in SEQ ID NO: 30;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 23;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 24;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 25; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 26; or(d) the following six CDRs for heavy and light chains defined according to the IMGT numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 31 or 54;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 32;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 33;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 34;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 35; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 26.

24. The use of claim 22, wherein the anti-human Nectin-4 antibody, or antigen-binding fragment thereof, comprises:(a) the following six CDRs for heavy and light chains defined according to the Chothia numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 3;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 4;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 5;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 6;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 7; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 8; or(b) the following six CDRs for heavy and light chains defined according to the Abm numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 9;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 10;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 5;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 6;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 7; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 8; or(c) the following six CDRs for heavy and light chains defined according to the Kabat numbering system:127NAI-5006826801V1CDR-H1 comprising the sequence as set forth in SEQ ID NO: 11;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 12 or 63;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 5;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 6;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 7; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 8; or(d) the following six CDRs for heavy and light chains defined according to the IMGT numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 13;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 14;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 15;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 16;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 17; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 8.

25. The use of claim 22, wherein the anti -human Nectin-4 antibody, or antigen-binding fragment thereof, comprises:(a) the following six CDRs for heavy and light chains defined according to the Chothia numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 39;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 40;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 41;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 42;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 43; andCDR-L3 comprising SEQ ID NO: 8 or a variant thereof; or(b) the following six CDRs for heavy and light chains defined according to the Abm numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 45;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 46;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 41;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 42;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 43; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 8; or128NAI-5006826801V1(c) the following six CDRs for heavy and light chains defined according to the Kabat numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 47;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 48 or 64;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 41;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 42;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 43; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 8; or(d) the following six CDRs for heavy and light chains defined according to the IMGT numbering system:CDR-H1 comprising the sequence as set forth in SEQ ID NO: 49;CDR-H2 comprising the sequence as set forth in SEQ ID NO: 50;CDR-H3 comprising the sequence as set forth in SEQ ID NO: 51;CDR-L1 comprising the sequence as set forth in SEQ ID NO: 52;CDR-L2 comprising the sequence as set forth in SEQ ID NO: 53; andCDR-L3 comprising the sequence as set forth in SEQ ID NO: 8.

26. The use of any one of claims 22 to 25, wherein the anti-human Nectin-4 antibody, or antigen-binding fragment thereof, comprises: a VH having the sequence as set forth in SEQ ID NO: 19 or SEQ ID NO: 57 and a VL having the sequence as set forth in SEQ ID NO: 20 or SEQ ID NO: 58; a VH having the sequence as set forth in SEQ ID NO: 1 or SEQ ID NO: 55 and a VL having the sequence as set forth in SEQ ID NO: 2 or SEQ ID NO: 56; or a VH having the sequence as set forth in SEQ ID NO: 37 or SEQ ID NO: 59 and a VL having the sequence as set forth in SEQ ID NO: 38 or SEQ ID NO: 60.

27. The use of any one of claims 22 to 26, wherein the anti-human Nectin-4 antibody, or antigen-binding fragment thereof, comprises:(1) a VH having the sequence as set forth in SEQ ID NO: 57 and a VL having the sequence as set forth in SEQ ID NO: 58;(2) a VH having the sequence as set forth in SEQ ID NO: 1 and a VL having the sequence as set forth in SEQ ID NO: 2;(3) a VH having the sequence as set forth in SEQ ID NO: 19 and a VL having the sequence as set forth in SEQ ID NO: 20;129NAI-5006826801V1(4) a VH having the sequence as set forth in SEQ ID NO: 37 and a VL having the sequence as set forth in SEQ ID NO: 38;(5) a VH having the sequence as set forth in SEQ ID NO: 55 and a VL having the sequence as set forth in SEQ ID NO: 56; or(6) a VH having the sequence as set forth in SEQ ID NO: 59 and a VL having the sequence as set forth in SEQ ID NO: 60.

28. The use of any one of claims 22 to 27, wherein the anti-human Nectin-4 antibody, or antigen-binding fragment thereof, further comprises: a human immunoglobulin heavy chain constant region or a variant thereof, and a light chain constant region or a variant thereof, wherein the variant has a substitution, deletion, or addition of one or more amino acids compared to the wild-type sequence from which it is derived, optionally wherein the variant has a substitution, deletion, or addition of up to 5 amino acids compared to the wild-type sequence from which it was derived.

29. The use of any one of claims 22 to 27, wherein the anti-human Nectin-4 antibody, or antigen-binding fragment thereof, further comprises a heavy chain constant region having the sequence as set forth in SEQ ID NO: 61 and a light chain constant region having the sequence as set forth in SEQ ID NO: 62.

