Antibody drug conjugate as well as preparation method and application thereof

Abstract

The invention relates to an antibody drug conjugate as well as a preparation method and application thereof, in particular to an antibody drug conjugate for treating PTK7 positive cancer, which comprises a PTK7 antibody and a drug-connector molecule coupled to the antibody. In some embodiments, the antibody is humanized, has excellent binding activity to PTK7 positive cells, and can efficiently deliver a drug to the PTK7 positive cells. In some embodiments, the drug comprises a DNA topoisomerase inhibitor. The antibody drug conjugate provided by the invention has a better drug-antibody coupling ratio, and has a very good targeted killing effect on lung cancer, breast cancer, epidermal cancer, ovarian cancer, esophageal cancer and the like. Therefore, the invention further provides a preparation method of the antibody drug conjugate and application of the antibody drug conjugate in treatment of PTK7 positive cancers.

Description

[0001] Related applications

[0002] This application claims priority to Chinese patent application No. 202311284321.4 filed on September 28, 2023 and Chinese patent application No. 202211263034.0 filed on October 14, 2022, the entire disclosures of which are incorporated herein by reference.

[0003] Sequence Listing

[0004] This application contains a computer-readable sequence listing, which is filed with this application in XML format, the entire contents of which are incorporated herein by reference. The sequence listing is an XML file titled "14463-051-228_SEQ_LISTING.xml", created on September 14, 2023, and is 63,238 bytes in size. Technical Field

[0005] The present application relates to the field of targeted therapy, and specifically to an antibody-drug conjugate for treating PTK7-positive cancers. Specifically, the antibody-drug conjugate comprises a PTK7 antibody and a drug-linker molecule coupled to the antibody. In some embodiments, the antibody is humanized and has excellent binding activity to PTK7-positive cells, enabling efficient drug delivery to PTK7-positive cells. In some embodiments, the drug comprises a DNA topoisomerase inhibitor. The antibody-drug conjugate of the present application has an excellent drug-antibody coupling ratio and exhibits excellent targeted killing effects against lung cancer, breast cancer, epidermal cancer, ovarian cancer, and esophageal cancer. Therefore, the present application further provides a method for preparing the antibody-drug conjugate and its use in treating PTK7-positive cancers. Background Art

[0006] Cancer is one of the leading causes of death today. It is a disease caused by the malignant transformation of healthy cells, triggered by genetic alterations such as chromosomal translocations and mutations in tumor suppressor genes and growth factor receptors, leading to malignant cell proliferation. Defective apoptosis, or programmed cell death, further promotes the malignant transformation of cells that leads to cancer. According to the latest assessment by the International Agency for Research on Cancer (IARC), an agency of the World Health Organization (WHO), 19.29 million new cases of cancer were diagnosed worldwide in 2020, including 10.06 million in men and 9.23 million in women. In 2020, 9.96 million deaths from cancer were reported worldwide, including 5.53 million in men and 4.43 million in women. In this century, cancer is expected to surpass cardiovascular disease as the leading cause of premature death in most countries.

[0007] PTK7 (protein tyrosine kinase 7) belongs to the receptor tyrosine kinase family and lacks kinase activity due to mutations in its kinase domain. The PTK7 protein consists of seven extracellular immunoglobulin domains, a transmembrane region, and an intracellular tyrosine kinase domain. Its ligand is unknown. Its extracellular domain can be cleaved by ADAM and MT1-MMP proteases, producing soluble fragments. The concentration of free PTK7 in healthy human serum is approximately 12.4 ± 3.3 ng / ml; in cancer patients, the concentration is higher, approximately 24.6 ± 3.8 ng / ml.

[0008] PTK7 is expressed in a variety of solid tumors: reportedly overexpressed in 47.4% of NSCLC, 45.1% of ovarian cancer, 28.6% of TNBC, and 60% of esophageal cancer. Studies have shown that PTK7 expression promotes tumor cell growth, and knocking out the PTK7 gene reduces tumor burden in mice. High PTK7 expression is positively correlated with disease stage and lymph node metastasis, and negatively correlated with survival. Currently, research is underway with therapies targeting PTK7, including antibody-drug conjugates and CAR-T cells. Preclinical studies have demonstrated high efficacy against PTK7-overexpressing tumor cell lines. Strong tumor suppressor activity has been demonstrated in human cell-derived xenograft (CDX) and patient-derived tumor xenograft (PDX) models. Cofetuzumab Pelidotin, an ADC targeting PTK7 developed by Pfizer, has demonstrated favorable safety and preliminary efficacy in Phase I clinical trials.

[0009] ADC drugs consist of antibodies, bioactive molecules, and linkers. The bioactive molecules are covalently coupled to the antibodies via the linkers. Antibodies (such as monoclonal antibodies) can specifically recognize targets on the surface of tumor cells, guiding the ADC to the cancer cell surface and allowing the ADC to enter the cancer cell through endocytosis. The bioactive molecules are then released inside the cancer cell, killing the cancer cells while minimizing damage to normal tissue cells. Developing antibodies with improved targeting, hydrophilicity, and endocytosis activity for coupling to cytotoxic small molecules to create ADC drugs with a wider therapeutic window will provide patients with better treatment options. Summary of the Invention

[0010] The present application relates to an antibody-drug conjugate for treating PTK7-positive cancers, and exemplarily discloses an antibody-drug conjugate with the general formula Ab-[MLED] using humanized antibodies 101A6HZ, 101A6HZm and 64A10HZ as targeting moieties. xAn antibody-drug conjugate with the structure shown. Results showed that the conjugate had a relatively good drug-antibody coupling ratio and excellent binding activity against PTK7-positive cells, with a strong targeted killing effect against PTK7-positive cancers, such as lung cancer, breast cancer, epidermal cancer, ovarian cancer, and esophageal cancer. Therefore, the present application provides an antibody-drug conjugate for treating cancers with high PTK7 expression, a pharmaceutical composition containing the antibody-drug conjugate, and their use in treating cancers with high PTK7 expression.

[0011] Antibody Drug Conjugates

[0012] In one aspect, the present application provides an antibody drug conjugate having the formula Ab-[MLED] x The structure shown, wherein:

[0013] Ab is an antibody or antigen-binding fragment thereof that specifically binds to human tyrosine kinase 7 (PTK7);

[0014] M is a linker site connected to the antibody or antigen-binding fragment thereof;

[0015] L is a connector connecting M and E;

[0016] E is a structural fragment connecting L and D;

[0017] D is the cytotoxic drug fragment;

[0018] x is selected from 1 to 10.

[0019] In some embodiments, the antibody or antigen-binding fragment thereof comprises:

[0020] (1) The following heavy chain variable region (VH) and / or light chain variable region (VL), wherein the CDRs are defined according to the Chothia numbering system:

[0021] (1a) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 11 or a variant thereof, CDR-H2 of SEQ ID NO: 12 or a variant thereof, and CDR-H3 of SEQ ID NO: 13 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 14 or a variant thereof, CDR-L2 of SEQ ID NO: 15 or a variant thereof, and CDR-L3 of SEQ ID NO: 16 or a variant thereof; or

[0022] (1b) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 27 or a variant thereof, CDR-H2 of SEQ ID NO: 28 or a variant thereof, and CDR-H3 of SEQ ID NO: 29 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 30 or a variant thereof, CDR-L2 of SEQ ID NO: 31 or a variant thereof, and CDR-L3 of SEQ ID NO: 32 or a variant thereof;

[0023] wherein the variant described in any one of (1a) and (1b) has at least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with the sequence from which it is derived, or the variant has one or more amino acid substitutions, deletions or additions (e.g., substitutions, deletions or additions of 1, 2 or 3 amino acids) compared to the sequence from which it is derived; preferably, the substitutions are conservative substitutions;

[0024] or,

[0025] (2) the following heavy chain variable region (VH) and / or light chain variable region (VL), wherein the CDRs are defined according to the Kabat numbering system:

[0026] (2a) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 17 or a variant thereof, CDR-H2 of SEQ ID NO: 18 or 19 or a variant thereof, and CDR-H3 of SEQ ID NO: 13 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 14 or a variant thereof, CDR-L2 of SEQ ID NO: 15 or a variant thereof, and CDR-L3 of SEQ ID NO: 16 or a variant thereof; or

[0027] (2b) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 33 or a variant thereof, CDR-H2 of SEQ ID NO: 34 or 35 or a variant thereof, and CDR-H3 of SEQ ID NO: 29 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 30 or a variant thereof, CDR-L2 of SEQ ID NO: 31 or a variant thereof, and CDR-L3 of SEQ ID NO: 32 or a variant thereof;

[0028] wherein the variant described in any one of (2a) and (2b) has at least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with the sequence from which it is derived, or the variant has one or more amino acid substitutions, deletions or additions (e.g., substitutions, deletions or additions of 1, 2 or 3 amino acids) compared to the sequence from which it is derived; preferably, the substitutions are conservative substitutions;

[0029] or,

[0030] (3) The following heavy chain variable region (VH) and / or light chain variable region (VL), wherein the CDRs are defined according to the IMGT numbering system:

[0031] (3a) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 20 or a variant thereof, CDR-H2 of SEQ ID NO: 21 or a variant thereof, and CDR-H3 of SEQ ID NO: 22 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 23 or a variant thereof, CDR-L2 of SEQ ID NO: 24 or a variant thereof, and CDR-L3 of SEQ ID NO: 16 or a variant thereof; or

[0032] (3b) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 36 or a variant thereof, CDR-H2 of SEQ ID NO: 37 or a variant thereof, and CDR-H3 of SEQ ID NO: 38 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 39 or a variant thereof, CDR-L2 of SEQ ID NO: 40 or a variant thereof, and CDR-L3 of SEQ ID NO: 32 or a variant thereof;

[0033] wherein the variant described in any one of (3a) and (3b) has at least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with the sequence from which it is derived, or the variant has one or more amino acid substitutions, deletions or additions (e.g., substitutions, deletions or additions of 1, 2 or 3 amino acids) compared to the sequence from which it is derived; preferably, the substitutions are conservative substitutions;

[0034] or,

[0035] (4) The following heavy chain variable region (VH) and / or light chain variable region (VL), wherein the CDRs are defined according to the AbM numbering system:

[0036] (4a) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 25 or a variant thereof, CDR-H2 of SEQ ID NO: 26 or a variant thereof, and CDR-H3 of SEQ ID NO: 13 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 14 or a variant thereof, CDR-L2 of SEQ ID NO: 15 or a variant thereof, and CDR-L3 of SEQ ID NO: 16 or a variant thereof; or

[0037] (4b) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 41 or a variant thereof, CDR-H2 of SEQ ID NO: 42 or a variant thereof, and CDR-H3 of SEQ ID NO: 29 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 30 or a variant thereof, CDR-L2 of SEQ ID NO: 31 or a variant thereof, and CDR-L3 of SEQ ID NO: 32 or a variant thereof;

[0038] Wherein, the variant described in any one of (4a) and (4b) has at least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity compared with the sequence from which it is derived, or the variant has one or more amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) compared with the sequence from which it is derived; preferably, the substitutions are conservative substitutions.

[0039] In some embodiments, the antibody or antigen-binding fragment thereof comprises:

[0040] (a) VH or a variant thereof shown in SEQ ID NO: 1, and / or VL or a variant thereof shown in SEQ ID NO: 2;

[0041] (b) VH or a variant thereof shown in SEQ ID NO: 3, and / or VL or a variant thereof shown in SEQ ID NO: 4; or

[0042] (c) VH or a variant thereof shown in SEQ ID NO: 5, and / or VL or a variant thereof shown in SEQ ID NO: 6;

[0043] (d) VH or a variant thereof shown in SEQ ID NO: 7, and / or VL or a variant thereof shown in SEQ ID NO: 8;

[0044] (e) VH or a variant thereof shown in SEQ ID NO: 9, and / or VL or a variant thereof shown in SEQ ID NO: 10;

[0045] wherein the variant has at least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity compared to the sequence from which it is derived, or the variant has one or more amino acid substitutions, deletions or additions (e.g., 1, 2, 3, 4 or 5 amino acid substitutions, deletions or additions) compared to the sequence from which it is derived; preferably, the substitutions are conservative substitutions.

[0046] In some embodiments, the antibody or antigen-binding fragment thereof further comprises:

[0047] (a) a heavy chain constant region (CH) of a human immunoglobulin, or a variant thereof, which has one or more amino acid substitutions, deletions or additions (e.g., up to 20, up to 15, up to 10, or up to 5 amino acid substitutions, deletions or additions; e.g., 1, 2, 3, 4 or 5 amino acid substitutions, deletions or additions) compared to the wild-type sequence from which it is derived; and

[0048] (b) a light chain constant region (CL) of a human immunoglobulin, or a variant thereof, which has one or more amino acid substitutions, deletions or additions (e.g., up to 20, up to 15, up to 10, or up to 5 amino acid substitutions, deletions or additions; for example, 1, 2, 3, 4 or 5 amino acid substitutions, deletions or additions) compared to the wild-type sequence from which it is derived.

[0049] In some embodiments, the heavy chain constant region is an IgG heavy chain constant region, such as an IgG1, IgG2, IgG3 or IgG4 heavy chain constant region, such as a human IgG1 heavy chain constant region or a human IgG4 heavy chain constant region. In some embodiments, the antibody or antigen-binding fragment thereof comprises a heavy chain constant region (CH) as set forth in SEQ ID NO: 43 or a variant thereof, wherein the variant has up to 20 conservative substitutions of amino acids compared to SEQ ID NO: 43 (e.g., up to 15, up to 10, or up to 5 conservative substitutions of amino acids; e.g., 1, 2, 3, 4 or 5 conservative substitutions of amino acids).

[0050] In some embodiments, the light chain constant region is a kappa light chain constant region. In some embodiments, the antibody or antigen-binding fragment thereof comprises a light chain constant region (CL) as set forth in SEQ ID NO: 44, or a variant thereof, wherein the variant has up to 20 conservative amino acid substitutions (e.g., up to 15, up to 10, or up to 5 conservative amino acid substitutions; e.g., 1, 2, 3, 4, or 5 conservative amino acid substitutions) compared to SEQ ID NO: 44.

[0051] In some embodiments, the antibody or antigen-binding fragment thereof comprises a heavy chain constant region (CH) as shown in SEQ ID NO: 43 or 45 and a light chain constant region (CL) as shown in SEQ ID NO: 44.

[0052] In some embodiments, the antibody or antigen-binding fragment thereof comprises:

[0053] (1) a heavy chain comprising the VH sequence of SEQ ID NO: 1 and the heavy chain constant region (CH) of SEQ ID NO: 43, and a light chain comprising the VL sequence of SEQ ID NO: 2 and the light chain constant region (CL) of SEQ ID NO: 44;

[0054] (2) a heavy chain comprising a VH sequence of SEQ ID NO: 3 and a heavy chain constant region (CH) of SEQ ID NO: 43 or 45, and a light chain comprising a VL sequence of SEQ ID NO: 4 and a light chain constant region (CL) of SEQ ID NO: 44;

[0055] (3) a heavy chain comprising the VH sequence of SEQ ID NO: 5 and the heavy chain constant region (CH) of SEQ ID NO: 43, and a light chain comprising the VL sequence of SEQ ID NO: 6 and the light chain constant region (CL) of SEQ ID NO: 44;

[0056] (4) a heavy chain comprising the VH sequence of SEQ ID NO: 7 and the heavy chain constant region (CH) of SEQ ID NO: 43, and a light chain comprising the VL sequence of SEQ ID NO: 8 and the light chain constant region (CL) of SEQ ID NO: 44; or

[0057] (5) A heavy chain comprising the VH sequence shown in SEQ ID NO: 9 and the heavy chain constant region (CH) shown in SEQ ID NO: 43, and a light chain comprising the VL sequence shown in SEQ ID NO: 10 and the light chain constant region (CL) shown in SEQ ID NO: 44.

[0058] In the antibody-drug conjugate, the cytotoxic drug can be linked to the antibody or antigen-binding fragment thereof via a linker (such as the "MLE" fragment shown in this application).

[0059] In some embodiments, M is Wherein ring A is a 5-6 membered alicyclic heterocyclic ring, or a 5-20 membered aromatic ring system, wherein the alicyclic heterocyclic ring and the aromatic ring system are optionally substituted by one or more selected from oxy (=O), halogen, cyano, amino, carboxyl, thiol and C 1-6 Alkyl group substituted; M1 is selected from single bond and C 1-20 Alkylene, C 2-20 Alkenylene or C 2-20 Alkynylidene.

[0060] In some embodiments, M is wherein Ring A is a 5-membered alicyclic heterocycle, a 6-membered heteroaromatic ring, or a polycyclic ring formed by connecting one or more 6-membered heteroaromatic rings to a benzene ring via a single bond, wherein the alicyclic heterocycle is optionally substituted with one or more selected from oxy (=O), halogen and C 1-4 Alkyl group substituted; M1 is selected from single bond and C 3-10 Alkylene, C 3-10 Alkenylene or C 3-10 Alkynylidene.

[0061] In some embodiments, M is wherein ring A is selected from M1 is selected from a single bond and C 5-8 Alkylene, C 5-8 Alkenylene or C 5-8 Alkynylidene.

[0062] In some embodiments, M is selected from the following structures:

[0063]

[0064] In some embodiments, M is selected from the following structures:

[0065]

[0066] In some embodiments, L is selected from the group consisting of one or more of the following: C 1-6 Alkylene, -N(R')-, Carbonyl, -O-, Val, Cit, Phe, Lys, Lys(COCH2CH2(OCH2CH2) sOCH3), D-Val, Leu, Gly, Ala, Asn, Val-Cit, Val-Ala, Val-Lys, Val-Lys(Ac), Phe-Lys, Phe-Lys(Ac), D-Val-Leu- Lys, Gly-Gly-Arg, Ala-Ala-Asn, Ala-Ala-Ala, Val-Lys-Ala, Val-Lys-Gly, Gly-Gly-Gly, Gly-Gly-Phe-Gly (SEQ ID NO:48), Gly-Phe-Leu-Gly (SEQ ID NO:49), Gly-Gly-Val-Ala (SEQ ID NO:50), Gly-Gly-Gly-Gly-Gly (SEQ ID NO:51), Where R' represents hydrogen, C 1-6 Alkyl or alkyl containing -(CH2CH2O)r-; r is selected from an integer of 1-10; s is selected from an integer of 1-20.

[0067] In some embodiments, L is selected from the group consisting of one or more of the following: C 1-6 Alkylene, -N(R')-, Carbonyl, -O-, Val, Cit, Phe, Lys, Lys(COCH2CH2(OCH2CH2) s OCH3), D-Val, Leu, Gly, Ala, Asn, Val-Cit, Val-Ala, Val-Lys, Val-Lys(Ac), Phe-Lys, Phe-Lys(Ac), D-Val-Leu- Lys, Gly-Gly-Arg, Ala-Ala-Asn, Ala-Ala-Ala, Val-Lys-Ala, Val-Lys-Gly, Gly-Gly-Gly, Gly-Gly-Phe-Gly (SEQ ID NO:48), Gly-Phe-Leu-Gly (SEQ ID NO:49), Gly-Gly-Val-Ala (SEQ ID NO:50), Gly-Gly-Gly-Gly-Gly (SEQ ID NO:51), Where R' represents hydrogen, C 1-6 Alkyl or alkyl containing -(CH2CH2O)r-; r is selected from an integer of 1-10; s is selected from an integer of 1-20.

[0068] In some embodiments, L is selected from the group consisting of one or more of the following: C 1-6Alkylene, -NH-, Val, Cit, Phe, Lys, Lys(COCH2CH2(OCH2CH2) s OCH3), Gly, Val-Cit, Gly-Gly-Phe-Gly (SEQ ID NO: 48), wherein s is selected from an integer of 1-20.

[0069] In some embodiments, L is selected from the group consisting of one or more of the following: C 1-6 Alkylene, -NH-, Val, Cit, Phe, Lys, Lys(COCH2CH2(OCH2CH2) s OCH3), Gly, Val-Cit, Gly-Gly-Phe-Gly (SEQ ID NO: 48), wherein s is selected from an integer of 1-20.

[0070] In some embodiments, L is selected from the following structures:

[0071]

[0072] wherein s is selected from an integer of 1-20.

[0073] In some embodiments, L is selected from the following structures:

[0074]

[0075] wherein s is selected from an integer of 1-20.

[0076] In some embodiments, L is selected from the following structures:

[0077]

[0078] In some embodiments, L is selected from the following structures:

[0079]

[0080]

[0081] In some embodiments, L is selected from the following structures:

[0082]

[0083] In some embodiments, L is selected from the following structures:

[0084]

[0085] In some embodiments, E is a single bond, -NH-CH2-, -NH-CH2-O-CH2-CO-,

[0086] In some embodiments, E is a single bond, -NH-CH2-, -NH-CH2-O-CH2-CO-,

[0087] In some embodiments, E is -NH-CH2-,

[0088] In some embodiments, E is -NH-CH2- or

[0089] In some embodiments, M is selected from the following structures:

[0090]

[0091] L is selected from the following structures:

[0092]

[0093] E is -NH-CH2,

[0094] In some embodiments, M is selected from the following structures:

[0095]

[0096] L is selected from the following structures:

[0097]

[0098] E is -NH-CH2,

[0099] In some embodiments, Selected from the following structures:

[0100]

[0101]

[0102]

[0103]

[0104] In some embodiments, Selected from the following structures:

[0105]

[0106]

[0107]

[0108]

[0109] In some embodiments, Selected from the following structures:

[0110]

[0111]

[0112] In some embodiments, Selected from the following structures:

[0113]

[0114]

[0115] In some embodiments, the cytotoxic drug is selected from microtubule inhibitors, DNA intercalators, DNA topoisomerase inhibitors and RNA polymerase inhibitors. In some embodiments, the microtubule inhibitor is an auristatin compound or a maytansine compound. In some embodiments, the DNA intercalator is a pyrrolobenzodiazepine (PBD). In some embodiments, the DNA topoisomerase inhibitor is a topoisomerase I inhibitor (e.g., camptothecin, hydroxycamptothecin, 9-aminocamptothecin, SN-38, irinotecan, topotecan, belotecan, or rubitecan) or a topoisomerase II inhibitor (e.g., doxorubicin, PNU-159682, multicarmycin, daunorubicin, mitoxantrone, podophyllotoxin, or etoposide). In some embodiments, the RNA polymerase inhibitor is α-amanitin or a pharmaceutically acceptable salt, ester, or analog thereof.

[0116] The cytotoxic drugs disclosed in this application generally contain a variety of functional groups, such as hydroxyl (-OH), carboxyl (-COOH), sulfhydryl (-SH), primary amine (-NH2), secondary amine (-NR A H) or tertiary amine (-NR B R C ), where R A 、R B 、R C These represent only non-hydrogen substituents on N, through which the cytotoxic drug can be attached to the linker in the conjugate.

[0117] In some embodiments, the cytotoxic drug is linked to E in the antibody drug conjugate through a -OH, -SH, primary amine group, secondary amine group, or tertiary amine group on the cytotoxic drug.

[0118] In some embodiments, the cytotoxic drug is

[0119] In some embodiments, the cytotoxic drug is selected from the following Formula I and Formula II:

[0120]

[0121] Wherein, R1, R2 are each independently selected from C 1-6 Alkyl and halogen;

[0122] R3 is selected from H and -CO-CH2OH;

[0123] R4 and R5 are each independently selected from H, halogen and hydroxyl; or R4 and R5 are connected to the connected carbon atom to form a 5-6 membered oxygen-containing heterocyclic ring;

[0124] R6 is selected from hydrogen or -C 1-4 Alkylene-NR a R b ;

[0125] R7 is selected from hydrogen, C 1-6 Alkyl and -C 1-4 Alkylene-NR a R b ;

[0126] where R a 、R b Each occurrence is independently selected from H, C 1-6 Alkyl, -SO2-C 1-6 Alkyl and -CO-C 1-6 alkyl.

[0127] In some embodiments, the cytotoxic drug is selected from the following Formula I and Formula II:

[0128]

[0129] Wherein, R1, R2 are each independently selected from C 1-6 Alkyl and halogen;

[0130] R3 is selected from H and -CO-CH2OH;

[0131] R4 and R5 are each independently selected from H, halogen and hydroxyl; or R4 and R5 are connected to the connected carbon atom to form a 5-6 membered oxygen-containing heterocyclic ring;

[0132] R6 is selected from hydrogen or -C1-4 Alkylene-NR a R b ;

[0133] R7 is selected from C 1-6 Alkyl and -C 1-4 Alkylene-NR a R b ;

[0134] where R a 、R b Each occurrence is independently selected from H, C 1-6 Alkyl, -SO2-C 1-6 Alkyl and -CO-C 1-6 alkyl.

[0135] In some embodiments, the cytotoxic drug is selected from the following compounds:

[0136]

[0137] In some embodiments, the cytotoxic drug is selected from the following compounds:

[0138]

[0139]

[0140] The fragment of the cytotoxic drug obtained by connecting the cytotoxic drug to the linker is the formula Ab-[MLED] x In some embodiments, D is a monovalent structure obtained by losing one H from -OH, -NH2 or a secondary amine group on the cytotoxic drug.

[0141] In some embodiments, D is selected from the following structures:

[0142]

[0143]

[0144] In some embodiments, the antibody drug conjugate is selected from ADC A-01 to ADC A-26, ADCB-01 to ADC B-06, and ADC C-01 shown below:

[0145] ADC A-01

[0146]

[0147] ADC A-02

[0148]

[0149] ADC A-03

[0150]

[0151] ADC A-04

[0152]

[0153] ADC A-05

[0154]

[0155] ADC A-06

[0156]

[0157] ADC A-07

[0158]

[0159] ADC A-08

[0160]

[0161] ADC A-09

[0162]

[0163] ADC A-10

[0164]

[0165] ADC A-11

[0166]

[0167] ADC A-12

[0168]

[0169] ADC A-13

[0170]

[0171] ADC A-14

[0172]

[0173] ADC A-15

[0174]

[0175] ADC A-16

[0176]

[0177] ADC A-17

[0178]

[0179] ADC A-18

[0180]

[0181] ADC A-19

[0182]

[0183] ADC A-20

[0184]

[0185] ADC A-21

[0186]

[0187] ADC A-22

[0188]

[0189] ADC A-23

[0190]

[0191] ADC A-24

[0192]

[0193] ADC A-25

[0194]

[0195] ADC A-26

[0196]

[0197] ADC B-01

[0198]

[0199] ADC B-02

[0200]

[0201] ADC B-03

[0202]

[0203] ADC B-04

[0204]

[0205] ADC B-05

[0206]

[0207] ADC B-06

[0208]

[0209] and ADC C-01

[0210]

[0211] Wherein, HA is any antibody disclosed in this application, Represents the connection between a thiol group in an antibody or antigen-binding fragment thereof and a linker.

[0212] In some embodiments, the antibody drug conjugate is selected from ADC A-01 to ADC A-25, ADCB-01 to ADC B-06, and ADC C-01 shown below:

[0213] ADC A-01

[0214]

[0215] ADC A-02

[0216]

[0217] ADC A-03

[0218]

[0219] ADC A-04

[0220]

[0221] ADC A-05

[0222]

[0223] ADC A-06

[0224]

[0225] ADC A-07

[0226]

[0227] ADC A-08

[0228]

[0229] ADC A-09

[0230]

[0231] ADC A-10

[0232]

[0233] ADC A-11

[0234]

[0235] ADC A-12

[0236]

[0237] ADC A-13

[0238]

[0239] ADC A-14

[0240]

[0241] ADC A-15

[0242]

[0243] ADC A-16

[0244]

[0245] ADC A-17

[0246]

[0247] ADC A-18

[0248]

[0249] ADC A-19

[0250]

[0251] ADC A-20

[0252]

[0253] ADC A-21

[0254]

[0255] ADC A-22

[0256]

[0257] ADC A-23

[0258]

[0259] ADC A-24

[0260]

[0261] ADC A-25

[0262]

[0263] ADC B-01

[0264]

[0265] ADC B-02

[0266]

[0267] ADC B-03

[0268]

[0269] ADC B-04

[0270]

[0271] ADC B-05

[0272]

[0273] ADC B-06

[0274]

[0275] and ADC C-01

[0276]

[0277] Wherein, HA is any antibody disclosed in the present invention, Represents the connection between a thiol group in an antibody or antigen-binding fragment thereof and a linker.

[0278] In certain embodiments, the specific connection mode is a connection or bond formed between a thiol group in the antibody or antigen-binding fragment thereof and a linker.

[0279] In another embodiment, the HA is an antibody comprising:

[0280] (1a) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 11, CDR-H2 of SEQ ID NO: 12, and CDR-H3 of SEQ ID NO: 13; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 14, CDR-L2 of SEQ ID NO: 15, and CDR-L3 of SEQ ID NO: 16; or

[0281] (1b) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 27, CDR-H2 of SEQ ID NO: 28, and CDR-H3 of SEQ ID NO: 29; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 30, CDR-L2 of SEQ ID NO: 31, and CDR-L3 of SEQ ID NO: 32;

[0282] (2a) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 17, CDR-H2 of SEQ ID NO: 18 or 19, and CDR-H3 of SEQ ID NO: 13; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 14, CDR-L2 of SEQ ID NO: 15, and CDR-L3 of SEQ ID NO: 16; or

[0283] (2b) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 33, CDR-H2 of SEQ ID NO: 34 or 35, and CDR-H3 of SEQ ID NO: 29; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 30, CDR-L2 of SEQ ID NO: 31, and CDR-L3 of SEQ ID NO: 32;

[0284] (3a) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 20, CDR-H2 of SEQ ID NO: 21, and CDR-H3 of SEQ ID NO: 22; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 23, CDR-L2 of SEQ ID NO: 24, and CDR-L3 of SEQ ID NO: 16; or

[0285] (3b) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 36, CDR-H2 of SEQ ID NO: 37, and CDR-H3 of SEQ ID NO: 38; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 39, CDR-L2 of SEQ ID NO: 40, and CDR-L3 of SEQ ID NO: 32;

[0286] (4a) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 25, CDR-H2 of SEQ ID NO: 26, and CDR-H3 of SEQ ID NO: 13; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 14, CDR-L2 of SEQ ID NO: 15, and CDR-L3 of SEQ ID NO: 16; or

[0287] (4b) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO:41, CDR-H2 of SEQ ID NO:42, and CDR-H3 of SEQ ID NO:29; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO:30, CDR-L2 of SEQ ID NO:31, and CDR-L3 of SEQ ID NO:32.

[0288] In another embodiment, the HA comprises: the VH shown in SEQ ID NO: 1, and / or, the VL shown in SEQ ID NO: 2.

[0289] In another embodiment, the HA comprises: the VH shown in SEQ ID NO: 3, and / or, the VL shown in SEQ ID NO: 4.

[0290] In another embodiment, the HA comprises: the VH shown in SEQ ID NO: 5, and / or, the VL shown in SEQ ID NO: 6.

[0291] In another embodiment, the HA comprises: the VH shown in SEQ ID NO: 7, and / or, the VL shown in SEQ ID NO: 8.

[0292] In another embodiment, the HA comprises: the VH shown in SEQ ID NO: 9, and / or, the VL shown in SEQ ID NO: 10.

[0293] In another embodiment, the HA comprises: (1) a heavy chain comprising a VH sequence shown in SEQ ID NO: 1 and a heavy chain constant region (CH) shown in SEQ ID NO: 43, and a light chain comprising a VL sequence shown in SEQ ID NO: 2 and a light chain constant region (CL) shown in SEQ ID NO: 44.

[0294] In another embodiment, the HA comprises: (2) a heavy chain comprising a VH having a sequence as shown in SEQ ID NO: 3 and a heavy chain constant region (CH) as shown in SEQ ID NO: 43 or 45, and a light chain comprising a VL having a sequence as shown in SEQ ID NO: 4 and a light chain constant region (CL) as shown in SEQ ID NO: 44.

[0295] In another embodiment, the HA comprises: (3) a heavy chain comprising a VH sequence shown in SEQ ID NO: 5 and a heavy chain constant region (CH) shown in SEQ ID NO: 43, and a light chain comprising a VL sequence shown in SEQ ID NO: 6 and a light chain constant region (CL) shown in SEQ ID NO: 44.

[0296] In another embodiment, the HA comprises: (4) a heavy chain comprising a VH sequence shown in SEQ ID NO: 7 and a heavy chain constant region (CH) shown in SEQ ID NO: 43, and a light chain comprising a VL sequence shown in SEQ ID NO: 8 and a light chain constant region (CL) shown in SEQ ID NO: 44.

[0297] In another embodiment, the HA comprises: (5) a heavy chain comprising a VH sequence shown in SEQ ID NO: 9 and a heavy chain constant region (CH) shown in SEQ ID NO: 43, and a light chain comprising a VL sequence shown in SEQ ID NO: 10 and a light chain constant region (CL) shown in SEQ ID NO: 44.

[0298] In another embodiment, the HA comprises: a heavy chain (HC) having the amino acid sequence shown in SEQ ID NO: 46, and a light chain (LC) having the amino acid sequence shown in SEQ ID NO: 47. In some embodiments, the antibody or antigen-binding fragment thereof further comprises:

[0299] (a) a heavy chain constant region (CH) of a human immunoglobulin, or a variant thereof, which has one or more amino acid substitutions, deletions or additions (e.g., up to 20, up to 15, up to 10, or up to 5 amino acid substitutions, deletions or additions; e.g., 1, 2, 3, 4 or 5 amino acid substitutions, deletions or additions) compared to the wild-type sequence from which it is derived; and

[0300] (b) a light chain constant region (CL) of a human immunoglobulin, or a variant thereof, which has one or more amino acid substitutions, deletions or additions (e.g., up to 20, up to 15, up to 10, or up to 5 amino acid substitutions, deletions or additions; for example, 1, 2, 3, 4 or 5 amino acid substitutions, deletions or additions) compared to the wild-type sequence from which it is derived.

[0301] In some embodiments, the HA in the antibody drug conjugate is an antibody or antigen-binding fragment, wherein the VH is a VH of SEQ ID NO: 1, 3, 5, 7 or 9 and the VL is a VL of SEQ ID NO: 2, 4, 6, 8 or 10, for example, the VH of the antibody or antigen-binding fragment is a VH of SEQ ID NO: 3 and the CH is a CH of SEQ ID NO: 43 or 45, the VL is a VL of SEQ ID NO: 4 and the CL is a CL of SEQ ID NO: 44; and

[0302] Represents the connection between a thiol group in an antibody or antigen-binding fragment thereof and a linker.

[0303] In certain embodiments of the HA or antibody drug conjugates described herein, the heavy chain constant region may comprise a C-terminal lysine, or lack a C-terminal lysine or a C-terminal glycine-lysine dipeptide. In some embodiments of the antibodies or antigen-binding fragments described herein, the N-terminal amino acid of the antibody variable region may be cyclized to pyroglutamate.

[0304] In some embodiments of the antibodies or antigen-binding fragments described herein, the N-terminal amino acid of the antibody variable region can be cyclized to pyroglutamic acid.

[0305] Thus, the composition may comprise various antibody drug conjugates, wherein each species may independently comprise a C-terminal lysine, lack a C-terminal lysine, lack a C-terminal glycine-lysine, and / or comprise an N-terminal glutamine or glutamic acid, or pyroglutamic acid resulting from cyclization of the N-terminal amino acid.

[0306] In certain embodiments, the composition may comprise various antibody drug conjugates, wherein each species may independently comprise a C-terminal lysine, lack a C-terminal lysine, lack a C-terminal glycine-lysine, and / or comprise an N-terminal glutamine or glutamic acid, or pyroglutamic acid resulting from cyclization of the N-terminal amino acid.

[0307] Therefore, in certain embodiments, the present invention further provides a composition comprising the ADC disclosed herein, wherein the major ADC species in the composition comprises: (i) an antibody with a lysine residue deleted from the C-terminus of the heavy chain; (ii) an antibody with glutamine, glutamic acid, or pyroglutamate at the N-terminus of the heavy chain; or (iii) an antibody with a lysine residue deleted from the C-terminus of the heavy chain and glutamine, glutamic acid, or pyroglutamate at the N-terminus of the heavy chain.

[0308] In certain embodiments, the present invention further provides a composition comprising the ADC disclosed herein, wherein the major ADC species in the composition comprises: (i) an antibody with a lysine residue deleted at the C-terminus of the heavy chain; (ii) an antibody with glutamine, glutamic acid, or pyroglutamate at the N-terminus of the heavy chain; or (iii) an antibody with a lysine residue deleted at the C-terminus of the heavy chain and glutamine, glutamic acid, or pyroglutamate at the N-terminus of the heavy chain.

[0309] It is known to those skilled in the art that pyroglutamic acid is the conjugate acid of pyroglutamate and is in equilibrium with pyroglutamate in solution.

[0310] Composition

[0311] In another aspect, the present application provides a composition of an antibody drug conjugate (ADC) as described herein. The composition may comprise a plurality of ADCs as described herein, wherein each ADC comprises a drug linker as described herein. Thus, the composition is characterized by a drug-to-antibody ratio (DAR) in the range of about 1 to about 10. Methods for determining the DAR are well known to those skilled in the art and include methods such as reverse phase chromatography or HPLC-MS.

[0312] In some embodiments, the DAR value (drug-antibody conjugate ratio) of the antibody drug conjugate composition is 1-10, for example: 1-2, 1-3, 1-4, 1-5, 1-6, 1-7, 1-8, 1-9, 1-10, 2-3, 2-4, 2-5, 2-6, 2-7, 2-8, 2-9, 2-10, 3-4, 3-5, 3-6, 3-7, 3-8, 3-9, 3-10, 4-5, 4-6, 4-7, 4-8, 4-9, 4 ~10, 5~6, 5~7, 5~8, 5~9, 5~10, 6~7, 6~8, 6~9, 6~10, 7~8, 7~9, 7~10, 8~9, 8~10, or 9~10, preferably 1~8, preferably 3~9, for example, 3.0~3.5, 3.0~4.0, 3.0~4.5, 3.0~5.0, 3.0~5.5, 3.0~6.0, 3.5~4.0, 3.5~4.5, 3.5~5.0, 3.5~5. 5,3.5~6.0,3.5~6.5,3.5~7.0,3.5~7.5,3.5~8.0,4.0~4.5,4.0~5.0,4.0~5.5,4.0~6.0,4.0~6.5,4.0~7.0,4.0~7.5,4.0~8.0,4.5~5.0,4.5~5.5,4.5~6.0,4.5~6.5,4.5~7.0,4.5~7.5,4.5~8.0,5.0 ~5.5, 5.0~6.0, 5.0~6.5, 5.0~7.0, 5.0~7.5, 5.0~8.0, 5.5~6.0, 5.5~6.5, 5.5~7.0, 5.5~7.5, 5.5~8.0, 6.0~6.5, 6.0~7.0, 6.0~7.5, 6.0~8.5, 6.5~7.0, 6.5~7.5, 6.5~8.5, 7.0~7.5, 7.0~9.0 or 7.5~9.0.

[0313] In one embodiment, the DAR value is 8. In one embodiment, the DAR value is 7 to 9. In one embodiment, the DAR value is 7 to 8. In other embodiments, the DAR value of the antibody drug conjugate composition comprising ADC A-05 is about 8. In other embodiments, the DAR value of the antibody drug conjugate composition comprising ADC A-05 is 7 to 9. In other embodiments, the DAR value of the antibody drug conjugate composition comprising ADC A-05 is 7 to 8. In other embodiments, the DAR value of the antibody drug conjugate composition comprising ADC B-01 is 7 to 8. In other embodiments, the DAR value of the antibody drug conjugate composition comprising ADC C-01 is 2 to 4. In other embodiments, the DAR value of the antibody drug conjugate composition comprising ADC B-04 is 7 to 9. In other embodiments, the DAR value of the antibody drug conjugate composition comprising ADC A-26 is 7 to 9.

[0314] Drug-linker

[0315] Those skilled in the art will appreciate that the antibody-drug conjugates described herein can be prepared modularly. For example, a free-form "drug-linker" (which can be understood as M'-LED, where M' is the structural form of M before covalently linking to the antibody or its antigen-binding fragment) is first obtained, and then covalently linked to the antibody or its antigen-binding fragment to obtain the antibody-drug conjugate described herein. Accordingly, M' in the free-form "drug-linker" is linked to one or more sulfhydryl (-SH), amino (-NH2) or carboxyl (-COOH) groups on the antibody or its antigen-binding fragment by a substitution reaction (e.g., removal of structures such as -SO2Me or -Br thereon) or by an addition reaction.

[0316] In another aspect, the present invention provides a drug-linker having a structure shown in formula M'-LED, wherein:

[0317] M' is Lg is a leaving group for nucleophilic substitution reaction (e.g., halogen, methanesulfonyl, fluorophenol or ), or is hydroxyl (-OH), thiol (-SH) or amino (-NH2); or, Lg forms an unsaturated double bond with the adjacent atoms on ring A; Ring A is a 5-6 membered alicyclic heterocyclic ring, or a 5-20 membered aromatic ring system, wherein the alicyclic heterocyclic ring and the aromatic ring system are optionally substituted with one or more selected from oxy (=O), halogen, cyano, amino, carboxyl, thiol and C 1-6 Alkyl group substituted; M1 is selected from single bond and C 1-20 Alkylene, C 2-20 Alkenylene or C 2-20 Alkynylidene;

[0318] The L, E and D structures are as defined in any of the above antibody drug conjugates.

[0319] In some embodiments, M' is Lg is a methylsulfonyl group, or Lg forms a carbon-carbon double bond with the adjacent atoms on ring A; Ring A is a 5-membered alicyclic heterocycle, a 6-membered heteroaromatic ring, or a polycyclic ring formed by connecting one or more 6-membered heteroaromatic rings to a benzene ring via a single bond, wherein the alicyclic heterocycle is optionally substituted with one or more selected from oxy (=O), halogen and C 1-4 Alkyl group substituted; M1 is selected from single bond and C 3-10 Alkylene, C 3-10 Alkenylene or C 3-10 Alkynylidene.

[0320] In some embodiments, M' is Selected from M1 is selected from a single bond and C 5-8 Alkylene, C 5-8 Alkenylene or C 5-8 Alkynylidene.

[0321] In some embodiments, M' is selected from

[0322] In some embodiments, M' is

[0323] In some embodiments, the free form of the "drug-linker" is selected from A-01 to A-26, B-01 to B-06, and C-01 shown below:

[0324] A-01:

[0325] A-02:

[0326] A-03:

[0327] A-04:

[0328] A-05:

[0329] A-06:

[0330] A-07:

[0331] A-08:

[0332] A-09:

[0333] A-10:

[0334] A-11:

[0335] A-12:

[0336] A-13:

[0337] A-14:

[0338] A-15:

[0339] A-16:

[0340] A-17:

[0341] A-18:

[0342] A-19:

[0343] A-20:

[0344] A-21:

[0345] A-22:

[0346] A-23:

[0347] A-24:

[0348] A-25:

[0349] A-26:

[0350] B-01:

[0351] B-02:

[0352] B-03:

[0353] B-04:

[0354] B-05:

[0355] B-06:

[0356] C-01:

[0357] In some embodiments, the free form of the "drug-linker" is selected from A-01 to A-25, B-01 to B-06, and C-01 shown below:

[0358] A-01:

[0359] A-02:

[0360] A-03:

[0361] A-04:

[0362] A-05:

[0363] A-06:

[0364] A-07:

[0365] A-08:

[0366] A-09:

[0367] A-10:

[0368] A-11:

[0369] A-12:

[0370] A-13:

[0371] A-14:

[0372] A-15:

[0373] A-16:

[0374] A-17:

[0375] A-18:

[0376] A-19:

[0377] A-20:

[0378] A-21:

[0379] A-22:

[0380] A-23:

[0381] A-24:

[0382] A-25:

[0383] B-01:

[0384] B-02:

[0385] B-03:

[0386] B-04:

[0387] B-05:

[0388] B-06:

[0389] C-01:

[0390] Pharmaceutical composition

[0391] In another aspect, the present application provides a pharmaceutical composition comprising the antibody-drug conjugate described in any one of the foregoing items and optionally the drug-linker described in any one of the foregoing items, and one or more pharmaceutically acceptable excipients.

[0392] The antibody drug conjugates described herein are generally formulated in a unit injectable form together with a pharmaceutically acceptable parenteral vehicle for parenteral use, such as bolus injection, intravenous injection, intratumoral injection, etc. Optionally, the antibody drug conjugate having the desired purity is mixed with a pharmaceutically acceptable diluent, carrier, excipient or stabilizer in the form of a lyophilized agent or solution (Remington's Pharmaceutical Sciences (1980) 16 th The antibody drug conjugates described herein or pharmaceutical compositions containing the same can be administered by any route appropriate to the subject to be treated.

[0393] application

[0394] The antibody-drug conjugates or pharmaceutical compositions thereof described herein can be used to treat a variety of diseases or conditions, such as PTK7-overexpressing cancers, including solid tumors or hematological malignancies, such as lung cancer, breast cancer, epidermal cancer (e.g., squamous cell carcinoma of the skin, oral squamous cell carcinoma), ovarian cancer, esophageal cancer (e.g., esophageal squamous cell carcinoma), etc.

[0395] Therefore, the present application provides the use of an antibody-drug conjugate, a drug-linker, or a pharmaceutical composition containing the same in the preparation of a drug for treating cancers with high PTK7 expression.

[0396] At the same time, the present application also provides a method for treating PTK7-overexpressing cancer, comprising the steps of administering to a subject in need thereof an effective amount of any of the above-described antibody-drug conjugates, drug linkers, or pharmaceutical compositions containing the same. In a specific embodiment, the cancer comprises a solid tumor or a hematological malignancy. In another embodiment, the cancer is lung cancer, breast cancer, epidermal cancer (e.g., squamous cell carcinoma of the skin, oral squamous cell carcinoma), ovarian cancer, esophageal cancer (e.g., esophageal squamous cell carcinoma), etc. BRIEF DESCRIPTION OF THE DRAWINGS

[0397] Figure 1A : Pharmacological efficacy testing of different antibody-drug conjugates in the NCI-H358 model.

[0398] Figure 1B : Body weight detection of NCI-H358 model with different antibody-drug conjugates.

[0399] Figure 2A : Pharmacological efficacy testing of different antibody-drug conjugates A431 models.

[0400] Figure 2B : Body weight detection of different antibody-drug conjugate A431 models.

[0401] Figure 3A : Detection of binding of anti-human PTK7 antibody-drug conjugate to OVCAR3 cells.

[0402] Figure 3B : Detection of binding of anti-human PTK7 antibody-drug conjugates to NCI-H358 cells.

[0403] Figure 3C : Detection of binding of anti-human PTK7 antibody-drug conjugate to HCC1806 cells.

[0404] Figure 3D : Detection of binding of anti-human PTK7 antibody-drug conjugates to A431 cells.

[0405] Figure 3E : Detection of binding of anti-human PTK7 antibody-drug conjugates to NCI-H520 cells.

[0406] Figure 3F : Detection of binding of anti-human PTK7 antibody-drug conjugates to DU4475 cells.

[0407] Figure 4A : pHrodo assay for endocytosis of antibody-drug conjugates in NCI-H358.

[0408] Figure 4B : pHrodo assay of antibody drug conjugate endocytosis in HCC1806.

[0409] Figure 4C : pHrodo assay for endocytosis of antibody-drug conjugates in OVCAR3.

[0410] Figure 4D : pHrodo assay for endocytosis of antibody-drug conjugates in NCI-H520.

[0411] Figure 4E : pHrodo assay of antibody drug conjugate endocytosis in DU4475.

[0412] Figure 5A : Detection of anti-human PTK7 conjugated drugs killing FADU cells.

[0413] Figure 5B : Detection of anti-human PTK7 conjugated drugs killing HCC1806 cells.

[0414] Figure 5C : Detection of anti-human PTK7 conjugate drug killing DU4475 cells.

[0415] Figure 6A : Pharmacological efficacy testing of antibody-drug conjugate HCC1806 model.

[0416] Figure 6B : Body weight detection of antibody-drug conjugate HCC1806 model.

[0417] Figure 7A : Efficacy testing of antibody-drug conjugate OVCAR3 model.

[0418] Figure 7B : Body weight detection of antibody-drug conjugate OVCAR3 model.

[0419] Figure 8A : Efficacy testing of antibody-drug conjugates in the NCI-H146 model.

[0420] Figure 8B : Body weight detection of antibody-drug conjugate NCI-H146 model.

[0421] Specific description

[0422] definition

[0423] Unless otherwise defined below, the meanings of all technical and scientific terms used herein are intended to be the same as those generally understood by those skilled in the art. Reference to the techniques used herein is intended to refer to techniques generally understood in the art, including variations of those techniques that are obvious to those skilled in the art or replacements with equivalent techniques. Furthermore, laboratory procedures such as genomics, nucleic acid chemistry, and molecular biology used herein are conventional procedures widely used in the corresponding fields. Although it is believed that the following terms are well understood by those skilled in the art, the following definitions are still set forth to better explain the present invention.

[0424] The term "antibody" refers to an immunoglobulin molecule typically composed of two pairs of polypeptide chains, each pair having one light chain (LC) and one heavy chain (HC). Antibody light chains can be classified as kappa (κ) and lambda (λ) light chains. Heavy chains can be classified as μ, δ, γ, α, or ε, 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 connected by a "J" region of approximately 12 or more amino acids, with heavy chains also containing a "D" region of approximately 3 or more amino acids. Each heavy chain consists of a heavy chain variable region (VH) and a heavy chain constant region (CH). The heavy chain constant region is composed of three domains: CH1, CH2, and CH3. Each light chain consists of a light chain variable region (VL) and a light chain constant region (CL). The light chain constant region is composed of one domain, CL. The constant domain is not directly involved in the binding of antibodies to antigens, but exhibits a variety of effector functions, such as mediating the binding of immunoglobulins to host tissues or factors, including various cells of the immune system (e.g., effector cells) and the first component (C1q) of the classical complement system. The VH and VL regions can also be subdivided into regions of high variability, called complementarity determining regions (CDRs), interspersed with more conserved regions called framework regions (FRs). Each VH and VL consists of three CDRs and four FRs, arranged in the following order: FR1, CDR1, FR2, CDR2, FR3, CDR3, FR4 from the amino terminus to the carboxyl terminus. The variable regions (VH and VL) of each heavy chain / light chain pair form the antigen binding site, respectively. The allocation of amino acids to each region or domain can follow various numbering systems known in the art.

[0425] The term "complementarity determining region" or "CDR" refers to the amino acid residues in the variable region of an antibody that are responsible for antigen binding. The variable region of the heavy chain and light chain each contains three CDRs, designated CDR1, CDR2, and CDR3. The precise boundaries of these CDRs can be defined according to various numbering systems known in the art, for example, as defined in the Kabat numbering system (Kabat et al., Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, Md., 1991), the Chothia numbering system (Chothia & Lesk (1987) J. Mol. Biol. 196:901-917; Chothia et al. (1989) Nature 342:878-883), the IMGT numbering system (Lefranc et al., Dev. Comparat. Immunol. 27:55-77, 2003), or the AbM numbering system (Martin ACR, Cheetham JC, Rees AR (1989) Modelling antibody hypervariable loops: A combined algorithm. Proc Natl Acad Sci USA 86:9268-9272). For a given antibody, a person skilled in the art will readily identify the CDRs defined by each numbering system. Furthermore, the correspondence between different numbering systems is well known to those skilled in the art (e.g., see Lefranc et al., Dev. Comparat. Immunol. 27:55-77, 2003).

[0426] In the present invention, the CDRs contained in an antibody or antigen-binding fragment thereof can be determined according to various numbering systems known in the art, such as those defined by the Kabat, Chothia, IMGT, or AbM numbering systems. In certain embodiments, the CDRs contained in an antibody or antigen-binding fragment thereof are defined by the Chothia numbering system.

[0427] The following general rules (published at www.bioinf.org.uk: Professor Andrew CR Martin's research group) can be used to define CDRs in antibody sequences, which include amino acids that specifically interact with amino acids that make up the epitope to which the antibody binds. In rare cases, these generally constant features do not appear; however, Cys residues are the most conserved feature.