30. The use of any one of claims 22 to 29, wherein the at least one therapeutic agent is a cytotoxic agent.

31. The use of claim 30, wherein the antibody conjugate has the structure shown in Formula (I),A-(L-D)y(i); wherein, A represents the antibody or antigen-binding fragment thereof according to any one of claims 1 to 8;130NAI-5006826801V1L comprises an amino acid or a peptide consisting of 2-10 amino acids, or L is selected fromof position 1 and position 2 represents the position where L and A are connected, and the other represents the position where L and D are connected;D is selected from the group consisting of cytotoxic agents, therapeutic antibodies, radioisotopes, oligonucleotides and analogs thereof, biologically active peptides, protein toxins and enzymes; preferably, D is a molecule with anti-tumor biological activity;131NAI-5006826801V1y is an integer of 1 to 10.

32. The use of claim 31, wherein the antibody conjugate is formed by connecting the (L-D) moiety of Formula (1) with a sulfhydryl group of A through a thioether bond,Formula (1), wherein the antibody conjugate has the structure of Formula (2):132NAI-5006826801V1Formula (2), wherein y is an integer from 1 to 10, and A represents the antibody or antigen-binding fragment thereof according to any one of claims 1 to 8.

33. The use of any one of claims 22 to 32, wherein the molar ratio (DAR value) of therapeutic agent to the antibody, or antigen-binding fragment thereof, in the composition is a decimal or integer between 1 and 10, optionally wherein the DAR is a decimal or integer between 1 and 8.

34. The method of claim 12 or the use of claim 33, wherein the DAR value is 7.0, 7.03, 7.1, 7.12, 7.2, 7.3, 7.4, 7.5, 7.6, 7.7, 7.8, 7.9 or 8.0.

35. The method of claim 12 or the use of claim 33, wherein the DAR value is 7.71.

36. The use of any one of claims 22 to 35, wherein the age of the patient is greater than or equal to 18 years of age.

37. The use of any one of claims 22 to 36, wherein the cancer is selected from the group consisting of urothelial carcinoma (UC), head and neck squamous cell carcinoma (HNSCC), cervical cancer (CC), endometrial carcinoma (EC), and triple-negative breast cancer (TNBC).

38. The use of any one of claims 22 to 37, wherein the cancer is selected from the group consisting of cervical cancer (CC) and endometrial carcinoma (EC).

39. The use of claim 38, wherein the cancer is (i) locally recurrent unresectable and / or (ii) metastatic.133NAI-5006826801V140. The use of any one of claims 37 to 39, wherein the patient was previously treated for the cancer.

41. The use of claim 40, wherein the patient was previously treated with a chemotherapy, an anti-PD-l / anti-PD-Ll therapy, tisotumab vedotin, and / or enfortumab vedotin.

42. The use of any one of claims 22-41, wherein the antibody conjugate is administered to the patient via intravenous infusion.

43. The use of any one of claims 22-42, wherein the antibody conjugate is administered to the patient at a dose of about 4 mg / kg or about 5 mg / kg.

44. The use of any one of claims 22-43, wherein the antibody conjugate is administered to the patient every two weeks (Q2W).

45. The method of any one of claims 1 to 21, 34 or 35, or the use of any one of claims 22 to 44, wherein the antibody conjugate or medicament is administered at least twice.

46. A kit comprising the antibody conjugate for use of any one of claims 22 to 35, and instructions to administer the antibody conjugate to a patient having a cancer.

47. The kit of claim 46, wherein the age of the patient is greater than or equal to 18 years of age.

48. The kit of claims 46 or 47, wherein the cancer is selected from the group consisting of urothelial carcinoma (UC), head and neck squamous cell carcinoma (HNSCC), cervical cancer (CC), endometrial carcinoma (EC), and triple-negative breast cancer (TNBC).

49. The kit of any one of claims 46 to 48, wherein the cancer is selected from the group consisting of cervical cancer (CC) and endometrial carcinoma (EC).

50. The kit of claims 48 or 49, wherein the cancer is (i) locally recurrent unresectable and / or (ii) metastatic.134NAI-5006826801V151. The method of any one of claims 1 to 21, 34 ,35, or 45, the use of any one of claims 22 to 45, or the kit of any one of claims 46 to 50, wherein the antibody conjugate or medicament is used in combination with one or more therapeutic agents selected from the group consisting of antineoplastic agents, chemotherapeutic agents, radioisotopes, immune checkpoint inhibitors.

52. The method of any one of claims 1 to 21, 34 ,35, or 45 the use of any one of claims 22 to 45, or the kit of any one of claims 46 to 50, wherein the antibody conjugate or medicament is used in combination with one or more therapeutic regimens selected from the group consisting of surgery, radiotherapy, chemotherapy, targeted therapy, immunotherapy, hormone therapy, angiogenesis inhibition, and palliative care.

53. The antibody conjugate of any one of claims 22 to 35, for use in method of any one of claims 1-21, 34 ,35, or 45, the use of any one of claims 22 to 45, or the kit of any one of claims 46 to 50.

54. The method of any one of claims 1-21, 34 ,35, or 45, the use of any one of claims 22 to 45, the kit of any one of claims 46 to 50, or the antibody conjugate of claim 51 wherein the antihuman Nectin-4 antibody, or antigen-binding fragment thereof, is the anti-human Nectin-4 antibody.135NAI-5006826801V1

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