[0428]

[0429] The entire amino acid sequence of a VH is generally numbered according to Kabat, while the three CDRs within the variable region can be defined according to any of the above numbering systems. In certain embodiments, amino acid positions in a VH can be numbered sequentially starting from amino acid position 1 and continuing to the end of the sequence, or numbered according to Kabat. Unless otherwise indicated, amino acid positions in VH and VL described herein are defined according to sequential numbering.

[0430] Amino acid positions in the heavy chain constant region can be numbered sequentially starting from amino acid position 1 and continuing to the end of the sequence, or numbered according to Eu. The amino acid sequence of the IgG1 heavy chain constant region has 330 amino acids, numbered sequentially from 1 to 330. The corresponding sequence numbered according to Eu begins at position 118 and ends at position 447. Unless otherwise indicated, amino acid positions in the heavy and light chains described herein are defined according to sequential numbering.

[0431] The term "framework region" or "FR" residues refers to those amino acid residues in the variable region of an antibody other than the CDR residues as defined above.

[0432] The term "antigen-binding fragment" of an antibody refers to polypeptides that are fragments of an antibody, such as polypeptides that are fragments of a full-length antibody, which retain the ability to specifically bind to the same antigen bound by the full-length antibody and / or compete with the full-length antibody for specific binding to the antigen, and are also referred to as "antigen-binding portions." See generally, Fundamental Immunology, Ch. 7 (Paul, W., ed., 2nd ed., Raven Press, NY (1989), which is incorporated herein by reference in its entirety for all purposes. Antigen-binding fragments of antibodies can be produced by recombinant DNA techniques or by enzymatic or chemical cleavage of intact antibodies. Non-limiting examples of antigen-binding fragments include Fab fragments, Fab' fragments, F(ab)'2 fragments, F(ab)'3 fragments, Fd, Fv, scFv, di-scFv, (scFv)2, disulfide-stabilized Fv proteins ("dsFv"), single domain antibodies (sdAbs, nanobodies), and polypeptides that comprise at least a portion of an antibody sufficient to confer specific antigen-binding ability on the polypeptide. Engineered antibody variants are reviewed in Holliger et al., 2005; Nat Biotechnol, 23: 1126-1136.

[0433] The term "Fd" means an antibody fragment consisting of the VH and CH1 domains; the term "dAb fragment" means an antibody fragment consisting of the VH domain (Ward et al., Nature 341:544-546 (1989)); the term "Fab fragment" means an antibody fragment consisting of the VL, VH, CL and CH1 domains; the term "F(ab')2 fragment" means an antibody fragment comprising two Fab fragments linked by a disulfide bridge at the hinge region; the term "Fab' fragment" means a fragment obtained after reducing the disulfide bonds linking the two heavy chain fragments in the F(ab')2 fragment, consisting of one complete light chain and the Fd fragment (consisting of the VH and CH1 domains) of the heavy chain.

[0434] The term "Fv" refers to an antibody fragment consisting of the VL and VH domains of a single arm of an antibody. The Fv fragment is generally considered to be the smallest antibody fragment that can form a complete antigen-binding site. It is generally believed that the six CDRs confer antigen-binding specificity to an antibody. However, even a single variable region (e.g., an Fd fragment, which contains only three CDRs specific for an antigen) can recognize and bind to an antigen, although its affinity may be lower than that of the complete binding site.

[0435] The term "Fc" refers to an antibody fragment formed by disulfide bonds between the second and third constant regions of the first heavy chain and the second and third constant regions of the second heavy chain. The Fc fragment of an antibody has various functions but is not involved in antigen binding.

[0436] The term "scFv" refers to a single polypeptide chain comprising a VL and VH domain, wherein the VL and VH are connected by a linker (see, e.g., Bird et al., Science 242: 423-426 (1988); Huston et al., Proc. Natl. Acad. Sci. USA 85: 5879-5883 (1988); and Pluckthun, The Pharmacology of Monoclonal Antibodies, Vol. 113, Roseburg and Moore, eds., Springer-Verlag, New York, pp. 269-315 (1994)). Such scFv molecules may have the general structure: NH2-VL-linker-VH-COOH or NH2-VH-linker-VL-COOH. Suitable linkers in the art include repeated GGGGS (SEQ ID NO: 55) amino acid sequences or variants thereof. For example, a linker having the amino acid sequence (GGGGS)4 (SEQ ID NO: 56) can be used, although variants thereof can also be used (Holliger et al. (1993), Proc. Natl. Acad. Sci. USA 90: 6444-6448). Other linkers useful in the present invention are described by Alfthan et al. (1995), Protein Eng. 8: 725-731, Choi et al. (2001), Eur. J. Immunol. 31: 94-106, Hu et al. (1996), Cancer Res. 56: 3055-3061, Kipriyanov et al. (1999), J. Mol. Biol. 293: 41-56, and Roovers et al. (2001), Cancer Immunol. In some cases, a disulfide bond may also be present between the VH and VL of the scFv. In certain embodiments, the VH and VL domains can be positioned relative to each other in any suitable arrangement. For example, a VH-VH-COOH domain comprising NH2-VH-VH-COOH, NH 2- VL-VL-COOH scFv.

[0437] The term "single-domain antibody (sdAb)" has the meaning generally understood by those skilled in the art, and refers to an antibody fragment composed of a single monomeric variable antibody domain (e.g., a single heavy chain variable region), and such an antibody fragment retains the ability to specifically bind to the same antigen as the full-length antibody (Holt, L. et al., Trends in Biotechnology, 21(11):484-490, 2003). Single-domain antibodies are also called nanobodies.

[0438] Each of the above antibody fragments retains the ability to specifically bind to the same antigen as the full-length antibody, and / or the ability to compete with the full-length antibody for specific binding to the antigen.

[0439] Herein, unless the context clearly indicates otherwise, when referring to the term "antibody", it includes not only intact antibodies, but also antigen-binding fragments of antibodies.

[0440] Antigen-binding fragments of antibodies (e.g., those described above) can be obtained from a given antibody (e.g., an antibody provided herein) using conventional techniques known to those skilled in the art (e.g., recombinant DNA technology or enzymatic or chemical cleavage methods), and the antigen-binding fragments of antibodies can be screened for specificity in the same manner as for intact antibodies.

[0441] The term "murine antibody" refers to antibodies obtained by fusing B cells from immunized mice with myeloma cells, screening for murine hybrid fusion cells that can both proliferate indefinitely and secrete antibodies, followed by screening, antibody preparation, and antibody purification; or refers to antibodies secreted by plasma cells formed by the differentiation and proliferation of B cells in mice after antigen invasion.

[0442] The term "humanized antibody" refers to a non-human antibody that has been genetically engineered and whose amino acid sequence has been modified to increase the homology with the sequence of a human antibody. Generally speaking, all or part of the CDR region of a humanized antibody comes from a non-human antibody (donor antibody), and all or part of the non-CDR region (e.g., variable region FR and / or constant region) comes from a human immunoglobulin (recipient antibody). Humanized antibodies generally retain the expected properties of the donor antibody, including but not limited to, antigen specificity, affinity, reactivity, ability to increase immune cell activity, ability to enhance immune response, etc. The donor antibody can be a mouse, rat, rabbit or non-human primate (e.g., cynomolgus monkey) antibody with the expected properties (e.g., antigen specificity, affinity, reactivity, ability to increase immune cell activity and / or ability to enhance immune response).

[0443] The term "identity" is used to refer to the match of sequences between two polypeptides or between two nucleic acids. When a position in both sequences being compared is occupied by the same base or amino acid monomer subunit (e.g., a position in each of the two DNA molecules is occupied by adenine, or a position in each of the two polypeptides is occupied by lysine), then the molecules are identical at that position. "Percent identity" between two sequences is the function obtained by the following formula: the number of matching positions shared by the two sequences divided by the number of positions compared × 100. For example, if 6 out of 10 positions in two sequences match, then the two sequences have 60% identity. For example, the DNA sequences CTGACT and CAGGTT share 50% identity (3 out of 6 total positions match). Typically, two sequences are compared when they are aligned for maximum identity. Such alignment can be achieved, for example, by using the method of Needleman et al. (1970) J. Mol. Biol. 48:443-453, which can be conveniently performed using a computer program such as the Align program (DNAstar, Inc.). The percent identity between two amino acid sequences can also be determined using the algorithm of E. Meyers and W. Miller (Comput. Appl Biosci., 4:11-17 (1988)), which has been incorporated into the ALIGN program (version 2.0), using a PAM120 weight residue table, a gap length penalty of 12, and a gap penalty of 4. In addition, the percent identity between two amino acid sequences can be determined using the Needleman and Wunsch (J Mol. Biol. 48:444-453 (1970)) algorithm, which has been incorporated into the GAP program in the GCG software package (available at www.gcg.com), using either a Blossum 62 matrix or a PAM250 matrix and a gap weight of 16, 14, 12, 10, 8, 6, or 4 and a length weight of 1, 2, 3, 4, 5, or 6.

[0444] The term "conservative substitution" means an amino acid substitution that does not adversely affect or change the expected properties of the protein / polypeptide comprising the amino acid sequence. For example, conservative substitutions can be introduced by standard techniques known in the art such as site-directed mutagenesis and PCR-mediated mutagenesis. Conservative amino acid substitutions include substitutions in which amino acid residues are substituted with amino acid residues having similar side chains, such as substitutions with residues that are physically or functionally similar to the corresponding amino acid residues (e.g., having similar size, shape, charge, chemical properties, including the ability to form covalent bonds or hydrogen bonds, etc.). Families of amino acid residues with similar side chains have been defined in the art. These families include amino acids with basic side chains (e.g., lysine, arginine, and histidine), acidic side chains (e.g., aspartic acid, glutamic acid), uncharged polar side chains (e.g., glycine, asparagine, glutamine, serine, threonine, tyrosine, cysteine, tryptophan), nonpolar side chains (e.g., alanine, valine, leucine, isoleucine, proline, phenylalanine, methionine), beta-branched side chains (e.g., threonine, valine, isoleucine), and aromatic side chains (e.g., tyrosine, phenylalanine, tryptophan, histidine). Thus, it is preferred to replace the corresponding amino acid residue with another amino acid residue from the same side chain family. Methods for identifying conservative amino acid substitutions are well known in the art (see, e.g., Brummell et al., Biochem. 32: 1180-1187 (1993); Kobayashi et al. Protein Eng. 12(10): 879-884 (1999); and Burks et al. Proc. Natl Acad. Set USA 94: 412-417 (1997), which are incorporated herein by reference).

[0445] The twenty conventional amino acids referred to herein are denoted according to conventional usage. See, for example, Immunology-A Synthesis (2nd Edition, E.S. Golub and D.R. Gren, Eds., Sinauer Associates, Sunderland, Mass. (1991)), which is incorporated herein by reference. In the present invention, amino acids are generally represented by single-letter and three-letter abbreviations known in the art. For example, alanine can be represented by A or Ala.

[0446] The term "pharmaceutically acceptable carrier and / or excipient" refers to a carrier and / or excipient that is pharmacologically and / or physiologically compatible with the subject and the active ingredient, which is well known in the art (see, for example, Remington's Pharmaceutical Sciences. Edited by Gennaro AR, 19th edition. Pennsylvania: Mack Publishing Company, 1995), and includes, but is not limited to, pH regulators, surfactants, adjuvants, ionic strength enhancers, diluents, agents that maintain osmotic pressure, agents that delay absorption, and preservatives. For example, pH regulators include, but are not limited to, phosphate buffers. Surfactants include, but are not limited to, cationic, anionic, or nonionic surfactants, such as Tween-80. Ionic strength enhancers include, but are not limited to, sodium chloride. Preservatives include, but are not limited to, various antibacterial and antifungal agents, such as parabens, chlorobutanol, phenol, sorbic acid, and the like. Agents that maintain osmotic pressure include, but are not limited to, sugars, sodium chloride, and the like. Agents that delay absorption include, but are not limited to, monostearate and gelatin. Diluents include, but are not limited to, water, aqueous buffers (such as buffered saline), alcohols, and polyols (such as glycerol). Preservatives include, but are not limited to, various antibacterial and antifungal agents, such as thimerosal, 2-phenoxyethanol, parabens, chlorobutanol, phenol, sorbic acid, and the like. Stabilizers have the meanings generally understood by those skilled in the art, and can stabilize the desired activity of the active ingredient in the drug, including, but not limited to, sodium glutamate, gelatin, SPGA, sugars (such as sorbitol, mannitol, starch, sucrose, lactose, dextran, or glucose), amino acids (such as glutamic acid, glycine), proteins (such as dry whey, albumin, or casein), or degradation products thereof (such as whey protein hydrolyzate), and the like.

[0447] The terms "comprises," "comprising," "having," "containing," or "involving," and other variations thereof herein, are inclusive or open-ended and do not exclude additional unrecited elements or method steps.

[0448] The term "effective amount" refers to an amount sufficient to achieve the desired clinical effect in the individual being treated. For example, this may be the amount necessary to alleviate any particular disease symptom or to inhibit or reduce the severity of the disease or its complications. Determining such an effective amount is entirely within the capabilities of those skilled in the art. For example, the effective amount for therapeutic use will depend on the severity of the disease to be treated, the general state of the patient's own immune system, the patient's general condition (e.g., age, weight, and sex), the mode of administration of the drug, and other treatments administered simultaneously. The therapeutically effective amount of an ADC may vary according to the following factors: the severity of the disease to be treated, the general state of the patient's own immune system, the patient's general condition such as age, weight, and sex, the mode of administration of the drug, and other treatments administered simultaneously.

[0449] The term "treatment" refers to methods implemented to obtain a beneficial or desired clinical outcome in a subject (human or animal subject) who exhibits symptoms of a disease or has been diagnosed with a disease ("in need of treatment"). For purposes of the present invention, beneficial or desired clinical outcomes include, but are not limited to, relief of symptoms, reduction in the extent of the disease, stabilization of the disease state (i.e., no worsening), delay or slowing of disease progression, improvement or alleviation of the disease state, and relief of symptoms (whether partial or complete), whether detectable or not. In addition, "treatment" may also refer to prolonging survival compared to the expected survival if not receiving treatment.

[0450] As used herein, the term "subject" refers to a mammal, such as a primate mammal, such as a human. In certain embodiments, the subject (eg, human) suffers from a tumor, or is at risk of suffering from the above-mentioned disease.

[0451] The terms "cancer" and "tumor" are used interchangeably to refer to a broad category of diseases characterized by the uncontrolled growth of abnormal cells in the body. Uncontrolled cell division may lead to the formation of malignant tumors, or cells that invade neighboring tissues and may metastasize to distant sites in the body via the lymphatic system or bloodstream. Cancer includes both benign and malignant cancers, as well as dormant tumors or micrometastases. Cancer also includes hematologic malignancies.

[0452] The term "hematological malignancy" includes lymphomas, leukemias, myelomas or lymphoid malignancies, as well as tumors of the spleen and lymph nodes. Exemplary lymphomas include B-cell lymphomas and T-cell lymphomas. B-cell lymphomas include, for example, Hodgkin's lymphoma. T-cell lymphomas include, for example, cutaneous T-cell lymphoma. Hematological malignancies also include leukemias, such as secondary leukemias or acute lymphoblastic leukemia. Hematological malignancies also include myeloma (e.g., multiple myeloma) and other blood and / or B-cell or T-cell related cancers.

[0453] The term "alkyl" refers to a group obtained by removing one hydrogen atom from a straight-chain or branched hydrocarbon group, for example, "C 1-20 Alkyl", "C 1-10 Alkyl", "C 1-6 Alkyl", "C 1-4 Alkyl", "C 1-3alkyl”, etc., specific examples include, but are not limited to, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, n-pentyl, isopentyl, 2-methylbutyl, neopentyl, 1-ethylpropyl, n-hexyl, isohexyl, 3-methylpentyl, 2-methylpentyl, 1-methylpentyl, 3,3-dimethylbutyl, 2,2-dimethylbutyl, 1,1-dimethylbutyl, 1,2-dimethylbutyl, 1,3-dimethylbutyl, 2,3-dimethylbutyl, 2-ethylbutyl, 1,2-dimethylpropyl, etc.

[0454] The term "alkylene" refers to a group obtained by removing two hydrogen atoms from a straight or branched hydrocarbon group, for example, "C 1-20 Alkylene", "C 1-10 Alkylene", "C 3-10 Alkylene", "C 5-8 Alkylene", "C 1-6 Alkylene", "C 1-4 Alkylene", "C 1-3 Specific examples include, but are not limited to, methylene, ethylene, 1,3-propylene, 1,4-butylene, 1,5-pentylene or 1,6-hexylene.

[0455] The term "alkenylene" refers to a divalent group derived from a straight or branched hydrocarbon group containing at least one carbon-carbon double bond losing two hydrogen atoms, including, for example, "C 2-20 Alkenylene", "C 3-10 Alkenylene", "C 5-8 Examples include, but are not limited to, vinylene, 1-propenylene, 2-propenylene, 1-butenylene, 2-butenylene, 1,3-butadienylene, 1-pentenylene, 2-pentenylene, 3-pentenylene, 1,3-pentadienylene, 1,4-pentadienylene, 1-hexenylene, 2-hexenylene, 3-hexenylene, 1,4-hexadienylene, and the like.

[0456] The term "alkynylene" refers to a divalent group derived from a straight or branched hydrocarbon group containing at least one carbon-carbon triple bond that loses two hydrogen atoms. 2-20 Alkynylidene", "C 3-10 Alkynylidene", "C 5-8 Examples include, but are not limited to, ethynylene, 1-propynylene, 2-propynylene, 1-butynylene, 2-butynylene, 1,3-butadiynylene, 1-pentynylene, 2-pentynylene, 3-pentynylene, 1,3-pentadiynylene, 1,4-pentadiynylene, 1-hexynylene, 2-hexynylene, 3-hexynylene, 1,4-hexadiynylene, and the like.

[0457] The term "aliphatic heterocycle" refers to a saturated or partially saturated cyclic structure containing at least one ring member selected from N, O, and S. Specific examples include, but are not limited to, 5-6 membered aliphatic heterocycles, 5-6 membered nitrogen-containing aliphatic heterocycles, 5-6 membered oxygen-containing aliphatic heterocycles, and the like, such as tetrahydrofuran, pyrrolidine, piperidine, tetrahydropyran, and the like.

[0458] The term "heteroaromatic ring" refers to an aromatic ring structure containing at least one ring member selected from N, O and S. Specific examples include, but are not limited to, 5-6 membered aromatic heterocycles, 5-6 membered nitrogen-containing aromatic heterocycles, 5-6 membered oxygen-containing aromatic heterocycles, and the like, such as furan, thiophene, pyrrole, thiazole, isothiazole, thiadiazole, oxazole, isoxazole, oxadiazole, imidazole, pyrazole, 1,2,3-triazole, 1,2,4-triazole, 1,2,3-oxadiazole, 1,2,4-oxadiazole, 1,2,5-oxadiazole, 1,3,4-oxadiazole, pyridine, pyrimidine, pyridazine, pyrazine, 1,2,3-triazine, 1,3,5-triazine, 1,2,4,5-tetrazine, and the like.

[0459] The term "aromatic ring system" refers to a monocyclic or polycyclic ring system comprising at least one aromatic ring (e.g., a benzene ring, etc.) or heteroaromatic ring (e.g., a pyrimidine ring, etc.), two or more aromatic rings and / or heteroaromatic rings may form a fused ring or be connected by a single bond (e.g., dipyrimidinylphenyl, etc.), and the aromatic ring system may be divalent or higher valent (e.g., trivalent or tetravalent), for example, a 5-20 membered aromatic ring system.

[0460] As used herein, "Cit" is an abbreviation for citrulline.

[0461] As used herein, the terms "about" or "approximately" when used in connection with a numerical variable generally mean that the value of the variable is within the range of experimental error (eg, within a 95% confidence interval about the mean) or within ±10%.

[0462] It is important to note that if there is a discrepancy between the depicted structure and the name of that structure, the depicted structure should be given greater weight.

[0463] Antibody Drug Conjugates

[0464] In one aspect, the present application provides an antibody drug conjugate having the formula Ab-[MLED] x The structure shown, wherein:

[0465] Ab is an antibody or antigen-binding fragment thereof that specifically binds to human tyrosine kinase 7 (PTK7);

[0466] M is a linker site connected to the antibody or antigen-binding fragment thereof;

[0467] L is a connector connecting M and E;

[0468] E is a structural fragment connecting L and D;

[0469] D is the cytotoxic drug fragment;

[0470] x is selected from 1 to 10.

[0471] In some embodiments, the antibody or antigen-binding fragment thereof comprises:

[0472] (1) The following heavy chain variable region (VH) and / or light chain variable region (VL), wherein the CDRs are defined according to the Chothia numbering system:

[0473] (1a) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 11 or a variant thereof, CDR-H2 of SEQ ID NO: 12 or a variant thereof, and CDR-H3 of SEQ ID NO: 13 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 14 or a variant thereof, CDR-L2 of SEQ ID NO: 15 or a variant thereof, and CDR-L3 of SEQ ID NO: 16 or a variant thereof; or

[0474] (1b) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 27 or a variant thereof, CDR-H2 of SEQ ID NO: 28 or a variant thereof, and CDR-H3 of SEQ ID NO: 29 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 30 or a variant thereof, CDR-L2 of SEQ ID NO: 31 or a variant thereof, and CDR-L3 of SEQ ID NO: 32 or a variant thereof;

[0475] wherein the variant described in any one of (1a) and (1b) has at least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with the sequence from which it is derived, or the variant has one or more amino acid substitutions, deletions or additions (e.g., substitutions, deletions or additions of 1, 2 or 3 amino acids) compared to the sequence from which it is derived; preferably, the substitutions are conservative substitutions;

[0476] or,

[0477] (2) the following heavy chain variable region (VH) and / or light chain variable region (VL), wherein the CDRs are defined according to the Kabat numbering system:

[0478] (2a) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 17 or a variant thereof, CDR-H2 of SEQ ID NO: 18 or 19 or a variant thereof, and CDR-H3 of SEQ ID NO: 13 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 14 or a variant thereof, CDR-L2 of SEQ ID NO: 15 or a variant thereof, and CDR-L3 of SEQ ID NO: 16 or a variant thereof; or

[0479] (2b) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 33 or a variant thereof, CDR-H2 of SEQ ID NO: 34 or 35 or a variant thereof, and CDR-H3 of SEQ ID NO: 29 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 30 or a variant thereof, CDR-L2 of SEQ ID NO: 31 or a variant thereof, and CDR-L3 of SEQ ID NO: 32 or a variant thereof;

[0480] wherein the variant described in any one of (2a) and (2b) has at least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with the sequence from which it is derived, or the variant has one or more amino acid substitutions, deletions or additions (e.g., substitutions, deletions or additions of 1, 2 or 3 amino acids) compared to the sequence from which it is derived; preferably, the substitutions are conservative substitutions;

[0481] or,

[0482] (3) The following heavy chain variable region (VH) and / or light chain variable region (VL), wherein the CDRs are defined according to the IMGT numbering system:

[0483] (3a) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 20 or a variant thereof, CDR-H2 of SEQ ID NO: 21 or a variant thereof, and CDR-H3 of SEQ ID NO: 22 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 23 or a variant thereof, CDR-L2 of SEQ ID NO: 24 or a variant thereof, and CDR-L3 of SEQ ID NO: 16 or a variant thereof; or

[0484] (3b) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 36 or a variant thereof, CDR-H2 of SEQ ID NO: 37 or a variant thereof, and CDR-H3 of SEQ ID NO: 38 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 39 or a variant thereof, CDR-L2 of SEQ ID NO: 40 or a variant thereof, and CDR-L3 of SEQ ID NO: 32 or a variant thereof;

[0485] wherein the variant described in any one of (3a) and (3b) has at least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with the sequence from which it is derived, or the variant has one or more amino acid substitutions, deletions or additions (e.g., substitutions, deletions or additions of 1, 2 or 3 amino acids) compared to the sequence from which it is derived; preferably, the substitutions are conservative substitutions;

[0486] or,

[0487] (4) The following heavy chain variable region (VH) and / or light chain variable region (VL), wherein the CDRs are defined according to the AbM numbering system:

[0488] (4a) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 25 or a variant thereof, CDR-H2 of SEQ ID NO: 26 or a variant thereof, and CDR-H3 of SEQ ID NO: 13 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 14 or a variant thereof, CDR-L2 of SEQ ID NO: 15 or a variant thereof, and CDR-L3 of SEQ ID NO: 16 or a variant thereof; or

[0489] (4b) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 41 or a variant thereof, CDR-H2 of SEQ ID NO: 42 or a variant thereof, and CDR-H3 of SEQ ID NO: 29 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 30 or a variant thereof, CDR-L2 of SEQ ID NO: 31 or a variant thereof, and CDR-L3 of SEQ ID NO: 32 or a variant thereof;

[0490] Wherein, the variant described in any one of (4a) and (4b) has at least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity compared with the sequence from which it is derived, or the variant has one or more amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) compared with the sequence from which it is derived; preferably, the substitutions are conservative substitutions.

[0491] In some embodiments, the antibody or antigen-binding fragment thereof comprises:

[0492] (1a) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 11 or a variant thereof, CDR-H2 of SEQ ID NO: 12 or a variant thereof, and CDR-H3 of SEQ ID NO: 13 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 14 or a variant thereof, CDR-L2 of SEQ ID NO: 15 or a variant thereof, and CDR-L3 of SEQ ID NO: 16 or a variant thereof; or

[0493] (1b) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 27 or a variant thereof, CDR-H2 of SEQ ID NO: 28 or a variant thereof, and CDR-H3 of SEQ ID NO: 29 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 30 or a variant thereof, CDR-L2 of SEQ ID NO: 31 or a variant thereof, and CDR-L3 of SEQ ID NO: 32 or a variant thereof;

[0494] wherein the variant described in any one of (1a) and (1b) has at least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with the sequence from which it is derived, or the variant has one or more amino acid substitutions, deletions or additions (e.g., substitutions, deletions or additions of 1, 2 or 3 amino acids) compared to the sequence from which it is derived; preferably, the substitutions are conservative substitutions;

[0495] or,

[0496] (2a) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 17 or a variant thereof, CDR-H2 of SEQ ID NO: 18 or 19 or a variant thereof, and CDR-H3 of SEQ ID NO: 13 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 14 or a variant thereof, CDR-L2 of SEQ ID NO: 15 or a variant thereof, and CDR-L3 of SEQ ID NO: 16 or a variant thereof; or

[0497] (2b) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 33 or a variant thereof, CDR-H2 of SEQ ID NO: 34 or 35 or a variant thereof, and CDR-H3 of SEQ ID NO: 29 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 30 or a variant thereof, CDR-L2 of SEQ ID NO: 31 or a variant thereof, and CDR-L3 of SEQ ID NO: 32 or a variant thereof;

[0498] wherein the variant described in any one of (2a) and (2b) has at least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with the sequence from which it is derived, or the variant has one or more amino acid substitutions, deletions or additions (e.g., substitutions, deletions or additions of 1, 2 or 3 amino acids) compared to the sequence from which it is derived; preferably, the substitutions are conservative substitutions;

[0499] or,

[0500] (3a) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 20 or a variant thereof, CDR-H2 of SEQ ID NO: 21 or a variant thereof, and CDR-H3 of SEQ ID NO: 22 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 23 or a variant thereof, CDR-L2 of SEQ ID NO: 24 or a variant thereof, and CDR-L3 of SEQ ID NO: 16 or a variant thereof; or

[0501] (3b) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 36 or a variant thereof, CDR-H2 of SEQ ID NO: 37 or a variant thereof, and CDR-H3 of SEQ ID NO: 38 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 39 or a variant thereof, CDR-L2 of SEQ ID NO: 40 or a variant thereof, and CDR-L3 of SEQ ID NO: 32 or a variant thereof;

[0502] wherein the variant described in any one of (3a) and (3b) has at least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with the sequence from which it is derived, or the variant has one or more amino acid substitutions, deletions or additions (e.g., substitutions, deletions or additions of 1, 2 or 3 amino acids) compared to the sequence from which it is derived; preferably, the substitutions are conservative substitutions;

[0503] or,

[0504] (4a) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 25 or a variant thereof, CDR-H2 of SEQ ID NO: 26 or a variant thereof, and CDR-H3 of SEQ ID NO: 13 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 14 or a variant thereof, CDR-L2 of SEQ ID NO: 15 or a variant thereof, and CDR-L3 of SEQ ID NO: 16 or a variant thereof; or

[0505] (4b) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 41 or a variant thereof, CDR-H2 of SEQ ID NO: 42 or a variant thereof, and CDR-H3 of SEQ ID NO: 29 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 30 or a variant thereof, CDR-L2 of SEQ ID NO: 31 or a variant thereof, and CDR-L3 of SEQ ID NO: 32 or a variant thereof;

[0506] Wherein, the variant described in any one of (4a) and (4b) has at least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity compared with the sequence from which it is derived, or the variant has one or more amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) compared with the sequence from which it is derived; preferably, the substitutions are conservative substitutions.

[0507] In some embodiments, the antibody or antigen-binding fragment thereof comprises:

[0508] (1a) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 11, CDR-H2 of SEQ ID NO: 12, and CDR-H3 of SEQ ID NO: 13; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 14, CDR-L2 of SEQ ID NO: 15, and CDR-L3 of SEQ ID NO: 16; or

[0509] (1b) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 27, CDR-H2 of SEQ ID NO: 28, and CDR-H3 of SEQ ID NO: 29; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 30, CDR-L2 of SEQ ID NO: 31, and CDR-L3 of SEQ ID NO: 32;

[0510] or,

[0511] (2a) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 17, CDR-H2 of SEQ ID NO: 18 or 19, and CDR-H3 of SEQ ID NO: 13; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 14, CDR-L2 of SEQ ID NO: 15, and CDR-L3 of SEQ ID NO: 16; or

[0512] (2b) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 33, CDR-H2 of SEQ ID NO: 34 or 35, and CDR-H3 of SEQ ID NO: 29; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 30, CDR-L2 of SEQ ID NO: 31, and CDR-L3 of SEQ ID NO: 32;

[0513] or,

[0514] (3a) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 20, CDR-H2 of SEQ ID NO: 21, and CDR-H3 of SEQ ID NO: 22; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 23, CDR-L2 of SEQ ID NO: 24, and CDR-L3 of SEQ ID NO: 16; or

[0515] (3b) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 36, CDR-H2 of SEQ ID NO: 37, and CDR-H3 of SEQ ID NO: 38; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 39, CDR-L2 of SEQ ID NO: 40, and CDR-L3 of SEQ ID NO: 32;

[0516] or,

[0517] (4a) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 25, CDR-H2 of SEQ ID NO: 26, and CDR-H3 of SEQ ID NO: 13; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 14, CDR-L2 of SEQ ID NO: 15, and CDR-L3 of SEQ ID NO: 16; or

[0518] (4b) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO:41, CDR-H2 of SEQ ID NO:42, and CDR-H3 of SEQ ID NO:29; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO:30, CDR-L2 of SEQ ID NO:31, and CDR-L3 of SEQ ID NO:32.

[0519] In some embodiments, the antibody or antigen-binding fragment thereof comprises: (1a) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with a sequence of SEQ ID NO: 11, CDR-H2 with a sequence of SEQ ID NO: 12, and CDR-H3 with a sequence of SEQ ID NO: 13; and / or, a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with a sequence of SEQ ID NO: 14, CDR-L2 with a sequence of SEQ ID NO: 15, and CDR-L3 with a sequence of SEQ ID NO: 16.

[0520] In some embodiments, the antibody or antigen-binding fragment thereof comprises: (1b) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with a sequence of SEQ ID NO:27, CDR-H2 with a sequence of SEQ ID NO:28, and CDR-H3 with a sequence of SEQ ID NO:29; and / or, a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with a sequence of SEQ ID NO:30, CDR-L2 with a sequence of SEQ ID NO:31, and CDR-L3 with a sequence of SEQ ID NO:32.

[0521] In some embodiments, the antibody or antigen-binding fragment thereof comprises: (2a) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with a sequence of SEQ ID NO: 17, CDR-H2 with a sequence of SEQ ID NO: 18 or 19, and CDR-H3 with a sequence of SEQ ID NO: 13; and / or, a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with a sequence of SEQ ID NO: 14, CDR-L2 with a sequence of SEQ ID NO: 15, and CDR-L3 with a sequence of SEQ ID NO: 16.

[0522] In some embodiments, the antibody or antigen-binding fragment thereof comprises: (2b) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with a sequence of SEQ ID NO:33, CDR-H2 with a sequence of SEQ ID NO:34 or 35, and CDR-H3 with a sequence of SEQ ID NO:29; and / or, a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with a sequence of SEQ ID NO:30, CDR-L2 with a sequence of SEQ ID NO:31, and CDR-L3 with a sequence of SEQ ID NO:32.

[0523] In some embodiments, the antibody or antigen-binding fragment thereof comprises: (3a) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with a sequence of SEQ ID NO:20, CDR-H2 with a sequence of SEQ ID NO:21, and CDR-H3 with a sequence of SEQ ID NO:22; and / or, a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with a sequence of SEQ ID NO:23, CDR-L2 with a sequence of SEQ ID NO:24, and CDR-L3 with a sequence of SEQ ID NO:16.

[0524] In some embodiments, the antibody or antigen-binding fragment thereof comprises: (3b) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with a sequence of SEQ ID NO:36, CDR-H2 with a sequence of SEQ ID NO:37, and CDR-H3 with a sequence of SEQ ID NO:38; and / or, a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with a sequence of SEQ ID NO:39, CDR-L2 with a sequence of SEQ ID NO:40, and CDR-L3 with a sequence of SEQ ID NO:32.

[0525] In some embodiments, the antibody or antigen-binding fragment thereof comprises: (4a) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with a sequence of SEQ ID NO:25, CDR-H2 with a sequence of SEQ ID NO:26, and CDR-H3 with a sequence of SEQ ID NO:13; and / or, a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with a sequence of SEQ ID NO:14, CDR-L2 with a sequence of SEQ ID NO:15, and CDR-L3 with a sequence of SEQ ID NO:16.

[0526] In some embodiments, the antibody or antigen-binding fragment thereof comprises: (4b) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with a sequence of SEQ ID NO:41, CDR-H2 with a sequence of SEQ ID NO:42, and CDR-H3 with a sequence of SEQ ID NO:29; and / or, a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with a sequence of SEQ ID NO:30, CDR-L2 with a sequence of SEQ ID NO:31, and CDR-L3 with a sequence of SEQ ID NO:32.

[0527] In some embodiments, the antibody or antigen-binding fragment thereof comprises:

[0528] (a) VH or a variant thereof shown in SEQ ID NO: 1, and / or VL or a variant thereof shown in SEQ ID NO: 2;

[0529] (b) VH or a variant thereof shown in SEQ ID NO: 3, and / or VL or a variant thereof shown in SEQ ID NO: 4; or

[0530] (c) VH or a variant thereof shown in SEQ ID NO: 5, and / or VL or a variant thereof shown in SEQ ID NO: 6;

[0531] (d) VH or a variant thereof shown in SEQ ID NO: 7, and / or VL or a variant thereof shown in SEQ ID NO: 8;

[0532] (e) VH or a variant thereof shown in SEQ ID NO: 9, and / or VL or a variant thereof shown in SEQ ID NO: 10;

[0533] wherein the variant has at least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity compared to the sequence from which it is derived, or the variant has one or more amino acid substitutions, deletions or additions (e.g., 1, 2, 3, 4 or 5 amino acid substitutions, deletions or additions) compared to the sequence from which it is derived; preferably, the substitutions are conservative substitutions.

[0534] In some embodiments, the antibody or antigen-binding fragment thereof comprises:

[0535] (a) VH shown in SEQ ID NO: 1, and / or VL shown in SEQ ID NO: 2;

[0536] (b) VH shown in SEQ ID NO: 3, and / or VL shown in SEQ ID NO: 4;

[0537] (c) VH represented by SEQ ID NO: 5, and / or VL represented by SEQ ID NO: 6;

[0538] (d) VH shown in SEQ ID NO: 7, and / or VL shown in SEQ ID NO: 8; or

[0539] (e) VH represented by SEQ ID NO: 9, and / or VL represented by SEQ ID NO: 10.

[0540] In some embodiments, the antibody or antigen-binding fragment thereof comprises:

[0541] (a) VH shown in SEQ ID NO: 1, and / or VL shown in SEQ ID NO: 2.

[0542] In some embodiments, the antibody or antigen-binding fragment thereof comprises:

[0543] (b) VH represented by SEQ ID NO: 3, and / or VL represented by SEQ ID NO: 4.

[0544] In some embodiments, the antibody or antigen-binding fragment thereof comprises:

[0545] (c) VH represented by SEQ ID NO: 5, and / or VL represented by SEQ ID NO: 6.

[0546] In some embodiments, the antibody or antigen-binding fragment thereof comprises:

[0547] (d) VH represented by SEQ ID NO: 7, and / or VL represented by SEQ ID NO: 8.

[0548] In some embodiments, the antibody or antigen-binding fragment thereof comprises:

[0549] (e) VH represented by SEQ ID NO: 9, and / or VL represented by SEQ ID NO: 10.

[0550] In some embodiments, the antibody or antigen-binding fragment thereof comprises: a heavy chain variable region (VH) comprising CDR-H1, CDR-H2 and CDR-H3, wherein the CDR-H1, CDR-H2 and CDR-H3 respectively comprise the amino acid sequences of CDR-H1, CDR-H2 and CDR-H3 shown in VH having the amino acid sequence of SEQ ID NO: 1; and a light chain variable region (VL) comprising CDR-L1, CDR-L2 and CDR-L3, wherein the CDR-L1, CDR-L2 and CDR-L3 respectively comprise the amino acid sequences of CDR-L1, CDR-L2 and CDR-L3 shown in VL having the amino acid sequence of SEQ ID NO: 2.

[0551] In some embodiments, the antibody or antigen-binding fragment thereof comprises: a heavy chain variable region (VH) comprising CDR-H1, CDR-H2 and CDR-H3, wherein the CDR-H1, CDR-H2 and CDR-H3 respectively comprise the amino acid sequences of CDR-H1, CDR-H2 and CDR-H3 shown in VH having the amino acid sequence of SEQ ID NO: 3; and a light chain variable region (VL) comprising CDR-L1, CDR-L2 and CDR-L3, wherein the CDR-L1, CDR-L2 and CDR-L3 respectively comprise the amino acid sequences of CDR-L1, CDR-L2 and CDR-L3 shown in VL having the amino acid sequence of SEQ ID NO: 4.

[0552] In some embodiments, the antibody or antigen-binding fragment thereof comprises: a heavy chain variable region (VH) comprising CDR-H1, CDR-H2 and CDR-H3, wherein the CDR-H1, CDR-H2 and CDR-H3 respectively comprise the amino acid sequences of CDR-H1, CDR-H2 and CDR-H3 shown in VH having the amino acid sequence of SEQ ID NO:5; and a light chain variable region (VL) comprising CDR-L1, CDR-L2 and CDR-L3, wherein the CDR-L1, CDR-L2 and CDR-L3 respectively comprise the amino acid sequences of CDR-L1, CDR-L2 and CDR-L3 shown in VL having the amino acid sequence of SEQ ID NO:6.

[0553] In some embodiments, the antibody or antigen-binding fragment thereof comprises: a heavy chain variable region (VH) comprising CDR-H1, CDR-H2 and CDR-H3, wherein the CDR-H1, CDR-H2 and CDR-H3 respectively comprise the amino acid sequences of CDR-H1, CDR-H2 and CDR-H3 shown in VH having the amino acid sequence of SEQ ID NO:7; and a light chain variable region (VL) comprising CDR-L1, CDR-L2 and CDR-L3, wherein the CDR-L1, CDR-L2 and CDR-L3 respectively comprise the amino acid sequences of CDR-L1, CDR-L2 and CDR-L3 shown in VL having the amino acid sequence of SEQ ID NO:8.

[0554] In some embodiments, the antibody or antigen-binding fragment thereof comprises: a heavy chain variable region (VH) comprising CDR-H1, CDR-H2 and CDR-H3, wherein the CDR-H1, CDR-H2 and CDR-H3 respectively comprise the amino acid sequences of CDR-H1, CDR-H2 and CDR-H3 shown in VH having the amino acid sequence of SEQ ID NO:9; and a light chain variable region (VL) comprising CDR-L1, CDR-L2 and CDR-L3, wherein the CDR-L1, CDR-L2 and CDR-L3 respectively comprise the amino acid sequences of CDR-L1, CDR-L2 and CDR-L3 shown in VL having the amino acid sequence of SEQ ID NO:10.

[0555] In some embodiments, the antibody or antigen-binding fragment thereof further comprises:

[0556] (a) a heavy chain constant region (CH) of a human immunoglobulin, or a variant thereof, which has one or more amino acid substitutions, deletions or additions (e.g., up to 20, up to 15, up to 10, or up to 5 amino acid substitutions, deletions or additions; e.g., 1, 2, 3, 4 or 5 amino acid substitutions, deletions or additions) compared to the wild-type sequence from which it is derived; and

[0557] (b) a light chain constant region (CL) of a human immunoglobulin, or a variant thereof, which has one or more amino acid substitutions, deletions or additions (e.g., up to 20, up to 15, up to 10, or up to 5 amino acid substitutions, deletions or additions; for example, 1, 2, 3, 4 or 5 amino acid substitutions, deletions or additions) compared to the wild-type sequence from which it is derived.

[0558] In some embodiments, the heavy chain constant region is an IgG heavy chain constant region, such as an IgG1, IgG2, IgG3 or IgG4 heavy chain constant region, such as a human IgG1 heavy chain constant region or a human IgG4 heavy chain constant region. In some embodiments, the antibody or antigen-binding fragment thereof comprises a heavy chain constant region (CH) as set forth in SEQ ID NO: 43 or a variant thereof, wherein the variant has up to 20 conservative substitutions of amino acids compared to SEQ ID NO: 43 (e.g., up to 15, up to 10, or up to 5 conservative substitutions of amino acids; e.g., 1, 2, 3, 4 or 5 conservative substitutions of amino acids).

[0559] In some embodiments, the light chain constant region is a kappa light chain constant region. In some embodiments, the antibody or antigen-binding fragment thereof comprises a light chain constant region (CL) as set forth in SEQ ID NO: 44, or a variant thereof, wherein the variant has up to 20 conservative amino acid substitutions (e.g., up to 15, up to 10, or up to 5 conservative amino acid substitutions; e.g., 1, 2, 3, 4, or 5 conservative amino acid substitutions) compared to SEQ ID NO: 44.

[0560] In some embodiments, the antibody or antigen-binding fragment thereof comprises a heavy chain constant region (CH) as shown in SEQ ID NO: 43 or 45 and a light chain constant region (CL) as shown in SEQ ID NO: 44.

[0561] In some embodiments, the antibody or antigen-binding fragment thereof comprises:

[0562] (1) a heavy chain comprising the VH sequence of SEQ ID NO: 1 and the heavy chain constant region (CH) of SEQ ID NO: 43, and a light chain comprising the VL sequence of SEQ ID NO: 2 and the light chain constant region (CL) of SEQ ID NO: 44;

[0563] (2) a heavy chain comprising a VH sequence of SEQ ID NO: 3 and a heavy chain constant region (CH) of SEQ ID NO: 43 or 45, and a light chain comprising a VL sequence of SEQ ID NO: 4 and a light chain constant region (CL) of SEQ ID NO: 44;

[0564] (3) a heavy chain comprising the VH sequence of SEQ ID NO: 5 and the heavy chain constant region (CH) of SEQ ID NO: 43, and a light chain comprising the VL sequence of SEQ ID NO: 6 and the light chain constant region (CL) of SEQ ID NO: 44;

[0565] (4) a heavy chain comprising the VH sequence of SEQ ID NO: 7 and the heavy chain constant region (CH) of SEQ ID NO: 43, and a light chain comprising the VL sequence of SEQ ID NO: 8 and the light chain constant region (CL) of SEQ ID NO: 44; or

[0566] (5) A heavy chain comprising the VH sequence shown in SEQ ID NO: 9 and the heavy chain constant region (CH) shown in SEQ ID NO: 43, and a light chain comprising the VL sequence shown in SEQ ID NO: 10 and the light chain constant region (CL) shown in SEQ ID NO: 44.

[0567] In some embodiments, the antibody or antigen-binding fragment thereof comprises:

[0568] (1) A heavy chain comprising the VH sequence of SEQ ID NO: 1 and the heavy chain constant region (CH) of SEQ ID NO: 43, and a light chain comprising the VL sequence of SEQ ID NO: 2 and the light chain constant region (CL) of SEQ ID NO: 44.

[0569] In some embodiments, the antibody or antigen-binding fragment thereof comprises:

[0570] (2) A heavy chain comprising the VH sequence shown in SEQ ID NO: 3 and the heavy chain constant region (CH) shown in SEQ ID NO: 43 or 45, and a light chain comprising the VL sequence shown in SEQ ID NO: 4 and the light chain constant region (CL) shown in SEQ ID NO: 44.

[0571] In some embodiments, the antibody or antigen-binding fragment thereof comprises:

[0572] (3) A heavy chain comprising the VH sequence shown in SEQ ID NO: 5 and the heavy chain constant region (CH) shown in SEQ ID NO: 43, and a light chain comprising the VL sequence shown in SEQ ID NO: 6 and the light chain constant region (CL) shown in SEQ ID NO: 44.

[0573] In some embodiments, the antibody or antigen-binding fragment thereof comprises:

[0574] (4) A heavy chain comprising the VH sequence of SEQ ID NO: 7 and the heavy chain constant region (CH) of SEQ ID NO: 43, and a light chain comprising the VL sequence of SEQ ID NO: 8 and the light chain constant region (CL) of SEQ ID NO: 44.

[0575] In some embodiments, the antibody or antigen-binding fragment thereof comprises:

[0576] (5) A heavy chain comprising the VH sequence shown in SEQ ID NO: 9 and the heavy chain constant region (CH) shown in SEQ ID NO: 43, and a light chain comprising the VL sequence shown in SEQ ID NO: 10 and the light chain constant region (CL) shown in SEQ ID NO: 44.

[0577] In some embodiments, the antibody or antigen-binding fragment thereof comprises a heavy chain (HC) of SEQ ID NO: 46 and a light chain (LC) of SEQ ID NO: 47.

[0578] In certain embodiments, the antibody or antigen-binding fragment thereof comprises a heavy chain (HC) of SEQ ID NO: 52 and a light chain (LC) of SEQ ID NO: 47.

[0579] In certain embodiments, the antibody or antigen-binding fragment thereof comprises a heavy chain (HC) of SEQ ID NO: 53 and a light chain (LC) of SEQ ID NO: 47.

[0580] In certain embodiments, the antibody or antigen-binding fragment thereof comprises a heavy chain (HC) of SEQ ID NO: 54 and a light chain (LC) of SEQ ID NO: 47.

[0581] In certain embodiments of the antibodies or antibody drug conjugates described herein, the heavy chain constant region may comprise a C-terminal lysine, or lack a C-terminal lysine or a C-terminal glycine-lysine dipeptide. In certain embodiments of the antibodies or antibody drug conjugates described herein, the N-terminal amino acid of the antibody variable region may be cyclized to pyroglutamate.

[0582] In certain embodiments of the antibodies or antibody drug conjugates described herein, the N-terminal amino acid of the antibody variable region can be cyclized to pyroglutamic acid.

[0583] In certain embodiments, the antibodies or antigen-binding fragments disclosed herein include antibodies or antigen-binding fragments that specifically bind to an antigen and may include post-translational modifications thereof (e.g., C-terminal lysine trimming in the heavy chain, conversion of glutamine or glutamic acid to pyroglutamate or pyroglutamic acid), which may occur upon recombinant expression in a host cell (e.g., CHO cells) or during purification / storage.

[0584] Thus, the composition may comprise various antibody drug conjugates, wherein each species may independently comprise a C-terminal lysine, lack a C-terminal lysine, lack a C-terminal glycine-lysine, and / or comprise an N-terminal glutamine or glutamic acid, or pyroglutamate resulting from cyclization of the N-terminal amino acid.

[0585] In certain embodiments, the composition may comprise various antibody drug conjugates, wherein each species may independently comprise a C-terminal lysine, lack a C-terminal lysine, lack a C-terminal glycine-lysine, and / or comprise an N-terminal glutamine or glutamic acid, or pyroglutamic acid resulting from cyclization of the N-terminal amino acid.

[0586] Therefore, in certain embodiments, the present invention further provides a composition comprising an antibody drug conjugate as described herein, wherein the major antibody drug conjugate species in the composition include: (i) an antibody with a lysine residue deleted at the C-terminus of the heavy chain; (ii) an antibody with glutamine, glutamic acid, pyroglutamate or pyroglutamate at the N-terminus of the heavy chain; or (iii) an antibody with a lysine residue deleted at the C-terminus of the heavy chain and glutamine, glutamic acid, pyroglutamate or pyroglutamate at the N-terminus of the heavy chain.

[0587] Therefore, in certain embodiments, the present invention further provides a composition comprising an antibody drug conjugate as described herein, wherein the majority of the antibody drug conjugate species in the composition comprises an antibody having a lysine residue deleted from the C-terminus of the heavy chain and pyroglutamate or pyroglutamic acid at the N-terminus of the heavy chain.

[0588] In certain embodiments, the antibody or antigen-binding fragment thereof according to the present invention comprises a heavy chain comprising a VH having an amino acid sequence selected from the group consisting of SEQ ID NO: 1, 3, 5, 7 and 9.

[0589] In certain embodiments, the antibody or antigen-binding fragment thereof according to the present invention comprises a heavy chain comprising a VH having an amino acid sequence selected from SEQ ID NO: 1.

[0590] In certain embodiments, the antibody or antigen-binding fragment thereof according to the present invention comprises a heavy chain comprising a VH having an amino acid sequence selected from SEQ ID NO:3.

[0591] In certain embodiments, the antibody or antigen-binding fragment thereof according to the present invention comprises a heavy chain comprising a VH having an amino acid sequence selected from SEQ ID NO:5.

[0592] In certain embodiments, the antibody or antigen-binding fragment thereof according to the present invention comprises a heavy chain comprising a VH having an amino acid sequence selected from SEQ ID NO:7.

[0593] In certain embodiments, the antibody or antigen-binding fragment thereof according to the present invention comprises a heavy chain comprising a VH having an amino acid sequence selected from SEQ ID NO:9.

[0594] In certain embodiments, the antibody or antigen-binding fragment thereof according to the present invention comprises a heavy chain comprising a VH having an amino acid sequence selected from SEQ ID NO: 1, 3, 5, 7 and 9, wherein the N-terminal glutamine or glutamic acid of the antibody heavy chain variable region is cyclized to pyroglutamate.

[0595] In certain embodiments, the antibody or antigen-binding fragment thereof according to the present invention comprises a heavy chain comprising a VH having an amino acid sequence selected from SEQ ID NO: 1, 3, 5, 7 and 9, wherein the N-terminal glutamine or glutamic acid in the antibody heavy chain variable region is cyclized to pyroglutamic acid.

[0596] In certain embodiments, the antibody or antigen-binding fragment thereof according to the present invention comprises a heavy chain comprising a VH having an amino acid sequence selected from SEQ ID NO: 1, wherein the N-terminal glutamine or glutamic acid of the antibody heavy chain variable region is cyclized to pyroglutamate.

[0597] In certain embodiments, the antibody or antigen-binding fragment thereof according to the present invention comprises a heavy chain comprising a VH having an amino acid sequence selected from SEQ ID NO: 1, wherein the N-terminal glutamine or glutamic acid in the antibody heavy chain variable region is cyclized to pyroglutamic acid.

[0598] In certain embodiments, the antibody or antigen-binding fragment thereof according to the present invention comprises a heavy chain comprising a VH having an amino acid sequence selected from SEQ ID NO: 3, wherein the N-terminal glutamine or glutamic acid of the antibody heavy chain variable region is cyclized to pyroglutamate.

[0599] In certain embodiments, the antibody or antigen-binding fragment thereof according to the present invention comprises a heavy chain comprising a VH having an amino acid sequence selected from SEQ ID NO: 3, wherein the N-terminal glutamine or glutamic acid of the antibody heavy chain variable region is cyclized to pyroglutamic acid.

[0600] In certain embodiments, the antibody or antigen-binding fragment thereof according to the present invention comprises a heavy chain comprising a VH having an amino acid sequence selected from SEQ ID NO: 5, wherein the N-terminal glutamine or glutamic acid of the antibody heavy chain variable region has been cyclized to pyroglutamate.

[0601] In certain embodiments, the antibody or antigen-binding fragment thereof according to the present invention comprises a heavy chain comprising a VH having an amino acid sequence selected from SEQ ID NO: 5, wherein the N-terminal glutamine or glutamic acid of the antibody heavy chain variable region has been cyclized to pyroglutamic acid.

[0602] In certain embodiments, the antibody or antigen-binding fragment thereof according to the present invention comprises a heavy chain comprising a VH having an amino acid sequence selected from SEQ ID NO: 7, wherein the N-terminal glutamine or glutamic acid of the antibody heavy chain variable region has been cyclized to pyroglutamate.

[0603] In certain embodiments, the antibody or antigen-binding fragment thereof according to the present invention comprises a heavy chain comprising a VH having an amino acid sequence selected from SEQ ID NO: 7, wherein the N-terminal glutamine or glutamic acid of the antibody heavy chain variable region has been cyclized to pyroglutamic acid.

[0604] In certain embodiments, the antibody or antigen-binding fragment thereof according to the present invention comprises a heavy chain comprising a VH having an amino acid sequence selected from SEQ ID NO: 9, wherein the N-terminal glutamine or glutamic acid of the antibody heavy chain variable region has been cyclized to pyroglutamate.

[0605] In certain embodiments, the antibody or antigen-binding fragment thereof according to the present invention comprises a heavy chain comprising a VH having an amino acid sequence selected from SEQ ID NO: 9, wherein the N-terminal glutamine or glutamic acid of the antibody heavy chain variable region has been cyclized to pyroglutamic acid.

[0606] In certain embodiments, the antibody or antigen-binding fragment thereof according to the present invention comprises a heavy chain comprising a VL having an amino acid sequence selected from SEQ ID NO: 8, wherein the N-terminal glutamine or glutamic acid of the antibody light chain variable region has been cyclized to pyroglutamate.

[0607] In certain embodiments, the antibody or antigen-binding fragment thereof according to the present invention comprises a heavy chain comprising a VL having an amino acid sequence selected from SEQ ID NO: 8, wherein the N-terminal glutamine or glutamic acid of the antibody light chain variable region has been cyclized to pyroglutamic acid.

[0608] In certain embodiments, the antibody or antigen-binding fragment thereof according to the present invention comprises: a heavy chain comprising a VH having the amino acid sequence shown in SEQ ID NO: 7 and / or a light chain comprising a VL having the amino acid sequence shown in SEQ ID NO: 8, and wherein the N-terminal glutamine of SEQ ID NO: 7 and / or the N-terminal glutamic acid of SEQ ID NO: 8 has been cyclized to pyroglutamate.

[0609] In certain embodiments, the antibody or antigen-binding fragment thereof according to the present invention comprises: a heavy chain comprising a VH having the amino acid sequence shown in SEQ ID NO: 7 and / or a light chain comprising a VL having the amino acid sequence shown in SEQ ID NO: 8, and wherein the N-terminal glutamine of SEQ ID NO: 7 and / or the N-terminal glutamic acid of SEQ ID NO: 8 has been cyclized to pyroglutamic acid.

[0610] In certain embodiments, the antibody or antigen-binding fragment thereof comprises:

[0611] (a) the heavy chain (HC) of SEQ ID NO: 52 and the light chain (LC) of SEQ ID NO: 47;

[0612] (b) a heavy chain (HC) of SEQ ID NO: 53 and a light chain (LC) of SEQ ID NO: 47; or

[0613] (c) The heavy chain (HC) of SEQ ID NO: 54 and the light chain (LC) of SEQ ID NO: 47.

[0614] In certain embodiments, the antibodies or antigen-binding fragments thereof of the present invention can be obtained by expressing nucleic acid molecules encoding the heavy and light chains of the antibodies or antigen-binding fragments thereof in host cells, wherein the antibodies or antigen-binding fragments thereof comprise:

[0615] (a) a VH having the amino acid sequence shown in SEQ ID NO: 1 or a variant thereof and / or a VL having the amino acid sequence shown in SEQ ID NO: 2 or a variant thereof;

[0616] (b) a VH having the amino acid sequence shown in SEQ ID NO: 3 or a variant thereof and / or a VL having the amino acid sequence shown in SEQ ID NO: 4 or a variant thereof;

[0617] (c) a VH having the amino acid sequence shown in SEQ ID NO: 5 or a variant thereof and / or a VL having the amino acid sequence shown in SEQ ID NO: 6 or a variant thereof;

[0618] (d) a VH having the amino acid sequence shown in SEQ ID NO: 7 or a variant thereof and / or a VL having the amino acid sequence shown in SEQ ID NO: 8 or a variant thereof; or

[0619] (e) a VH having the amino acid sequence shown in SEQ ID NO: 9 or a variant thereof and / or a VL having the amino acid sequence shown in SEQ ID NO: 10 or a variant thereof;

[0620] wherein the variant has at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% sequence identity compared to the sequence from which it is derived, or has one or more amino acid substitutions, deletions or additions (e.g., substitutions, deletions or additions of 1, 2, 3, 4 or 5 amino acids) compared to the sequence from which it is derived.

[0621] In certain embodiments, the substitution is a conservative substitution. In certain embodiments, the antibody or antigen-binding fragment thereof of the present invention is obtained by expressing a nucleic acid molecule encoding the heavy and light chains of the antibody or antigen-binding fragment thereof in a host cell, wherein the antibody or antigen-binding fragment thereof comprises:

[0622] (a) a VH having the amino acid sequence shown in SEQ ID NO: 1 and / or a VL having the amino acid sequence shown in SEQ ID NO: 2;

[0623] (b) a VH having the amino acid sequence shown in SEQ ID NO: 3 and / or a VL having the amino acid sequence shown in SEQ ID NO: 4;

[0624] (c) a VH having the amino acid sequence shown in SEQ ID NO: 5 and / or a VL having the amino acid sequence shown in SEQ ID NO: 6;

[0625] (d) a VH having the amino acid sequence shown in SEQ ID NO: 7 and / or a VL having the amino acid sequence shown in SEQ ID NO: 8; or

[0626] (e) VH having the amino acid sequence shown in SEQ ID NO: 9 and / or VL having the amino acid sequence shown in SEQ ID NO: 10.

[0627] In certain embodiments, the antibodies or antigen-binding fragments thereof of the present invention are obtained by expressing a nucleic acid molecule encoding the heavy and light chains of the antibodies or antigen-binding fragments thereof in a host cell, wherein the antibodies or antigen-binding fragments thereof comprise:

[0628] (1) a heavy chain comprising the VH sequence shown in SEQ ID NO: 1 and the heavy chain constant region (CH) shown in SEQ ID NO: 43, and a light chain comprising the VL sequence shown in SEQ ID NO: 2 and the light chain constant region (CL) shown in SEQ ID NO: 44;

[0629] (2) a heavy chain comprising the VH sequence shown in SEQ ID NO: 3 and the heavy chain constant region (CH) shown in SEQ ID NO: 43 or 45, and a light chain comprising the VL sequence shown in SEQ ID NO: 4 and the light chain constant region (CL) shown in SEQ ID NO: 44;

[0630] (3) a heavy chain comprising the VH sequence shown in SEQ ID NO: 5 and the heavy chain constant region (CH) shown in SEQ ID NO: 43, and a light chain comprising the VL sequence shown in SEQ ID NO: 6 and the light chain constant region (CL) shown in SEQ ID NO: 44;

[0631] (4) a heavy chain comprising the VH sequence shown in SEQ ID NO: 7 and the heavy chain constant region (CH) shown in SEQ ID NO: 43, and a light chain comprising the VL sequence shown in SEQ ID NO: 8 and the light chain constant region (CL) shown in SEQ ID NO: 44;

[0632] (5) a heavy chain comprising the VH sequence shown in SEQ ID NO: 9 and the heavy chain constant region (CH) shown in SEQ ID NO: 43, and a light chain comprising the VL sequence shown in SEQ ID NO: 10 and the light chain constant region (CL) shown in SEQ ID NO: 44;

[0633] (6) a heavy chain (HC) having the amino acid sequence shown in SEQ ID NO: 46 and a light chain (LC) having the amino acid sequence shown in SEQ ID NO: 47;

[0634] (7) VH shown in SEQ ID NO: 1 and VL shown in SEQ ID NO: 2;

[0635] (8) VH shown in SEQ ID NO: 3 and VL shown in SEQ ID NO: 4;

[0636] (9) VH shown in SEQ ID NO: 5 and VL shown in SEQ ID NO: 6;

[0637] (10) VH shown in SEQ ID NO: 7 and VL shown in SEQ ID NO: 8; or

[0638] (11) VH shown in SEQ ID NO: 9 and VL shown in SEQ ID NO: 10.

[0639] In certain embodiments, the antibodies or antigen-binding fragments thereof of the present invention are obtained by expressing nucleic acid molecules encoding the heavy and light chains of the antibodies or antigen-binding fragments thereof in host cells, wherein the antibodies or antigen-binding fragments thereof comprise: (a) VH or a variant thereof shown in SEQ ID NO: 1, and / or VL or a variant thereof shown in SEQ ID NO: 2.

[0640] In certain embodiments, the antibodies or antigen-binding fragments thereof of the present invention are obtained by expressing nucleic acid molecules encoding the heavy and light chains of the antibodies or antigen-binding fragments thereof in host cells, wherein the antibodies or antigen-binding fragments thereof comprise: (b) VH or a variant thereof shown in SEQ ID NO: 3, and / or VL or a variant thereof shown in SEQ ID NO: 4.

[0641] In certain embodiments, the antibodies or antigen-binding fragments thereof of the present invention are obtained by expressing nucleic acid molecules encoding the heavy and light chains of the antibodies or antigen-binding fragments thereof in host cells, wherein the antibodies or antigen-binding fragments thereof comprise: (c) VH or a variant thereof shown in SEQ ID NO: 5, and / or VL or a variant thereof shown in SEQ ID NO: 6.

[0642] In certain embodiments, the antibodies or antigen-binding fragments thereof of the present invention are obtained by expressing nucleic acid molecules encoding the heavy and light chains of the antibodies or antigen-binding fragments thereof in host cells, wherein the antibodies or antigen-binding fragments thereof comprise: (d) VH or a variant thereof shown in SEQ ID NO: 7, and / or VL or a variant thereof shown in SEQ ID NO: 8.

[0643] In certain embodiments, the antibodies or antigen-binding fragments thereof of the present invention are obtained by expressing nucleic acid molecules encoding the heavy and light chains of the antibodies or antigen-binding fragments thereof in host cells, wherein the antibodies or antigen-binding fragments thereof comprise: (e) VH or a variant thereof shown in SEQ ID NO: 9, and / or VL or a variant thereof shown in SEQ ID NO: 10.

[0644] In certain embodiments of antibodies, the heavy chain constant region disclosed herein may comprise a C-terminal lysine, or lack a C-terminal lysine or a C-terminal glycine-lysine dipeptide. In some embodiments of antibodies or antigen-binding fragments thereof, the N-terminal amino acid of the antibody or antigen-binding fragment thereof may be cyclized to a pyroglutamate salt.

[0645] In some embodiments of the antibody or antigen-binding fragment, the N-terminal amino acid of the antibody or antigen-binding fragment thereof can be cyclized to pyroglutamate.

[0646] Thus, in compositions comprising a particular antibody described herein, the composition can comprise various antibody drug conjugates, wherein each species can independently comprise a C-terminal lysine, lack a C-terminal lysine, lack a C-terminal glycine-lysine, and / or comprise an N-terminal glutamine or glutamic acid, or pyroglutamate resulting from cyclization of the N-terminal amino acid.

[0647] In some embodiments, in compositions comprising specific antibodies described herein, the compositions may comprise various antibody drug conjugates, wherein each species may independently comprise a C-terminal lysine, lack a C-terminal lysine, lack a C-terminal glycine-lysine, and / or comprise an N-terminal glutamine or glutamic acid, or pyroglutamic acid resulting from cyclization of the N-terminal amino acid.

[0648] In the antibody-drug conjugate, the cytotoxic drug can be linked to the antibody or antigen-binding fragment thereof via a linker (such as the "MLE" fragment shown in this application).

[0649] In some embodiments, M is Wherein ring A is a 5-6 membered alicyclic heterocyclic ring, or a 5-20 membered aromatic ring system, wherein the alicyclic heterocyclic ring and the aromatic ring system are optionally substituted by one or more selected from oxy (=O), halogen, cyano, amino, carboxyl, thiol and C 1-6 Alkyl group substituted; M1 is selected from single bond and C 1-20 Alkylene, C 2-20 Alkenylene or C 2-20 Alkynylidene.

[0650] In some embodiments, M is wherein Ring A is a 5-membered alicyclic heterocycle, a 6-membered heteroaromatic ring, or a polycyclic ring formed by connecting one or more 6-membered heteroaromatic rings to a benzene ring via a single bond, wherein the alicyclic heterocycle is optionally substituted with one or more selected from oxy (=O), halogen and C 1-4 Alkyl group substitution; M1 is selected from single bond, C 3-10 Alkylene, C 3-10 Alkenylene or C 3-10 Alkynylidene.

[0651] In some embodiments, M is wherein ring A is selected from M1 is selected from a single bond and C 5-8 Alkylene, C 5-8 Alkenylene or C 5-8 Alkynylidene.

[0652] In some embodiments, M is selected from the following structures:

[0653]

[0654] In some embodiments, M is selected from the following structures:

[0655]

[0656] In some embodiments, L is selected from the group consisting of one or more of the following: C 1-6 Alkylene, -N(R')-, Carbonyl, -O-, Val, Cit, Phe, Lys, Lys(COCH2CH2(OCH2CH2) s OCH3), D-Val, Leu, Gly, Ala, Asn, Val-Cit, Val-Ala, Val-Lys, Val-Lys(Ac), Phe-Lys, Phe-Lys(Ac), D-Val-Leu- Lys, Gly-Gly-Arg, Ala-Ala-Asn, Ala-Ala-Ala, Val-Lys-Ala, Val-Lys-Gly, Gly-Gly-Gly, Gly-Gly-Phe-Gly (SEQ ID NO:48), Gly-Phe-Leu-Gly (SEQ ID NO:49), Gly-Gly-Val-Ala (SEQ ID NO:50), Gly-Gly-Gly-Gly-Gly (SEQ ID NO:51), Where R' represents hydrogen, C 1-6 Alkyl or alkyl containing -(CH2CH2O)r-; r is selected from an integer of 1-10; s is selected from an integer of 1-20.

[0657] In some embodiments, L is selected from the structure consisting of one or more of the following: 1-6 Alkylene, -NH-, Val, Cit, Phe, Lys, Lys(COCH2CH2(OCH2CH2) s OCH3), Gly, Val-Cit, Gly-Gly-Phe-Gly, wherein s is selected from an integer of 1-20.

[0658] In some embodiments, L is selected from the following structures:

[0659] wherein s is selected from an integer of 1-20.

[0660] In some embodiments, L is selected from the following structures:

[0661]

[0662] In some embodiments, L is selected from the following structures:

[0663]

[0664] In some embodiments, L is selected from the following structures:

[0665]

[0666] In some embodiments, E is a single bond, -NH-CH2-, -NH-CH2-O-CH2-CO-,

[0667] In some embodiments, E is a single bond, -NH-CH2-, -NH-CH2-O-CH2-CO-,

[0668] In some embodiments, E is -NH-CH2-,

[0669] In some embodiments, E is -NH-CH2- or

[0670] In some embodiments, M is selected from the following structures:

[0671]

[0672] L is selected from the following structures:

[0673]

[0674] E is -NH-CH2,

[0675] In some embodiments, Selected from the following structures:

[0676]

[0677]

[0678]

[0679]

[0680] In some embodiments, Selected from the following structures:

[0681]

[0682]

[0683] In some embodiments, Selected from the following structures:

[0684]

[0685]

[0686] In some embodiments, the cytotoxic drug is selected from microtubule inhibitors, DNA intercalators, DNA topoisomerase inhibitors and RNA polymerase inhibitors. In some embodiments, the microtubule inhibitor is an auristatin compound or a maytansine compound. In some embodiments, the DNA intercalator is a pyrrolobenzodiazepine (PBD). In some embodiments, the DNA topoisomerase inhibitor is a topoisomerase I inhibitor (e.g., camptothecin, hydroxycamptothecin, 9-aminocamptothecin, SN-38, irinotecan, topotecan, belotecan, or rubitecan) or a topoisomerase II inhibitor (e.g., doxorubicin, PNU-159682, multicarmycin, daunorubicin, mitoxantrone, podophyllotoxin, or etoposide). In some embodiments, the RNA polymerase inhibitor is α-amanitin or a pharmaceutically acceptable salt, ester, or analog thereof.

[0687] The cytotoxic drugs disclosed in this application generally contain a variety of functional groups, such as hydroxyl (-OH), carboxyl (-COOH), sulfhydryl (-SH), primary amine (-NH2), secondary amine (-NR A H) or tertiary amine (-NRB R C ), where R A 、R B 、R C These represent only non-hydrogen substituents on N, through which the cytotoxic drug can be attached to the linker in the conjugate.

[0688] In some embodiments, the cytotoxic drug is linked to E in the antibody drug conjugate through a -OH, -SH, primary amine group, secondary amine group, or tertiary amine group on the cytotoxic drug.

[0689] In some embodiments, the cytotoxic drug is

[0690] In some embodiments, the cytotoxic drug is selected from the following Formula I and Formula II:

[0691]

[0692] Wherein, R1, R2 are each independently selected from C 1-6 Alkyl and halogen;

[0693] R3 is selected from H and -CO-CH2OH;

[0694] R4 and R5 are each independently selected from H, halogen and hydroxyl; or R4 and R5 are connected to the connected carbon atom to form a 5-6 membered oxygen-containing heterocyclic ring;

[0695] R6 is selected from hydrogen or -C 1-4 Alkylene-NR a R b ;

[0696] R7 is selected from hydrogen, C 1-6 Alkyl and -C 1-4 Alkylene-NR a R b ;

[0697] where R a 、R b Each occurrence is independently selected from H, C 1-6 Alkyl, -SO2-C 1-6 Alkyl and -CO-C 1-6 alkyl.

[0698] In some embodiments, the cytotoxic drug is selected from the following compounds:

[0699]

[0700]

[0701] In some embodiments, the cytotoxic drug is selected from the following compounds:

[0702]

[0703] The fragment of the cytotoxic drug obtained by connecting the cytotoxic drug to the linker is the formula Ab-[MLED] x In some embodiments, D is a monovalent structure obtained by losing one H from -OH, -NH2 or a secondary amine group on the cytotoxic drug.

[0704] In some embodiments, D is selected from the following structures:

[0705]

[0706] In some embodiments, the antibody drug conjugate is selected from ADC A-01 to ADC A-26, ADCB-01 to ADC B-06, and ADC C-01 shown below:

[0707] ADC A-01

[0708]

[0709] ADC A-02

[0710]

[0711] ADC A-03

[0712]

[0713] ADC A-04

[0714]

[0715] ADC A-05

[0716]

[0717] ADC A-06

[0718]

[0719] ADC A-07

[0720]

[0721] ADC A-08

[0722]

[0723] ADC A-09

[0724]

[0725] ADC A-10

[0726]

[0727] ADC A-11

[0728]

[0729] ADC A-12

[0730]

[0731] ADC A-13

[0732]

[0733] ADC A-14

[0734]

[0735] ADC A-15

[0736]

[0737] ADC A-16

[0738]

[0739] ADC A-17

[0740]

[0741] ADC A-18

[0742]

[0743] ADC A-19

[0744]

[0745] ADC A-20

[0746]

[0747] ADC A-21

[0748]

[0749] ADC A-22

[0750]

[0751] ADC A-23

[0752]

[0753] ADC A-24

[0754]

[0755] ADC A-25

[0756]

[0757] ADC A-26

[0758]

[0759] ADC B-01

[0760]

[0761] ADC B-02

[0762]

[0763] ADC B-03

[0764]

[0765] ADC B-04

[0766]

[0767] ADC B-05

[0768]

[0769] ADC B-06

[0770]

[0771] ADC C-01

[0772]

[0773] Wherein, HA in each antibody-drug conjugate represents an antibody or antigen-binding fragment of VH shown in SEQ ID NO: 1, 3, 5, 7 or 9 and VL shown in SEQ ID NO: 2, 4, 6, 8 or 10, for example, an antibody or antigen-binding fragment of VH shown in SEQ ID NO: 3 and CH shown in SEQ ID NO: 43 or 45, VL shown in SEQ ID NO: 4 and CL shown in SEQ ID NO: 44;

[0774] It refers to the connection formed by the sulfhydryl group of the antibody or antigen-binding fragment thereof and the linker; wherein HA is linked through the sulfhydryl group of the antibody or antigen-binding fragment to form an antibody-drug conjugate.

[0775] In some embodiments, the antibody drug conjugate is selected from:

[0776]

[0777]

[0778] wherein the HA of each antibody drug conjugate is selected from:

[0779] (1) An antibody or antigen-binding fragment thereof comprising: a VH having the amino acid sequence of SEQ ID NO: 1 and a CH having the amino acid sequence of SEQ ID NO: 43 or 45, and a VL having the amino acid sequence of SEQ ID NO: 2 and a CL having the amino acid sequence of SEQ ID NO: 44;

[0780] (2) an antibody or antigen-binding fragment thereof comprising: a VH having the amino acid sequence of SEQ ID NO: 3 and a CH having the amino acid sequence of SEQ ID NO: 43 or 45, and a VL having the amino acid sequence of SEQ ID NO: 4 and a CL having the amino acid sequence of SEQ ID NO: 44;

[0781] (3) an antibody or antigen-binding fragment thereof comprising: a VH having the amino acid sequence of SEQ ID NO: 5 and a CH having the amino acid sequence of SEQ ID NO: 43 or 45, and a light chain comprising a VL having the sequence of SEQ ID NO: 6 and a light chain constant region (CL) of SEQ ID NO: 44;

[0782] (4) an antibody or antigen-binding fragment thereof comprising: a VH having the amino acid sequence of SEQ ID NO: 7 and a CH having the amino acid sequence of SEQ ID NO: 43 or 45, and a VL having the amino acid sequence of SEQ ID NO: 8 and a CL having the amino acid sequence of SEQ ID NO: 44; and

[0783] (5) An antibody or antigen-binding fragment thereof comprising: a VH having the amino acid sequence of SEQ ID NO: 9 and a CH having the amino acid sequence of SEQ ID NO: 43 or 45, and a VL having the amino acid sequence of SEQ ID NO: 10 and a CL having the amino acid sequence of SEQ ID NO: 44,

[0784] represents a connection formed by a thiol group in an antibody or antigen-binding fragment thereof and a linker, wherein

[0785] HA is linked to the sulfhydryl group of the antibody or its antigen-binding fragment to form an antibody-drug conjugate.

[0786] And x is 3 to 9.

[0787] In some embodiments, the antibody drug conjugate is selected from:

[0788] ADC A-05

[0789]

[0790] ADC A-26

[0791]

[0792] ADC B-01

[0793]

[0794] ADC B-04

[0795]

[0796] ADC C-01

[0797]

[0798] in,

[0799] represents a connection between a thiol group in an antibody or antigen-binding fragment thereof and a linker, wherein

[0800] HA is linked to the sulfhydryl group of the antibody or its antigen-binding fragment to form an antibody-drug conjugate.

[0801] x is 2 to 8, 3 to 4, or 7 to 8, and the HA in each antibody-drug conjugate is selected from an antibody or antigen-binding fragment comprising:

[0802] (1) The following heavy chain variable region (VH) and / or light chain variable region (VL), wherein the CDRs are defined according to the Chothia numbering system:

[0803] (1a) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 11 or a variant thereof, CDR-H2 of SEQ ID NO: 12 or a variant thereof, and CDR-H3 of SEQ ID NO: 13 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 14 or a variant thereof, CDR-L2 of SEQ ID NO: 15 or a variant thereof, and CDR-L3 of SEQ ID NO: 16 or a variant thereof; or

[0804] (1b) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 27 or a variant thereof, CDR-H2 of SEQ ID NO: 28 or a variant thereof, and CDR-H3 of SEQ ID NO: 29 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 30 or a variant thereof, CDR-L2 of SEQ ID NO: 31 or a variant thereof, and CDR-L3 of SEQ ID NO: 32 or a variant thereof;

[0805] wherein the variant described in any one of (1a) and (1b) has at least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with the sequence from which it is derived, or the variant has one or more amino acid substitutions, deletions or additions (e.g., substitutions, deletions or additions of 1, 2 or 3 amino acids) compared to the sequence from which it is derived; preferably, the substitutions are conservative substitutions;

[0806] or,

[0807] (2) the following heavy chain variable region (VH) and / or light chain variable region (VL), wherein the CDRs are defined according to the Kabat numbering system:

[0808] (2a) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 17 or a variant thereof, CDR-H2 of SEQ ID NO: 18 or 19 or a variant thereof, and CDR-H3 of SEQ ID NO: 13 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 14 or a variant thereof, CDR-L2 of SEQ ID NO: 15 or a variant thereof, and CDR-L3 of SEQ ID NO: 16 or a variant thereof; or

[0809] (2b) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 33 or a variant thereof, CDR-H2 of SEQ ID NO: 34 or 35 or a variant thereof, and CDR-H3 of SEQ ID NO: 29 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 30 or a variant thereof, CDR-L2 of SEQ ID NO: 31 or a variant thereof, and CDR-L3 of SEQ ID NO: 32 or a variant thereof;

[0810] wherein the variant described in any one of (2a) and (2b) has at least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with the sequence from which it is derived, or the variant has one or more amino acid substitutions, deletions or additions (e.g., substitutions, deletions or additions of 1, 2 or 3 amino acids) compared to the sequence from which it is derived; preferably, the substitutions are conservative substitutions;

[0811] or,

[0812] (3) The following heavy chain variable region (VH) and / or light chain variable region (VL), wherein the CDRs are defined according to the IMGT numbering system:

[0813] (3a) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 20 or a variant thereof, CDR-H2 of SEQ ID NO: 21 or a variant thereof, and CDR-H3 of SEQ ID NO: 22 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 23 or a variant thereof, CDR-L2 of SEQ ID NO: 24 or a variant thereof, and CDR-L3 of SEQ ID NO: 16 or a variant thereof; or

[0814] (3b) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 36 or a variant thereof, CDR-H2 of SEQ ID NO: 37 or a variant thereof, and CDR-H3 of SEQ ID NO: 38 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 39 or a variant thereof, CDR-L2 of SEQ ID NO: 40 or a variant thereof, and CDR-L3 of SEQ ID NO: 32 or a variant thereof;

[0815] wherein the variant described in any one of (3a) and (3b) has at least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with the sequence from which it is derived, or the variant has one or more amino acid substitutions, deletions or additions (e.g., substitutions, deletions or additions of 1, 2 or 3 amino acids) compared to the sequence from which it is derived; preferably, the substitutions are conservative substitutions;

[0816] or,

[0817] (4) The following heavy chain variable region (VH) and / or light chain variable region (VL), wherein the CDRs are defined according to the AbM numbering system:

[0818] (4a) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 25 or a variant thereof, CDR-H2 of SEQ ID NO: 26 or a variant thereof, and CDR-H3 of SEQ ID NO: 13 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 14 or a variant thereof, CDR-L2 of SEQ ID NO: 15 or a variant thereof, and CDR-L3 of SEQ ID NO: 16 or a variant thereof; or

[0819] (4b) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 41 or a variant thereof, CDR-H2 of SEQ ID NO: 42 or a variant thereof, and CDR-H3 of SEQ ID NO: 29 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 30 or a variant thereof, CDR-L2 of SEQ ID NO: 31 or a variant thereof, and CDR-L3 of SEQ ID NO: 32 or a variant thereof;

[0820] Wherein, the variant described in any one of (4a) and (4b) has at least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity compared with the sequence from which it is derived, or the variant has one or more amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) compared with the sequence from which it is derived; preferably, the substitutions are conservative substitutions.

[0821] In another embodiment, the HA in each antibody drug conjugate is selected from an antibody or antigen-binding fragment comprising:

[0822] (a) VH or a variant thereof shown in SEQ ID NO: 1, and / or VL or a variant thereof shown in SEQ ID NO: 2;

[0823] (b) VH or a variant thereof shown in SEQ ID NO: 3, and / or VL or a variant thereof shown in SEQ ID NO: 4; or

[0824] (c) VH or a variant thereof shown in SEQ ID NO: 5, and / or VL or a variant thereof shown in SEQ ID NO: 6;

[0825] (d) VH or a variant thereof shown in SEQ ID NO: 7, and / or VL or a variant thereof shown in SEQ ID NO: 8;

[0826] (e) VH or a variant thereof shown in SEQ ID NO: 9, and / or VL or a variant thereof shown in SEQ ID NO: 10;

[0827] wherein the variant has at least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity compared to the sequence from which it is derived, or the variant has one or more amino acid substitutions, deletions or additions (e.g., 1, 2, 3, 4 or 5 amino acid substitutions, deletions or additions) compared to the sequence from which it is derived; preferably, the substitutions are conservative substitutions.

[0828] In another embodiment, the HA in each antibody drug conjugate is selected from an antibody or antigen-binding fragment comprising:

[0829] (a) VH shown in SEQ ID NO: 1, and / or VL shown in SEQ ID NO: 2;

[0830] (b) VH shown in SEQ ID NO: 3, and / or VL shown in SEQ ID NO: 4;

[0831] (c) VH represented by SEQ ID NO: 5, and / or VL represented by SEQ ID NO: 6;

[0832] (d) VH shown in SEQ ID NO: 7, and / or VL shown in SEQ ID NO: 8; and

[0833] (e) VH represented by SEQ ID NO: 9, and / or VL represented by SEQ ID NO: 10.

[0834] In another embodiment, the HA in each antibody-drug conjugate is an antibody or antigen-binding fragment comprising a heavy chain comprising a VH having an amino acid sequence selected from the group consisting of SEQ ID NOs: 1, 3, 5, 7, and 9.

[0835] In another embodiment, the HA in each antibody-drug conjugate is an antibody or antigen-binding fragment comprising a heavy chain comprising a VH having the amino acid sequence set forth in SEQ ID NO:1.

[0836] In another embodiment, the HA in each antibody-drug conjugate is an antibody or antigen-binding fragment comprising a heavy chain comprising a VH having the amino acid sequence set forth in SEQ ID NO:3.

[0837] In another embodiment, the HA in each antibody-drug conjugate is an antibody or antigen-binding fragment comprising a heavy chain comprising a VH having the amino acid sequence set forth in SEQ ID NO:5.

[0838] In another embodiment, the HA in each antibody-drug conjugate is an antibody or antigen-binding fragment comprising a heavy chain comprising a VH having the amino acid sequence set forth in SEQ ID NO:7.

[0839] In another embodiment, the HA in each antibody-drug conjugate is an antibody or antigen-binding fragment comprising a heavy chain comprising a VH having the amino acid sequence set forth in SEQ ID NO:9.

[0840] In another embodiment, the HA in each antibody-drug conjugate is an antibody or antigen-binding fragment comprising a heavy chain comprising a VH having an amino acid sequence selected from the group consisting of SEQ ID NOs: 1, 3, 5, 7, and 9, wherein the N-terminal glutamine or glutamic acid of the antibody variable region is cyclized to pyroglutamate.

[0841] In another embodiment, the HA in each antibody-drug conjugate is an antibody or antigen-binding fragment comprising a heavy chain comprising a VH having an amino acid sequence selected from SEQ ID NOs: 1, 3, 5, 7, and 9, wherein the N-terminal glutamine or glutamic acid of the antibody variable region is cyclized to pyroglutamate.

[0842] In another embodiment, the HA in each antibody-drug conjugate is an antibody or antigen-binding fragment comprising a heavy chain comprising a VH having the amino acid sequence shown in SEQ ID NO: 1, wherein the N-terminal glutamine or glutamic acid of the antibody variable region is cyclized to pyroglutamate.

[0843] In another embodiment, the HA in each antibody-drug conjugate is an antibody or antigen-binding fragment comprising a heavy chain comprising a VH having the amino acid sequence shown in SEQ ID NO: 1, wherein the N-terminal glutamine or glutamic acid of the antibody variable region is cyclized to pyroglutamic acid.

[0844] In another embodiment, the HA in each antibody-drug conjugate is an antibody or antigen-binding fragment comprising a heavy chain comprising a VH having the amino acid sequence shown in SEQ ID NO: 3, wherein the N-terminal glutamine or glutamic acid of the antibody variable region is cyclized to pyroglutamate.

[0845] In another embodiment, the HA in each antibody-drug conjugate is an antibody or antigen-binding fragment comprising a heavy chain comprising a VH having the amino acid sequence shown in SEQ ID NO: 3, wherein the N-terminal glutamine or glutamic acid of the antibody variable region is cyclized to pyroglutamic acid.

[0846] In another embodiment, the HA in each antibody-drug conjugate is an antibody or antigen-binding fragment comprising a heavy chain comprising a VH having the amino acid sequence shown in SEQ ID NO: 5, wherein the N-terminal glutamine or glutamic acid of the antibody variable region is cyclized to pyroglutamate.

[0847] In another embodiment, the HA in each antibody-drug conjugate is an antibody or antigen-binding fragment comprising a heavy chain comprising a VH having the amino acid sequence shown in SEQ ID NO: 5, wherein the N-terminal glutamine or glutamic acid of the antibody variable region is cyclized to pyroglutamic acid.

[0848] In another embodiment, the HA in each antibody-drug conjugate is an antibody or antigen-binding fragment comprising a heavy chain comprising a VH having the amino acid sequence shown in SEQ ID NO: 7, wherein the N-terminal glutamine or glutamic acid of the antibody variable region is cyclized to pyroglutamate.

[0849] In another embodiment, the HA in each antibody-drug conjugate is an antibody or antigen-binding fragment comprising a heavy chain comprising a VH having the amino acid sequence shown in SEQ ID NO: 7, wherein the N-terminal glutamine or glutamic acid of the antibody variable region is cyclized to pyroglutamic acid.

[0850] In another embodiment, the HA in each antibody-drug conjugate is an antibody or antigen-binding fragment comprising a heavy chain comprising a VH having the amino acid sequence set forth in SEQ ID NO:9, wherein the N-terminal glutamine or glutamic acid of the antibody variable region is cyclized to pyroglutamate.

[0851] In another embodiment, the HA in each antibody-drug conjugate is an antibody or antigen-binding fragment comprising a heavy chain comprising a VH having the amino acid sequence shown in SEQ ID NO: 9, wherein the N-terminal glutamine or glutamic acid of the antibody variable region is cyclized to pyroglutamic acid.

[0852] In another embodiment, the HA in each antibody-drug conjugate can be obtained by expressing in a host cell a nucleic acid molecule encoding the heavy and light chains of the antibody or antigen-binding fragment thereof, wherein the antibody or antigen-binding fragment thereof comprises:

[0853] (a) a VH having the amino acid sequence shown in SEQ ID NO: 1 or a variant thereof and / or a VL having the amino acid sequence shown in SEQ ID NO: 2 or a variant thereof;

[0854] (b) a VH having the amino acid sequence shown in SEQ ID NO: 3 or a variant thereof and / or a VL having the amino acid sequence shown in SEQ ID NO: 4 or a variant thereof;

[0855] (c) a VH having the amino acid sequence shown in SEQ ID NO: 5 or a variant thereof and / or a VL having the amino acid sequence shown in SEQ ID NO: 6 or a variant thereof;

[0856] (d) a VH having the amino acid sequence shown in SEQ ID NO: 7 or a variant thereof and / or a VL having the amino acid sequence shown in SEQ ID NO: 8 or a variant thereof; or

[0857] (e) a VH having the amino acid sequence shown in SEQ ID NO: 9 or a variant thereof and / or a VL having the amino acid sequence shown in SEQ ID NO: 10 or a variant thereof;

[0858] wherein the variant has at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% sequence identity compared to the sequence from which it is derived, or has one or more amino acid substitutions, deletions or additions (e.g., substitutions, deletions or additions of 1, 2, 3, 4 or 5 amino acids) compared to the sequence from which it is derived.

[0859] In another embodiment, the HA in each antibody-drug conjugate can be obtained by expressing in a host cell a nucleic acid molecule encoding the heavy and light chains of the antibody or antigen-binding fragment thereof, wherein the antibody or antigen-binding fragment thereof comprises:

[0860] (a) a VH having the amino acid sequence shown in SEQ ID NO: 1 and / or a VL having the amino acid sequence shown in SEQ ID NO: 2;

[0861] (b) a VH having the amino acid sequence shown in SEQ ID NO: 3 and / or a VL having the amino acid sequence shown in SEQ ID NO: 4;

[0862] (c) a VH having the amino acid sequence shown in SEQ ID NO: 5 and / or a VL having the amino acid sequence shown in SEQ ID NO: 6;

[0863] (d) a VH having the amino acid sequence shown in SEQ ID NO: 7 and / or a VL having the amino acid sequence shown in SEQ ID NO: 8; or

[0864] (e) VH having the amino acid sequence shown in SEQ ID NO: 9 and / or VL having the amino acid sequence shown in SEQ ID NO: 10.

[0865] In another embodiment, the HA in each antibody-drug conjugate can be obtained by expressing in a host cell a nucleic acid molecule encoding the heavy and light chains of the antibody or antigen-binding fragment thereof, wherein the antibody or antigen-binding fragment thereof comprises:

[0866] (a) VH having the amino acid sequence shown in SEQ ID NO: 1 or a variant thereof and / or VL having the amino acid sequence shown in SEQ ID NO: 2 or a variant thereof.

[0867] In another embodiment, the HA in each antibody-drug conjugate can be obtained by expressing in a host cell a nucleic acid molecule encoding the heavy and light chains of the antibody or antigen-binding fragment thereof, wherein the antibody or antigen-binding fragment thereof comprises:

[0868] (b) VH having the amino acid sequence shown in SEQ ID NO: 3 or a variant thereof and / or VL having the amino acid sequence shown in SEQ ID NO: 4 or a variant thereof.

[0869] In another embodiment, the HA in each antibody-drug conjugate can be obtained by expressing in a host cell a nucleic acid molecule encoding the heavy and light chains of the antibody or antigen-binding fragment thereof, wherein the antibody or antigen-binding fragment thereof comprises:

[0870] (c) VH having the amino acid sequence shown in SEQ ID NO: 5 or a variant thereof and / or VL having the amino acid sequence shown in SEQ ID NO: 6 or a variant thereof.

[0871] In another embodiment, the HA in each antibody-drug conjugate can be obtained by expressing in a host cell a nucleic acid molecule encoding the heavy and light chains of the antibody or antigen-binding fragment thereof, wherein the antibody or antigen-binding fragment thereof comprises:

[0872] (d) VH having the amino acid sequence shown in SEQ ID NO: 7 or a variant thereof and / or VL having the amino acid sequence shown in SEQ ID NO: 8 or a variant thereof.

[0873] In another embodiment, the HA in each antibody-drug conjugate can be obtained by expressing in a host cell a nucleic acid molecule encoding the heavy and light chains of the antibody or antigen-binding fragment thereof, wherein the antibody or antigen-binding fragment thereof comprises:

[0874] (e) VH having the amino acid sequence shown in SEQ ID NO: 9 or a variant thereof and / or VL having the amino acid sequence shown in SEQ ID NO: 10 or a variant thereof.

[0875] In another embodiment, the antibody drug conjugate is selected from:

[0876] ADC A-05

[0877]

[0878] ADC A-26

[0879]

[0880] ADC B-01

[0881]

[0882] ADC B-04

[0883]

[0884] ADC C-01

[0885]

[0886] in

[0887] The HA in the antibody-drug conjugate is selected from:

[0888] (1) an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 1 and a VL having the amino acid sequence of SEQ ID NO: 2, for example, an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 1 and a CH having the amino acid sequence of SEQ ID NO: 43 or 45, a VL having the amino acid sequence of SEQ ID NO: 2, and a CL having the amino acid sequence of SEQ ID NO: 44;

[0889] (2) an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence set forth in SEQ ID NO: 3 and a VL having the amino acid sequence set forth in SEQ ID NO: 4, for example, an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence set forth in SEQ ID NO: 3 and a CH having the amino acid sequence set forth in SEQ ID NO: 43 or 45, a VL having the amino acid sequence set forth in SEQ ID NO: 4, and a CL having the amino acid sequence set forth in SEQ ID NO: 44;

[0890] (3) an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 5 and a VL having the amino acid sequence of SEQ ID NO: 6, for example, an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 5 and a CH having the amino acid sequence of SEQ ID NO: 43 or 45, a VL having the amino acid sequence of SEQ ID NO: 6, and a CL having the amino acid sequence of SEQ ID NO: 44;

[0891] (4) an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 7 and a VL having the amino acid sequence of SEQ ID NO: 8, for example, an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 7 and a CH having the amino acid sequence of SEQ ID NO: 43 or 45, a VL having the amino acid sequence of SEQ ID NO: 8, and a CL having the amino acid sequence of SEQ ID NO: 44;

[0892] (5) an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 9 and a VL having the amino acid sequence of SEQ ID NO: 10, for example, an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 9 and a CH having the amino acid sequence of SEQ ID NO: 43 or 45, a VL having the amino acid sequence of SEQ ID NO: 10, and a CL having the amino acid sequence of SEQ ID NO: 44;

[0893] represents a connection between a thiol group in an antibody or antigen-binding fragment thereof and a linker; wherein

[0894] HA is linked to the sulfhydryl groups of the antibody or antigen-binding fragment to form an antibody-drug conjugate.

[0895] In another embodiment, the antibody drug conjugate is:

[0896]

[0897] in

[0898] The HA in the antibody-drug conjugate is selected from:

[0899] (1) an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 1 and a VL having the amino acid sequence of SEQ ID NO: 2, for example, an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 1 and a CH having the amino acid sequence of SEQ ID NO: 43 or 45, a VL having the amino acid sequence of SEQ ID NO: 2, and a CL having the amino acid sequence of SEQ ID NO: 44;

[0900] (2) an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence set forth in SEQ ID NO: 3 and a VL having the amino acid sequence set forth in SEQ ID NO: 4, for example, an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence set forth in SEQ ID NO: 3 and a CH having the amino acid sequence set forth in SEQ ID NO: 43 or 45, a VL having the amino acid sequence set forth in SEQ ID NO: 4, and a CL having the amino acid sequence set forth in SEQ ID NO: 44;

[0901] (3) an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 5 and a VL having the amino acid sequence of SEQ ID NO: 6, for example, an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 5 and a CH having the amino acid sequence of SEQ ID NO: 43 or 45, a VL having the amino acid sequence of SEQ ID NO: 6, and a CL having the amino acid sequence of SEQ ID NO: 44;

[0902] (4) an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 7 and a VL having the amino acid sequence of SEQ ID NO: 8, for example, an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 7 and a CH having the amino acid sequence of SEQ ID NO: 43 or 45, a VL having the amino acid sequence of SEQ ID NO: 8, and a CL having the amino acid sequence of SEQ ID NO: 44;

[0903] (5) an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 9 and a VL having the amino acid sequence of SEQ ID NO: 10, for example, an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 9 and a CH having the amino acid sequence of SEQ ID NO: 43 or 45, a VL having the amino acid sequence of SEQ ID NO: 10, and a CL having the amino acid sequence of SEQ ID NO: 44;

[0904] represents a connection between a thiol group in an antibody or antigen-binding fragment thereof and a linker; and

[0905] HA is linked to the sulfhydryl groups of the antibody or antigen-binding fragment to form an antibody-drug conjugate.

[0906] In another embodiment, the antibody drug conjugate is:

[0907]

[0908] The HA in the antibody-drug conjugate is an antibody or antigen-binding fragment comprising:

[0909] (1) an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 1 and a VL having the amino acid sequence of SEQ ID NO: 2, for example, an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 1 and a CH having the amino acid sequence of SEQ ID NO: 43 or 45, a VL having the amino acid sequence of SEQ ID NO: 2, and a CL having the amino acid sequence of SEQ ID NO: 44;

[0910] represents a connection between a thiol group in the antibody or antigen-binding fragment thereof and a linker; and

[0911] HA is linked to the sulfhydryl groups of the antibody or antigen-binding fragment to form an antibody-drug conjugate.

[0912] In another embodiment, the antibody drug conjugate is:

[0913]

[0914] wherein the HA in the antibody-drug conjugate is an antibody or antigen-binding fragment comprising: (2) an antibody or antigen-binding fragment thereof comprising a VH having an amino acid sequence as shown in SEQ ID NO: 3 and a VL having an amino acid sequence as shown in SEQ ID NO: 4, for example, an antibody or antigen-binding fragment thereof comprising a VH having an amino acid sequence as shown in SEQ ID NO: 3 and a CH having an amino acid sequence as shown in SEQ ID NO: 43 or 45 and a VL having an amino acid sequence as shown in SEQ ID NO: 4 and a CL having an amino acid sequence as shown in SEQ ID NO: 44;

[0915] represents a connection between a thiol group in the antibody or antigen-binding fragment thereof and a linker; and

[0916] HA is linked to the sulfhydryl groups of the antibody or antigen-binding fragment to form an antibody-drug conjugate.

[0917] In another embodiment, the antibody drug conjugate is:

[0918]

[0919] wherein the HA in the antibody-drug conjugate is an antibody or antigen-binding fragment comprising: (3) an antibody or antigen-binding fragment thereof comprising a VH having an amino acid sequence as shown in SEQ ID NO: 5 and a VL having an amino acid sequence as shown in SEQ ID NO: 6, for example, an antibody or antigen-binding fragment thereof comprising a VH having an amino acid sequence as shown in SEQ ID NO: 5 and a CH having an amino acid sequence as shown in SEQ ID NO: 43 or 45 and a VL having an amino acid sequence as shown in SEQ ID NO: 6 and a CL having an amino acid sequence as shown in SEQ ID NO: 44;

[0920] represents a connection between a thiol group in an antibody or antigen-binding fragment thereof and a linker; and

[0921] HA is linked to the sulfhydryl groups of the antibody or antigen-binding fragment to form an antibody-drug conjugate.

[0922] In another embodiment, the antibody drug conjugate is:

[0923]

[0924] wherein the HA in the antibody-drug conjugate is an antibody or antigen-binding fragment comprising: (4) an antibody or antigen-binding fragment thereof comprising a VH having an amino acid sequence as shown in SEQ ID NO: 7 and a VL having an amino acid sequence as shown in SEQ ID NO: 8, for example, an antibody or antigen-binding fragment thereof comprising a VH having an amino acid sequence as shown in SEQ ID NO: 7 and a CH having an amino acid sequence as shown in SEQ ID NO: 43 or 45 and a VL having an amino acid sequence as shown in SEQ ID NO: 8 and a CL having an amino acid sequence as shown in SEQ ID NO: 44;

[0925] represents a connection between a thiol group in the antibody or antigen-binding fragment thereof and a linker; and

[0926] HA is linked to the sulfhydryl groups of the antibody or antigen-binding fragment to form an antibody-drug conjugate.

[0927] In another embodiment, the antibody drug conjugate is:

[0928]

[0929] wherein the HA in the antibody-drug conjugate is an antibody or antigen-binding fragment comprising: (5) an antibody or antigen-binding fragment thereof comprising a VH having an amino acid sequence as shown in SEQ ID NO: 9 and a VL having an amino acid sequence as shown in SEQ ID NO: 10, for example, an antibody or antigen-binding fragment thereof comprising a VH having an amino acid sequence as shown in SEQ ID NO: 9 and a CH having an amino acid sequence as shown in SEQ ID NO: 43 or 45 and a VL having an amino acid sequence as shown in SEQ ID NO: 10 and a CL having an amino acid sequence as shown in SEQ ID NO: 44;

[0930] represents a connection between a thiol group in the antibody or antigen-binding fragment thereof and a linker; and

[0931] HA is linked to the sulfhydryl groups of the antibody or antigen-binding fragment to form an antibody-drug conjugate.

[0932] In another embodiment, the antibody drug conjugate is:

[0933]

[0934] Wherein the HA in the antibody-drug conjugate is an antibody or antigen-binding fragment comprising: an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence set forth in SEQ ID NO: 3 and a VL having the amino acid sequence set forth in SEQ ID NO: 4, for example, an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence set forth in SEQ ID NO: 3, a CH having the amino acid sequence set forth in SEQ ID NO: 43 or 45, a VL having the amino acid sequence set forth in SEQ ID NO: 4, and a CL having the amino acid sequence set forth in SEQ ID NO: 44;

[0935] x is 3 to 4 or 7 to 9,

[0936] represents a connection between a thiol group in the antibody or antigen-binding fragment thereof and a linker; and

[0937] HA is linked to the sulfhydryl groups of the antibody or antigen-binding fragment to form an antibody-drug conjugate.

[0938] In another embodiment, the antibody drug conjugate is:

[0939]

[0940] wherein HA in the antibody-drug conjugate is an antibody or antigen-binding fragment comprising: an antibody or antigen-binding fragment comprising a VH having the amino acid sequence set forth in SEQ ID NO: 3 and a VL having the amino acid sequence set forth in SEQ ID NO: 4, for example, an antibody or antigen-binding fragment comprising a VH having the amino acid sequence set forth in SEQ ID NO: 3, a CH having the amino acid sequence set forth in SEQ ID NO: 43 or 45, a VL having the amino acid sequence set forth in SEQ ID NO: 4, and a CL having the amino acid sequence set forth in SEQ ID NO: 44; and x is 7 to 9,

[0941] represents a connection between a thiol group in the antibody or antigen-binding fragment thereof and a linker; and

[0942] HA is linked to the sulfhydryl groups of the antibody or antigen-binding fragment to form an antibody-drug conjugate.

[0943] In another embodiment, the antibody drug conjugate is:

[0944]

[0945] wherein the HA in the antibody-drug conjugate is an antibody or antigen-binding fragment comprising: an antibody or antigen-binding fragment comprising a VH having the amino acid sequence set forth in SEQ ID NO: 3 and a VL having the amino acid sequence set forth in SEQ ID NO: 4, for example, an antibody or antigen-binding fragment comprising a VH having the amino acid sequence set forth in SEQ ID NO: 3, a CH having the amino acid sequence set forth in SEQ ID NO: 43 or 45, a VL having the amino acid sequence set forth in SEQ ID NO: 4, and a CL having the amino acid sequence set forth in SEQ ID NO: 44; and x is 8 to 9,

[0946] represents a connection between a thiol group in the antibody or antigen-binding fragment thereof and a linker; and

[0947] HA is linked to the sulfhydryl groups of the antibody or antigen-binding fragment to form an antibody-drug conjugate.

[0948] In another embodiment, the antibody drug conjugate is:

[0949]

[0950] Wherein the HA in the antibody-drug conjugate is an antibody or antigen-binding fragment comprising: an antibody or antigen-binding fragment comprising a VH having the amino acid sequence set forth in SEQ ID NO: 3 and a VL having the amino acid sequence set forth in SEQ ID NO: 4, for example, an antibody or antigen-binding fragment comprising a VH having the amino acid sequence set forth in SEQ ID NO: 3, a CH having the amino acid sequence set forth in SEQ ID NO: 43 or 45, a VL having the amino acid sequence set forth in SEQ ID NO: 4, and a CL having the amino acid sequence set forth in SEQ ID NO: 44; and x is 8,

[0951] represents a connection between a thiol group in the antibody or antigen-binding fragment thereof and a linker; and

[0952] HA is linked to the sulfhydryl groups of the antibody or antigen-binding fragment to form an antibody-drug conjugate.

[0953] In another embodiment, the antibody drug conjugate is:

[0954]

[0955] wherein HA is an antibody or an antigen-binding fragment thereof comprising a heavy chain (HC) having the amino acid sequence shown in SEQ ID NO: 46 and a light chain (LC) having the amino acid sequence shown in SEQ ID NO: 47; and x is 3 to 4 or 7 to 9,

[0956] represents a connection between a thiol group in the antibody or antigen-binding fragment thereof and a linker; and

[0957] HA is linked to the sulfhydryl groups of the antibody or antigen-binding fragment to form an antibody-drug conjugate.

[0958] In another embodiment, the antibody drug conjugate is:

[0959]

[0960] wherein HA is an antibody or an antigen-binding fragment thereof comprising a heavy chain (HC) having the amino acid sequence shown in SEQ ID NO: 46 and a light chain (LC) having the amino acid sequence shown in SEQ ID NO: 47; and x is 8 to 9,

[0961] represents a connection between a thiol group in the antibody or antigen-binding fragment thereof and a linker; and

[0962] HA is linked to the sulfhydryl groups of the antibody or antigen-binding fragment to form an antibody-drug conjugate.

[0963] In another embodiment, the antibody drug conjugate is:

[0964]

[0965] wherein HA is an antibody or an antigen-binding fragment thereof comprising a heavy chain (HC) having the amino acid sequence shown in SEQ ID NO: 46 and a light chain (LC) having the amino acid sequence shown in SEQ ID NO: 47; and x is 7 to 8,

[0966] represents a connection between a thiol group in the antibody or antigen-binding fragment thereof and a linker; and

[0967] HA is linked to the sulfhydryl groups of the antibody or antigen-binding fragment to form an antibody-drug conjugate.

[0968] In another embodiment, the antibody drug conjugate is:

[0969]

[0970] wherein HA is an antibody or an antigen-binding fragment thereof comprising a heavy chain (HC) having the amino acid sequence set forth in SEQ ID NO: 46 and a light chain (LC) having the amino acid sequence set forth in SEQ ID NO: 47;

[0971] x is 7 to 8,

[0972] represents a connection between a thiol group in the antibody or antigen-binding fragment thereof and a linker; and

[0973] HA is linked to the sulfhydryl group of cysteine ​​in the antibody or antigen-binding fragment to form an antibody-drug conjugate.

[0974] In another embodiment, the antibody drug conjugate is:

[0975]

[0976] wherein HA is an antibody or an antigen-binding fragment thereof comprising a heavy chain (HC) having the amino acid sequence shown in SEQ ID NO: 46 and a light chain (LC) having the amino acid sequence shown in SEQ ID NO: 47; and x is 8,

[0977] represents a connection between a thiol group in the antibody or antigen-binding fragment thereof and a linker; and

[0978] HA is linked to the sulfhydryl group of cysteine ​​in the antibody or antigen-binding fragment to form an antibody-drug conjugate.

[0979] In another embodiment, the antibody drug conjugate is:

[0980]

[0981] wherein HA is an antibody or an antigen-binding fragment thereof comprising a heavy chain (HC) having the amino acid sequence shown in SEQ ID NO: 52, 53 or 54 and a light chain (LC) having the amino acid sequence shown in SEQ ID NO: 47;

[0982] represents the connection between the sulfhydryl group in the antibody or antigen-binding fragment thereof and the linker;

[0983] x is 7 to 8, and

[0984] HA is linked to the sulfhydryl group of cysteine ​​in the antibody or antigen-binding fragment to form an antibody-drug conjugate.

[0985] In another embodiment, the antibody drug conjugate is:

[0986]

[0987] Wherein HA is an antibody composed of two heavy chains and two light chains, each heavy chain (HC) consists of the amino acid sequence set forth in SEQ ID NO: 52, and each light chain (LC) consists of the amino acid sequence set forth in SEQ ID NO: 47;

[0988] represents the connection between the sulfhydryl group in the antibody or antigen-binding fragment thereof and the linker;

[0989] x is 7 to 8, and

[0990] HA is linked to the sulfhydryl group of cysteine ​​in the antibody or antigen-binding fragment to form an antibody-drug conjugate.

[0991] In another embodiment, the antibody drug conjugate is:

[0992]

[0993] wherein HA is an antibody or an antigen-binding fragment thereof comprising a heavy chain (HC) having the amino acid sequence set forth in SEQ ID NO: 46 and a light chain (LC) having the amino acid sequence set forth in SEQ ID NO: 47;

[0994] represents a connection between a thiol group in an antibody or antigen-binding fragment thereof and a linker; wherein

[0995] HA is linked to the sulfhydryl group of cysteine ​​in the antibody or antigen-binding fragment to form an antibody-drug conjugate;

[0996] And x is 8.

[0997] In another embodiment, the antibody drug conjugate is:

[0998]

[0999] Wherein the HA in the antibody-drug conjugate is selected from:

[1000] (1) an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 1 and a VL having the amino acid sequence of SEQ ID NO: 2, for example, an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 1 and a CH having the amino acid sequence of SEQ ID NO: 43 or 45, a VL having the amino acid sequence of SEQ ID NO: 2, and a CL having the amino acid sequence of SEQ ID NO: 44;

[1001] (2) an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence set forth in SEQ ID NO: 3 and a VL having the amino acid sequence set forth in SEQ ID NO: 4, for example, an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence set forth in SEQ ID NO: 3 and a CH having the amino acid sequence set forth in SEQ ID NO: 43 or 45, a VL having the amino acid sequence set forth in SEQ ID NO: 4, and a CL having the amino acid sequence set forth in SEQ ID NO: 44;

[1002] (3) an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 5 and a VL having the amino acid sequence of SEQ ID NO: 6, for example, an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 5 and a CH having the amino acid sequence of SEQ ID NO: 43 or 45, a VL having the amino acid sequence of SEQ ID NO: 6, and a CL having the amino acid sequence of SEQ ID NO: 44;

[1003] (4) an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 7 and a VL having the amino acid sequence of SEQ ID NO: 8, for example, an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 7 and a CH having the amino acid sequence of SEQ ID NO: 43 or 45, a VL having the amino acid sequence of SEQ ID NO: 8, and a CL having the amino acid sequence of SEQ ID NO: 44;

[1004] (5) An antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence shown in SEQ ID NO: 9 and a VL having the amino acid sequence shown in SEQ ID NO: 10, for example, an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence shown in SEQ ID NO: 9 and a CH having the amino acid sequence shown in SEQ ID NO: 43 or 45, a VL having the amino acid sequence shown in SEQ ID NO: 10, and a CL having the amino acid sequence shown in SEQ ID NO: 44; wherein

[1005] represents a connection between a thiol group in an antibody or antigen-binding fragment thereof and a linker; and

[1006] HA is linked to the sulfhydryl groups of the antibody or antigen-binding fragment to form an antibody-drug conjugate.

[1007] In another embodiment, the antibody drug conjugate is:

[1008]

[1009] The HA in the antibody-drug conjugate is an antibody or antigen-binding fragment comprising:

[1010] (1) an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 1 and a VL having the amino acid sequence of SEQ ID NO: 2, for example, an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 1 and a CH having the amino acid sequence of SEQ ID NO: 43 or 45, a VL having the amino acid sequence of SEQ ID NO: 2, and a CL having the amino acid sequence of SEQ ID NO: 44;

[1011] represents a connection between a thiol group in the antibody or antigen-binding fragment thereof and a linker; and

[1012] HA is linked to the sulfhydryl groups of the antibody or antigen-binding fragment to form an antibody-drug conjugate.

[1013] In another embodiment, the antibody drug conjugate is:

[1014]

[1015] wherein the HA in the antibody-drug conjugate is an antibody or antigen-binding fragment comprising: (2) an antibody or antigen-binding fragment thereof comprising a VH having an amino acid sequence as shown in SEQ ID NO: 3 and a VL having an amino acid sequence as shown in SEQ ID NO: 4, for example, an antibody or antigen-binding fragment thereof comprising a VH having an amino acid sequence as shown in SEQ ID NO: 3 and a CH having an amino acid sequence as shown in SEQ ID NO: 43 or 45 and a VL having an amino acid sequence as shown in SEQ ID NO: 4 and a CL having an amino acid sequence as shown in SEQ ID NO: 44;

[1016] represents a connection between a thiol group in the antibody or antigen-binding fragment thereof and a linker; and

[1017] HA is linked to the sulfhydryl groups of the antibody or antigen-binding fragment to form an antibody-drug conjugate.

[1018] In a further embodiment, the antibody drug conjugate is:

[1019]

[1020] wherein the HA in the antibody-drug conjugate is an antibody or antigen-binding fragment comprising: (3) an antibody or antigen-binding fragment thereof comprising a VH having an amino acid sequence as shown in SEQ ID NO: 5 and a VL having an amino acid sequence as shown in SEQ ID NO: 6, for example, an antibody or antigen-binding fragment thereof comprising a VH having an amino acid sequence as shown in SEQ ID NO: 5 and a CH having an amino acid sequence as shown in SEQ ID NO: 43 or 45 and a VL having an amino acid sequence as shown in SEQ ID NO: 6 and a CL having an amino acid sequence as shown in SEQ ID NO: 44;

[1021] represents a connection between a thiol group in an antibody or antigen-binding fragment thereof and a linker; and

[1022] HA is linked to the sulfhydryl groups of the antibody or antigen-binding fragment to form an antibody-drug conjugate.

[1023] In a further embodiment, the antibody drug conjugate is:

[1024]

[1025] wherein the HA in the antibody-drug conjugate is an antibody or antigen-binding fragment comprising: (4) an antibody or antigen-binding fragment thereof comprising a VH having an amino acid sequence as shown in SEQ ID NO: 7 and a VL having an amino acid sequence as shown in SEQ ID NO: 8, for example, an antibody or antigen-binding fragment thereof comprising a VH having an amino acid sequence as shown in SEQ ID NO: 7 and a CH having an amino acid sequence as shown in SEQ ID NO: 43 or 45 and a VL having an amino acid sequence as shown in SEQ ID NO: 8 and a CL having an amino acid sequence as shown in SEQ ID NO: 44;

[1026] represents a connection between a thiol group in the antibody or antigen-binding fragment thereof and a linker; and

[1027] HA is linked to the sulfhydryl groups of the antibody or antigen-binding fragment to form an antibody-drug conjugate.

[1028] In another embodiment, the antibody drug conjugate is:

[1029]

[1030] wherein the HA in the antibody-drug conjugate is an antibody or antigen-binding fragment comprising: (5) an antibody or antigen-binding fragment thereof comprising a VH having an amino acid sequence as shown in SEQ ID NO: 9 and a VL having an amino acid sequence as shown in SEQ ID NO: 10, for example, an antibody or antigen-binding fragment thereof comprising a VH having an amino acid sequence as shown in SEQ ID NO: 9 and a CH having an amino acid sequence as shown in SEQ ID NO: 43 or 45 and a VL having an amino acid sequence as shown in SEQ ID NO: 10 and a CL having an amino acid sequence as shown in SEQ ID NO: 44;

[1031] represents a connection between a thiol group in the antibody or antigen-binding fragment thereof and a linker; and

[1032] HA is linked to the sulfhydryl groups of the antibody or antigen-binding fragment to form an antibody-drug conjugate.

[1033] In another embodiment, the antibody drug conjugate is:

[1034]

[1035] Wherein the HA in the antibody-drug conjugate is an antibody or antigen-binding fragment comprising: an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence set forth in SEQ ID NO: 3 and a VL having the amino acid sequence set forth in SEQ ID NO: 4, for example, an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence set forth in SEQ ID NO: 3, a CH having the amino acid sequence set forth in SEQ ID NO: 43 or 45, a VL having the amino acid sequence set forth in SEQ ID NO: 4, and a CL having the amino acid sequence set forth in SEQ ID NO: 44;

[1036] x is 7 to 9,

[1037] represents a connection between a thiol group in the antibody or antigen-binding fragment thereof and a linker; and

[1038] HA is linked to the sulfhydryl groups of the antibody or antigen-binding fragment to form an antibody-drug conjugate.

[1039] In another embodiment, the antibody drug conjugate is:

[1040]

[1041] Wherein the HA in the antibody-drug conjugate is an antibody or antigen-binding fragment comprising: an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence set forth in SEQ ID NO: 3 and a VL having the amino acid sequence set forth in SEQ ID NO: 4, for example, an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence set forth in SEQ ID NO: 3, a CH having the amino acid sequence set forth in SEQ ID NO: 43 or 45, a VL having the amino acid sequence set forth in SEQ ID NO: 4, and a CL having the amino acid sequence set forth in SEQ ID NO: 44;

[1042] x is 7 to 8,

[1043] represents a connection between a thiol group in the antibody or antigen-binding fragment thereof and a linker; and

[1044] HA is linked to the sulfhydryl groups of the antibody or antigen-binding fragment to form an antibody-drug conjugate.

[1045] In another embodiment, the antibody drug conjugate is:

[1046]

[1047] Wherein the HA in the antibody-drug conjugate is an antibody or antigen-binding fragment comprising: an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence set forth in SEQ ID NO: 3 and a VL having the amino acid sequence set forth in SEQ ID NO: 4, for example, an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence set forth in SEQ ID NO: 3, a CH having the amino acid sequence set forth in SEQ ID NO: 43 or 45, a VL having the amino acid sequence set forth in SEQ ID NO: 4, and a CL having the amino acid sequence set forth in SEQ ID NO: 44;

[1048] x is 8,

[1049] represents a connection between a thiol group in the antibody or antigen-binding fragment thereof and a linker; and

[1050] HA is linked to the sulfhydryl groups of the antibody or antigen-binding fragment to form an antibody-drug conjugate.

[1051] In another embodiment, the antibody drug conjugate is:

[1052]

[1053] Wherein the HA in the antibody-drug conjugate is an antibody or antigen-binding fragment comprising: an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence set forth in SEQ ID NO: 3 and a VL having the amino acid sequence set forth in SEQ ID NO: 4, for example, an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence set forth in SEQ ID NO: 3, a CH having the amino acid sequence set forth in SEQ ID NO: 43 or 45, a VL having the amino acid sequence set forth in SEQ ID NO: 4, and a CL having the amino acid sequence set forth in SEQ ID NO: 44;

[1054] x is 7 to 9,

[1055] represents a connection between a thiol group in the antibody or antigen-binding fragment thereof and a linker; and

[1056] HA is linked to the sulfhydryl groups of the antibody or antigen-binding fragment to form an antibody-drug conjugate.

[1057] In a further embodiment, the antibody drug conjugate is:

[1058]

[1059] wherein HA is an antibody or an antigen-binding fragment thereof comprising a heavy chain (HC) having the amino acid sequence set forth in SEQ ID NO: 46 and a light chain (LC) having the amino acid sequence set forth in SEQ ID NO: 47;

[1060] x is 7 to 8,

[1061] represents a connection between a thiol group in the antibody or antigen-binding fragment thereof and a linker; and

[1062] HA is linked to the sulfhydryl groups of the antibody or antigen-binding fragment to form an antibody-drug conjugate.

[1063] In another embodiment, the antibody drug conjugate is:

[1064]

[1065] wherein HA is an antibody or an antigen-binding fragment thereof comprising a heavy chain (HC) having the amino acid sequence set forth in SEQ ID NO: 46 and a light chain (LC) having the amino acid sequence set forth in SEQ ID NO: 47;

[1066] x is 8,

[1067] represents a connection between a thiol group in the antibody or antigen-binding fragment thereof and a linker; and

[1068] HA is linked to the sulfhydryl groups of the antibody or antigen-binding fragment to form an antibody-drug conjugate.

[1069] In another embodiment, the antibody drug conjugate is:

[1070]

[1071] in

[1072] The HA in the antibody-drug conjugate is selected from:

[1073] (1) an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 1 and a VL having the amino acid sequence of SEQ ID NO: 2, for example, an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 1 and a CH having the amino acid sequence of SEQ ID NO: 43 or 45, a VL having the amino acid sequence of SEQ ID NO: 2, and a CL having the amino acid sequence of SEQ ID NO: 44;

[1074] (2) an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence set forth in SEQ ID NO: 3 and a VL having the amino acid sequence set forth in SEQ ID NO: 4, for example, an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence set forth in SEQ ID NO: 3 and a CH having the amino acid sequence set forth in SEQ ID NO: 43 or 45, a VL having the amino acid sequence set forth in SEQ ID NO: 4, and a CL having the amino acid sequence set forth in SEQ ID NO: 44;

[1075] (3) an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 5 and a VL having the amino acid sequence of SEQ ID NO: 6, for example, an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 5 and a CH having the amino acid sequence of SEQ ID NO: 43 or 45, a VL having the amino acid sequence of SEQ ID NO: 6, and a CL having the amino acid sequence of SEQ ID NO: 44;

[1076] (4) an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 7 and a VL having the amino acid sequence of SEQ ID NO: 8, for example, an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 7 and a CH having the amino acid sequence of SEQ ID NO: 43 or 45, a VL having the amino acid sequence of SEQ ID NO: 8, and a CL having the amino acid sequence of SEQ ID NO: 44;

[1077] (5) An antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence shown in SEQ ID NO: 9 and a VL having the amino acid sequence shown in SEQ ID NO: 10, for example, an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence shown in SEQ ID NO: 9 and a CH having the amino acid sequence shown in SEQ ID NO: 43 or 45, a VL having the amino acid sequence shown in SEQ ID NO: 10, and a CL having the amino acid sequence shown in SEQ ID NO: 44.

[1078] In another embodiment, the antibody drug conjugate is:

[1079]

[1080] The HA in the antibody-drug conjugate is an antibody or antigen-binding fragment comprising:

[1081] (1) an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 1 and a VL having the amino acid sequence of SEQ ID NO: 2, for example, an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 1 and a CH having the amino acid sequence of SEQ ID NO: 43 or 45, a VL having the amino acid sequence of SEQ ID NO: 2, and a CL having the amino acid sequence of SEQ ID NO: 44;

[1082] represents a connection between a thiol group in the antibody or antigen-binding fragment thereof and a linker; and

[1083] HA is linked to the sulfhydryl groups of the antibody or antigen-binding fragment to form an antibody-drug conjugate.

[1084] In another embodiment, the antibody drug conjugate is:

[1085]

[1086] wherein the HA in the antibody-drug conjugate is an antibody or antigen-binding fragment comprising: (2) an antibody or antigen-binding fragment thereof comprising a VH having an amino acid sequence as shown in SEQ ID NO: 3 and a VL having an amino acid sequence as shown in SEQ ID NO: 4, for example, an antibody or antigen-binding fragment thereof comprising a VH having an amino acid sequence as shown in SEQ ID NO: 3 and a CH having an amino acid sequence as shown in SEQ ID NO: 43 or 45 and a VL having an amino acid sequence as shown in SEQ ID NO: 4 and a CL having an amino acid sequence as shown in SEQ ID NO: 44;

[1087] represents a connection between a thiol group in the antibody or antigen-binding fragment thereof and a linker; and

[1088] HA is linked to the sulfhydryl groups of the antibody or antigen-binding fragment to form an antibody-drug conjugate.

[1089] In another embodiment, the antibody drug conjugate is:

[1090]

[1091] wherein the HA in the antibody-drug conjugate is an antibody or antigen-binding fragment comprising: (3) an antibody or antigen-binding fragment thereof comprising a VH having an amino acid sequence as shown in SEQ ID NO: 5 and a VL having an amino acid sequence as shown in SEQ ID NO: 6, for example, an antibody or antigen-binding fragment thereof comprising a VH having an amino acid sequence as shown in SEQ ID NO: 5 and a CH having an amino acid sequence as shown in SEQ ID NO: 43 or 45 and a VL having an amino acid sequence as shown in SEQ ID NO: 6 and a CL having an amino acid sequence as shown in SEQ ID NO: 44;

[1092] represents a connection between a thiol group in an antibody or antigen-binding fragment thereof and a linker; and

[1093] HA is linked to the sulfhydryl groups of the antibody or antigen-binding fragment to form an antibody-drug conjugate.

[1094] In another embodiment, the antibody drug conjugate is:

[1095]

[1096] wherein the HA in the antibody-drug conjugate is an antibody or antigen-binding fragment comprising: (4) an antibody or antigen-binding fragment thereof comprising a VH having an amino acid sequence as shown in SEQ ID NO: 7 and a VL having an amino acid sequence as shown in SEQ ID NO: 8, for example, an antibody or antigen-binding fragment thereof comprising a VH having an amino acid sequence as shown in SEQ ID NO: 7 and a CH having an amino acid sequence as shown in SEQ ID NO: 43 or 45 and a VL having an amino acid sequence as shown in SEQ ID NO: 8 and a CL having an amino acid sequence as shown in SEQ ID NO: 44;

[1097] represents a connection between a thiol group in the antibody or antigen-binding fragment thereof and a linker; and

[1098] HA is linked to the sulfhydryl groups of the antibody or antigen-binding fragment to form an antibody-drug conjugate.

[1099] In another embodiment, the antibody drug conjugate is:

[1100]

[1101] wherein the HA in the antibody-drug conjugate is an antibody or antigen-binding fragment comprising: (5) an antibody or antigen-binding fragment thereof comprising a VH having an amino acid sequence as shown in SEQ ID NO: 9 and a VL having an amino acid sequence as shown in SEQ ID NO: 10, for example, an antibody or antigen-binding fragment thereof comprising a VH having an amino acid sequence as shown in SEQ ID NO: 9 and a CH having an amino acid sequence as shown in SEQ ID NO: 43 or 45 and a VL having an amino acid sequence as shown in SEQ ID NO: 10 and a CL having an amino acid sequence as shown in SEQ ID NO: 44;

[1102] represents a connection between a thiol group in the antibody or antigen-binding fragment thereof and a linker; and

[1103] HA is linked to the sulfhydryl groups of the antibody or antigen-binding fragment to form an antibody-drug conjugate.

[1104] In another embodiment, the antibody drug conjugate is:

[1105]

[1106] wherein HA is an antibody or an antigen-binding fragment thereof comprising a heavy chain (HC) having the amino acid sequence shown in SEQ ID NO: 46 and a light chain (LC) having the amino acid sequence shown in SEQ ID NO: 47; and x is 7 to 8,

[1107] represents a connection between a thiol group in the antibody or antigen-binding fragment thereof and a linker; and

[1108] HA is linked to the sulfhydryl groups of the antibody or antigen-binding fragment to form an antibody-drug conjugate.

[1109] In another embodiment, the antibody drug conjugate is:

[1110]

[1111] in

[1112] The HA in the antibody-drug conjugate is selected from:

[1113] (1) an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 1 and a VL having the amino acid sequence of SEQ ID NO: 2, for example, an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 1 and a CH having the amino acid sequence of SEQ ID NO: 43 or 45, a VL having the amino acid sequence of SEQ ID NO: 2, and a CL having the amino acid sequence of SEQ ID NO: 44;

[1114] (2) an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence set forth in SEQ ID NO: 3 and a VL having the amino acid sequence set forth in SEQ ID NO: 4, for example, an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence set forth in SEQ ID NO: 3 and a CH having the amino acid sequence set forth in SEQ ID NO: 43 or 45, a VL having the amino acid sequence set forth in SEQ ID NO: 4, and a CL having the amino acid sequence set forth in SEQ ID NO: 44;

[1115] (3) an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 5 and a VL having the amino acid sequence of SEQ ID NO: 6, for example, an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 5 and a CH having the amino acid sequence of SEQ ID NO: 43 or 45, a VL having the amino acid sequence of SEQ ID NO: 6, and a CL having the amino acid sequence of SEQ ID NO: 44;

[1116] (4) an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 7 and a VL having the amino acid sequence of SEQ ID NO: 8, for example, an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 7 and a CH having the amino acid sequence of SEQ ID NO: 43 or 45, a VL having the amino acid sequence of SEQ ID NO: 8, and a CL having the amino acid sequence of SEQ ID NO: 44;

[1117] (5) an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 9 and a VL having the amino acid sequence of SEQ ID NO: 10, for example, an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 9 and a CH having the amino acid sequence of SEQ ID NO: 43 or 45, a VL having the amino acid sequence of SEQ ID NO: 10, and a CL having the amino acid sequence of SEQ ID NO: 44; wherein

[1118] represents a connection between a thiol group in an antibody or antigen-binding fragment thereof and a linker; and

[1119] HA is linked to the sulfhydryl groups of the antibody or antigen-binding fragment to form an antibody-drug conjugate.

[1120] In another embodiment, the antibody drug conjugate is:

[1121]

[1122] The HA in the antibody-drug conjugate is an antibody or antigen-binding fragment comprising:

[1123] (1) an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 1 and a VL having the amino acid sequence of SEQ ID NO: 2, for example, an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 1 and a CH having the amino acid sequence of SEQ ID NO: 43 or 45, a VL having the amino acid sequence of SEQ ID NO: 2, and a CL having the amino acid sequence of SEQ ID NO: 44;

[1124] represents a connection between a thiol group in the antibody or antigen-binding fragment thereof and a linker; and

[1125] HA is linked to the sulfhydryl groups of the antibody or antigen-binding fragment to form an antibody-drug conjugate.

[1126] In another embodiment, the antibody drug conjugate is:

[1127]

[1128] wherein the HA in the antibody-drug conjugate is an antibody or antigen-binding fragment comprising: (2) an antibody or antigen-binding fragment thereof comprising a VH having an amino acid sequence as shown in SEQ ID NO: 3 and a VL having an amino acid sequence as shown in SEQ ID NO: 4, for example, an antibody or antigen-binding fragment thereof comprising a VH having an amino acid sequence as shown in SEQ ID NO: 3 and a CH having an amino acid sequence as shown in SEQ ID NO: 43 or 45 and a VL having an amino acid sequence as shown in SEQ ID NO: 4 and a CL having an amino acid sequence as shown in SEQ ID NO: 44;

[1129] represents a connection between a thiol group in the antibody or antigen-binding fragment thereof and a linker; and

[1130] HA is linked to the sulfhydryl groups of the antibody or antigen-binding fragment to form an antibody-drug conjugate.

[1131] In another embodiment, the antibody drug conjugate is:

[1132]

[1133] wherein the HA in the antibody-drug conjugate is an antibody or antigen-binding fragment comprising: (3) an antibody or antigen-binding fragment thereof comprising a VH having an amino acid sequence as shown in SEQ ID NO: 5 and a VL having an amino acid sequence as shown in SEQ ID NO: 6, for example, an antibody or antigen-binding fragment thereof comprising a VH having an amino acid sequence as shown in SEQ ID NO: 5 and a CH having an amino acid sequence as shown in SEQ ID NO: 43 or 45 and a VL having an amino acid sequence as shown in SEQ ID NO: 6 and a CL having an amino acid sequence as shown in SEQ ID NO: 44;

[1134] represents a connection between a thiol group in an antibody or antigen-binding fragment thereof and a linker; and

[1135] HA is linked to the sulfhydryl groups of the antibody or antigen-binding fragment to form an antibody-drug conjugate.

[1136] In another embodiment, the antibody drug conjugate is:

[1137]

[1138] wherein the HA in the antibody-drug conjugate is an antibody or antigen-binding fragment comprising: (4) an antibody or antigen-binding fragment thereof comprising a VH having an amino acid sequence as shown in SEQ ID NO: 7 and a VL having an amino acid sequence as shown in SEQ ID NO: 8, for example, an antibody or antigen-binding fragment thereof comprising a VH having an amino acid sequence as shown in SEQ ID NO: 7 and a CH having an amino acid sequence as shown in SEQ ID NO: 43 or 45 and a VL having an amino acid sequence as shown in SEQ ID NO: 8 and a CL having an amino acid sequence as shown in SEQ ID NO: 44;

[1139] represents a connection between a thiol group in the antibody or antigen-binding fragment thereof and a linker; and

[1140] HA is linked to the sulfhydryl groups of the antibody or antigen-binding fragment to form an antibody-drug conjugate.

[1141] In another embodiment, the antibody drug conjugate is:

[1142]

[1143] wherein the HA in the antibody-drug conjugate is an antibody or antigen-binding fragment comprising: (5) an antibody or antigen-binding fragment thereof comprising a VH having an amino acid sequence as shown in SEQ ID NO: 9 and a VL having an amino acid sequence as shown in SEQ ID NO: 10, for example, an antibody or antigen-binding fragment thereof comprising a VH having an amino acid sequence as shown in SEQ ID NO: 9 and a CH having an amino acid sequence as shown in SEQ ID NO: 43 or 45 and a VL having an amino acid sequence as shown in SEQ ID NO: 10 and a CL having an amino acid sequence as shown in SEQ ID NO: 44;

[1144] represents a connection between a thiol group in the antibody or antigen-binding fragment thereof and a linker; and

[1145] HA is linked to the sulfhydryl groups of the antibody or antigen-binding fragment to form an antibody-drug conjugate.

[1146] In another embodiment, the antibody drug conjugate is:

[1147]

[1148] Wherein the HA in the antibody-drug conjugate is an antibody or antigen-binding fragment comprising: an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence set forth in SEQ ID NO: 3 and a VL having the amino acid sequence set forth in SEQ ID NO: 4, for example, an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence set forth in SEQ ID NO: 3, a CH having the amino acid sequence set forth in SEQ ID NO: 43 or 45, a VL having the amino acid sequence set forth in SEQ ID NO: 4, and a CL having the amino acid sequence set forth in SEQ ID NO: 44;

[1149] x is 7 to 9,

[1150] represents a connection between a thiol group in the antibody or antigen-binding fragment thereof and a linker; and

[1151] HA is linked to the sulfhydryl groups of the antibody or antigen-binding fragment to form an antibody-drug conjugate.

[1152] In another embodiment, the antibody drug conjugate is:

[1153]

[1154] Wherein the HA in the antibody-drug conjugate is an antibody or antigen-binding fragment comprising: an antibody or antigen-binding fragment comprising a VH having the amino acid sequence set forth in SEQ ID NO: 3 and a VL having the amino acid sequence set forth in SEQ ID NO: 4, for example, an antibody or antigen-binding fragment comprising a VH having the amino acid sequence set forth in SEQ ID NO: 3, a CH having the amino acid sequence set forth in SEQ ID NO: 43 or 45, a VL having the amino acid sequence set forth in SEQ ID NO: 4, and a CL having the amino acid sequence set forth in SEQ ID NO: 47;

[1155] x is 7 to 9,

[1156] represents a connection between a thiol group in the antibody or antigen-binding fragment thereof and a linker; and

[1157] HA is linked to the sulfhydryl groups of the antibody or antigen-binding fragment to form an antibody-drug conjugate.

[1158] In another embodiment, the antibody drug conjugate is:

[1159]

[1160] wherein HA is an antibody or an antigen-binding fragment thereof comprising a heavy chain (HC) having the amino acid sequence set forth in SEQ ID NO: 46 and a light chain (LC) having the amino acid sequence set forth in SEQ ID NO: 47;

[1161] x is 7 to 8,

[1162] represents a connection between a thiol group in the antibody or antigen-binding fragment thereof and a linker; and

[1163] HA is linked to the sulfhydryl groups of the antibody or antigen-binding fragment to form an antibody-drug conjugate.

[1164] In another embodiment, the antibody drug conjugate is:

[1165]

[1166] wherein HA is an antibody or an antigen-binding fragment thereof comprising a heavy chain (HC) having the amino acid sequence set forth in SEQ ID NO: 46 and a light chain (LC) having the amino acid sequence set forth in SEQ ID NO: 47;

[1167] x is 8 to 9,

[1168] represents a connection between a thiol group in the antibody or antigen-binding fragment thereof and a linker; and

[1169] HA is linked to the sulfhydryl groups of the antibody or antigen-binding fragment to form an antibody-drug conjugate.

[1170] In another embodiment, the antibody drug conjugate is:

[1171]

[1172] wherein HA is an antibody or an antigen-binding fragment thereof comprising a heavy chain (HC) having the amino acid sequence set forth in SEQ ID NO: 46 and a light chain (LC) having the amino acid sequence set forth in SEQ ID NO: 47;

[1173] x is 8,

[1174] represents a connection between a thiol group in the antibody or antigen-binding fragment thereof and a linker; and

[1175] HA is linked to the sulfhydryl groups of the antibody or antigen-binding fragment to form an antibody-drug conjugate.

[1176] In another embodiment, the antibody drug conjugate is:

[1177]

[1178] in

[1179] The HA in the antibody-drug conjugate is selected from:

[1180] (1) an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 1 and a VL having the amino acid sequence of SEQ ID NO: 2, for example, an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 1 and a CH having the amino acid sequence of SEQ ID NO: 43 or 45, a VL having the amino acid sequence of SEQ ID NO: 2, and a CL having the amino acid sequence of SEQ ID NO: 44;

[1181] (2) an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence set forth in SEQ ID NO: 3 and a VL having the amino acid sequence set forth in SEQ ID NO: 4, for example, an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence set forth in SEQ ID NO: 3 and a CH having the amino acid sequence set forth in SEQ ID NO: 43 or 45, a VL having the amino acid sequence set forth in SEQ ID NO: 4, and a CL having the amino acid sequence set forth in SEQ ID NO: 44;

[1182] (3) an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 5 and a VL having the amino acid sequence of SEQ ID NO: 6, for example, an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 5 and a CH having the amino acid sequence of SEQ ID NO: 43 or 45, a VL having the amino acid sequence of SEQ ID NO: 6, and a CL having the amino acid sequence of SEQ ID NO: 44;

[1183] (4) an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 7 and a VL having the amino acid sequence of SEQ ID NO: 8, for example, an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 7 and a CH having the amino acid sequence of SEQ ID NO: 43 or 45, a VL having the amino acid sequence of SEQ ID NO: 8, and a CL having the amino acid sequence of SEQ ID NO: 44;

[1184] (5) An antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence shown in SEQ ID NO: 9 and a VL having the amino acid sequence shown in SEQ ID NO: 10, for example, an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence shown in SEQ ID NO: 9 and a CH having the amino acid sequence shown in SEQ ID NO: 43 or 45, a VL having the amino acid sequence shown in SEQ ID NO: 10, and a CL having the amino acid sequence shown in SEQ ID NO: 44.

[1185] In a further embodiment, the antibody drug conjugate is:

[1186]

[1187] The HA in the antibody-drug conjugate is an antibody or antigen-binding fragment comprising:

[1188] (1) An antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence set forth in SEQ ID NO: 1 and a VL having the amino acid sequence set forth in SEQ ID NO: 2, for example, an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence set forth in SEQ ID NO: 1 and a CH having the amino acid sequence set forth in SEQ ID NO: 43 or 45, a VL having the amino acid sequence set forth in SEQ ID NO: 2, and a CL having the amino acid sequence set forth in SEQ ID NO: 44.

[1189] In another embodiment, the antibody drug conjugate is:

[1190]

[1191] Wherein the HA in the antibody-drug conjugate is an antibody or antigen-binding fragment comprising the following: (2) an antibody or antigen-binding fragment comprising a VH having an amino acid sequence as shown in SEQ ID NO: 3 and a VL having an amino acid sequence as shown in SEQ ID NO: 4, for example, an antibody or antigen-binding fragment comprising a VH having an amino acid sequence as shown in SEQ ID NO: 3 and a CH having an amino acid sequence as shown in SEQ ID NO: 43 or 45 and a VL having an amino acid sequence as shown in SEQ ID NO: 4 and a CL having an amino acid sequence as shown in SEQ ID NO: 44.

[1192] In another embodiment, the antibody drug conjugate is:

[1193]

[1194] Wherein the HA in the antibody-drug conjugate is an antibody or antigen-binding fragment comprising the following: (3) an antibody or antigen-binding fragment thereof comprising a VH having an amino acid sequence as shown in SEQ ID NO: 5 and a VL having an amino acid sequence as shown in SEQ ID NO: 6, for example, an antibody or antigen-binding fragment thereof comprising a VH having an amino acid sequence as shown in SEQ ID NO: 5 and a CH having an amino acid sequence as shown in SEQ ID NO: 43 or 45 and a VL having an amino acid sequence as shown in SEQ ID NO: 6 and a CL having an amino acid sequence as shown in SEQ ID NO: 44.

[1195] In another embodiment, the antibody drug conjugate is:

[1196]

[1197] The HA in the antibody-drug conjugate is an antibody or antigen-binding fragment comprising the following: (4) an antibody or antigen-binding fragment comprising a VH having an amino acid sequence as shown in SEQ ID NO: 7 and a VL having an amino acid sequence as shown in SEQ ID NO: 8, for example, an antibody or antigen-binding fragment comprising a VH having an amino acid sequence as shown in SEQ ID NO: 7 and a CH having an amino acid sequence as shown in SEQ ID NO: 43 or 45 and a VL having an amino acid sequence as shown in SEQ ID NO: 8 and a CL having an amino acid sequence as shown in SEQ ID NO: 44.

[1198] In another embodiment, the antibody drug conjugate is:

[1199]

[1200] Wherein the HA in the antibody-drug conjugate is an antibody or antigen-binding fragment comprising the following: (5) an antibody or antigen-binding fragment thereof comprising a VH having an amino acid sequence as shown in SEQ ID NO: 9 and a VL having an amino acid sequence as shown in SEQ ID NO: 10, for example, an antibody or antigen-binding fragment thereof comprising a VH having an amino acid sequence as shown in SEQ ID NO: 9 and a CH having an amino acid sequence as shown in SEQ ID NO: 43 or 45 and a VL having an amino acid sequence as shown in SEQ ID NO: 10 and a CL having an amino acid sequence as shown in SEQ ID NO: 44.

[1201] In another embodiment, the antibody drug conjugate is:

[1202]

[1203] wherein HA is an antibody or an antigen-binding fragment thereof comprising a heavy chain (HC) having the amino acid sequence shown in SEQ ID NO: 46 and a light chain (LC) having the amino acid sequence shown in SEQ ID NO: 47; and x is 2 to 5 or 3 to 4.

[1204] In some embodiments, x of the antibody-drug conjugate (ADC) is 1 to 10, e.g., 1 to 2, 1 to 3, 1 to 4, 1 to 5, 1 to 6, 1 to 7, 1 to 8, 1 to 9, 1 to 10, 2 to 3, 2 to 4, 2 to 5, 2 to 6, 2 to 7, 2 to 8, 2 to 9, 2 to 10, 3 to 4, 3 to 5, 3 to 6, 3 to 7, 3 to 8, 3 to 9, 3 to 10, 4 to 5, 4 to 6, 4 to 7, 4 to 8, 4 to 9, 4 to 10, 5 to 6, 5 to 7, 5 to 8, 5 to 9, 5 to 10, 6 to 7, 6 to 8, 6 to 9, 6 to 10, 7 to 8, 7 to 9, 7 to 10, 8 to 9, 8 to 10, or 9 to 10.

[1205] In some embodiments, x of the antibody-drug conjugate (ADC) is 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10.

[1206] In some embodiments, the antibodies disclosed herein are genetically engineered to contain one or more cysteine ​​or lysine residues or non-canonical amino acid substitutions of amino acids at defined positions within the antibody.

[1207] In some embodiments, the antibody disclosed herein is genetically engineered to include one or more cysteines or amino acid whose non-conventional amino acid substitutions in the restricted position within the antibody. Then, these cysteine ​​residues or non-conventional amino acid residues can be coupled to the drug linker by the sulfhydryl group of the cysteine ​​residues or the reactive group of non-conventional amino acid. Therefore, the antibody-drug conjugate of the present invention can include the replacement of one or more amino acids in the heavy chain or light chain of the antibody by cysteine ​​residues or non-conventional amino acid residues, which are then coupled to the drug linker disclosed herein. In a specific embodiment, the amino acid position that can be substituted is selected from position 152, 153, 171, 172, 173 and 375 (numbering according to the Eu numbering scheme) of the heavy chain constant domain and position 165 and 168 (numbering starting from the amino acid 1 of the N-terminal) of the light chain constant domain. In certain embodiments, cysteine ​​may replace one or more amino acids at positions 152, 153, 171, 172, 173, and 375 of the heavy chain constant domain (numbered according to the Eu numbering scheme) and positions 165 and 168 of the light chain constant region (numbered starting from amino acid 1 at the N-terminus). In certain embodiments, the antibody-drug conjugate comprises an S375C amino acid substitution coupled to a drug linker disclosed herein. In certain embodiments, the antibody comprises an S375C amino acid substitution and an E152C amino acid substitution, each coupled to a drug linker disclosed herein. In certain embodiments, the antibody comprises an S375C amino acid substitution and an S168C amino acid substitution, each coupled to a drug linker disclosed herein.In some embodiments, as explained herein, the DAR value (drug to antibody ratio) of the antibody drug conjugate is 1-10, for example: 1-2, 1-3, 1-4, 1-5, 1-6, 1-7, 1-8, 1-9, 1-10, 2-3, 2-4, 2-5, 2-6, 2-7, 2-8, 2-9, 2-10, 3-4, 3-5, 3-6, 3-7, 3-8, 3-9, 3-10, 4-5, 4-6, 4-7, 4-8, 4-9, 4-10, 5-6, 5-7, 5-8, 5-9, 5-10, 6-7, 6-8, 6-9, 6-10, 7-8, 7-9, 7-10, 8-9, 8-10, or 9-10, preferably 3-9, for example, 3.0-3.5, 3.0-4.0, 3.0-4.5, 3.0-5.0, 3.0-5.5, 3.0-6.0, 3.5-4.0, 3.5-4.5, 3.5-5.0, 3.5-5.5 , 3.5~6.0,3.5~6.5,3.5~7.0,3.5~7.5,3.5~8.0,4.0~4.5,4.0~5.0,4.0~5.5,4.0~6.0,4.0~6.5,4.0~7.0,4.0~7.5,4.0~8.0,4.5~5.0,4.5~5.5,4.5~6.0,4.5~6.5,4.5~7.0,4.5~7.5,4.5~8.0,5.0 ~5.5, 5.0~6.0, 5.0~6.5, 5.0~7.0, 5.0~7.5, 5.0~8.0, 5.5~6.0, 5.5~6.5, 5.5~7.0, 5.5~7.5, 5.5~8.0, 6.0~6.5, 6.0~7.0, 6.0~7.5, 6.0~8.5, 6.5~7.0, 6.5~7.5, 6.5~8.5, 7.0~7.5, 7.0~9.0 or 7.5~9.0.

[1208] Drug-linker

[1209] Those skilled in the art will appreciate that the antibody-drug conjugates described herein can be prepared modularly. For example, a free-form "drug-linker" (which can be understood as M'-LED, where M' is the structural form of M before covalently linking to the antibody or its antigen-binding fragment) is first obtained, and then the free-form "drug-linker" is covalently linked to the antibody or its antigen-binding fragment to obtain the antibody-drug conjugate described herein. Accordingly, one or more sulfhydryl (-SH), amino (-NH2) or carboxyl (-COOH) groups on the antibody or its antigen-binding fragment are linked to M' in the free-form "drug-linker" by a substitution reaction (e.g., removal of structures such as -SO2Me or -Br thereon) or by an addition reaction.

[1210] In another aspect, the present invention provides a drug-linker having a structure shown in formula M'-LED, wherein:

[1211] M' is Lg is a leaving group for a nucleophilic substitution reaction (e.g.,

[1212] Halogen, methylsulfonyl, fluorophenol or ), or is hydroxyl (-OH), thiol (-SH) or amino (-NH2); or, Lg forms an unsaturated double bond with the adjacent atoms on ring A; Ring A is a 5-6 membered alicyclic heterocyclic ring, or a 5-20 membered aromatic ring system, wherein the alicyclic heterocyclic ring and the aromatic ring system are optionally substituted with one or more selected from oxy (=O), halogen, cyano, amino, carboxyl, thiol and C 1-6 Alkyl group substituted; M1 is selected from single bond and C 1-20 Alkylene, C 2-20 Alkenylene or C 2-20 Alkynylidene;

[1213] The L, E and D structures are as defined in any of the above antibody drug conjugates.

[1214] In some embodiments, M' is Lg is a methylsulfonyl group, or Lg forms a carbon-carbon double bond with the adjacent atoms on ring A; Ring A is a 5-membered alicyclic heterocycle, a 6-membered heteroaromatic ring, or a polycyclic ring formed by connecting one or more 6-membered heteroaromatic rings to a benzene ring via a single bond, wherein the alicyclic heterocycle is optionally substituted with one or more selected from oxy (=O), halogen and C 1-4 Alkyl group substituted; M1 is selected from single bond and C 3-10 Alkylene, C 3-10 Alkenylene or C 3-10 Alkynylidene.

[1215] In some embodiments, M' is Selected from M1 is selected from a single bond and C 5-8 Alkylene, C 5-8 Alkenylene or C 5-8 Alkynylidene.

[1216] In some embodiments, M' is selected from

[1217] In some embodiments, M' is

[1218] In some embodiments, the free form of the "drug-linker" is selected from A-01 to A-26, B-01 to B-06, and C-01 shown below:

[1219] A-01:

[1220] A-02:

[1221] A-03:

[1222] A-04:

[1223] A-05:

[1224] A-06:

[1225] A-07:

[1226] A-08:

[1227] A-09:

[1228] A-10:

[1229] A-11:

[1230] A-12:

[1231] A-13:

[1232] A-14:

[1233] A-15:

[1234] A-16:

[1235] A-17:

[1236] A-18:

[1237] A-19:

[1238] A-20:

[1239] A-21:

[1240] A-22:

[1241] A-23:

[1242] A-24:

[1243] A-25:

[1244] A-26:

[1245] B-01:

[1246] B-02:

[1247] B-03:

[1248] B-04:

[1249] B-05:

[1250] B-06:

[1251] C-01:

[1252] The present invention also provides an ADC comprising an antibody that binds to PTK7, wherein the antibody is conjugated to a drug linker selected from the group consisting of A-01, A-02, A-03, A-04, A-05, A-06, A-07, A-08, A-09, A-10, A-12, A13, A-14, A15, A-16, A-17, A-18, A-19, A-20, A-21, A-22, A-23, A-24, A-25, A-26, B-01, B-02, B-03, B-04, B-05, B-06, and C-01 through non-canonical amino acid substitutions of cysteine ​​or lysine or amino acids within the antibody residues.

[1253] The present invention also provides an ADC comprising an antibody that binds to PTK7, wherein the antibody is conjugated to a drug linker selected from the group consisting of A-01, A-02, A-03, A-04, A-05, A-06, A-07, A-08, A-09, A-10, A-12, A13, A-14, A15, A-16, A-17, A-18, A-19, A-20, A-21, A-22, A-23, A-24, A-25, A-26, B-01, B-02, B-03, B-04, B-05, B-06, and C-01 through non-canonical amino acid substitutions of cysteine ​​or amino acids within the antibody residues.

[1254] In another embodiment of the ADC, the antibody is an antibody or antigen-binding fragment thereof selected from:

[1255] (1) The following heavy chain variable region (VH) and / or light chain variable region (VL), wherein the CDRs are defined according to the Chothia numbering system:

[1256] (1a) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 11 or a variant thereof, CDR-H2 of SEQ ID NO: 12 or a variant thereof, and CDR-H3 of SEQ ID NO: 13 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 14 or a variant thereof, CDR-L2 of SEQ ID NO: 15 or a variant thereof, and CDR-L3 of SEQ ID NO: 16 or a variant thereof; or

[1257] (1b) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 27 or a variant thereof, CDR-H2 of SEQ ID NO: 28 or a variant thereof, and CDR-H3 of SEQ ID NO: 29 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 30 or a variant thereof, CDR-L2 of SEQ ID NO: 31 or a variant thereof, and CDR-L3 of SEQ ID NO: 32 or a variant thereof;

[1258] wherein the variant described in any one of (1a) and (1b) has at least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with the sequence from which it is derived, or the variant has one or more amino acid substitutions, deletions or additions (e.g., substitutions, deletions or additions of 1, 2 or 3 amino acids) compared to the sequence from which it is derived; preferably, the substitutions are conservative substitutions;

[1259] or,

[1260] (2) the following heavy chain variable region (VH) and / or light chain variable region (VL), wherein the CDRs are defined according to the Kabat numbering system:

[1261] (2a) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 17 or a variant thereof, CDR-H2 of SEQ ID NO: 18 or 19 or a variant thereof, and CDR-H3 of SEQ ID NO: 13 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 14 or a variant thereof, CDR-L2 of SEQ ID NO: 15 or a variant thereof, and CDR-L3 of SEQ ID NO: 16 or a variant thereof; or

[1262] (2b) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 33 or a variant thereof, CDR-H2 of SEQ ID NO: 34 or 35 or a variant thereof, and CDR-H3 of SEQ ID NO: 29 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 30 or a variant thereof, CDR-L2 of SEQ ID NO: 31 or a variant thereof, and CDR-L3 of SEQ ID NO: 32 or a variant thereof;

[1263] wherein the variant described in any one of (2a) and (2b) has at least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with the sequence from which it is derived, or the variant has one or more amino acid substitutions, deletions or additions (e.g., substitutions, deletions or additions of 1, 2 or 3 amino acids) compared to the sequence from which it is derived; preferably, the substitutions are conservative substitutions;

[1264] or,

[1265] (3) The following heavy chain variable region (VH) and / or light chain variable region (VL), wherein the CDRs are defined according to the IMGT numbering system:

[1266] (3a) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 20 or a variant thereof, CDR-H2 of SEQ ID NO: 21 or a variant thereof, and CDR-H3 of SEQ ID NO: 22 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 23 or a variant thereof, CDR-L2 of SEQ ID NO: 24 or a variant thereof, and CDR-L3 of SEQ ID NO: 16 or a variant thereof; or

[1267] (3b) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 36 or a variant thereof, CDR-H2 of SEQ ID NO: 37 or a variant thereof, and CDR-H3 of SEQ ID NO: 38 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 39 or a variant thereof, CDR-L2 of SEQ ID NO: 40 or a variant thereof, and CDR-L3 of SEQ ID NO: 32 or a variant thereof;

[1268] wherein the variant described in any one of (3a) and (3b) has at least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with the sequence from which it is derived, or the variant has one or more amino acid substitutions, deletions or additions (e.g., substitutions, deletions or additions of 1, 2 or 3 amino acids) compared to the sequence from which it is derived; preferably, the substitutions are conservative substitutions;

[1269] or,

[1270] (4) The following heavy chain variable region (VH) and / or light chain variable region (VL), wherein the CDRs are defined according to the AbM numbering system:

[1271] (4a) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 25 or a variant thereof, CDR-H2 of SEQ ID NO: 26 or a variant thereof, and CDR-H3 of SEQ ID NO: 13 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 14 or a variant thereof, CDR-L2 of SEQ ID NO: 15 or a variant thereof, and CDR-L3 of SEQ ID NO: 16 or a variant thereof; or

[1272] (4b) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 41 or a variant thereof, CDR-H2 of SEQ ID NO: 42 or a variant thereof, and CDR-H3 of SEQ ID NO: 29 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 30 or a variant thereof, CDR-L2 of SEQ ID NO: 31 or a variant thereof, and CDR-L3 of SEQ ID NO: 32 or a variant thereof;

[1273] Wherein, the variant described in any one of (4a) and (4b) has at least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity compared with the sequence from which it is derived, or the variant has one or more amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) compared with the sequence from which it is derived; preferably, the substitutions are conservative substitutions.

[1274] In another embodiment of the ADC, the antibody or antigen-binding fragment is selected from:

[1275] (a) a VH having the amino acid sequence shown in SEQ ID NO: 1 and / or a VL having the amino acid sequence shown in SEQ ID NO: 2;

[1276] (b) a VH having the amino acid sequence shown in SEQ ID NO: 3 and / or a VL having the amino acid sequence shown in SEQ ID NO: 4;

[1277] (c) a VH having the amino acid sequence shown in SEQ ID NO: 5 and / or a VL having the amino acid sequence shown in SEQ ID NO: 6;

[1278] (d) a VH having the amino acid sequence shown in SEQ ID NO: 7 and / or a VL having the amino acid sequence shown in SEQ ID NO: 8; and

[1279] (e) VH having the amino acid sequence shown in SEQ ID NO: 9 and / or VL having the amino acid sequence shown in SEQ ID NO: 10.

[1280] In another embodiment of the ADC, the antibody or antigen-binding fragment thereof is selected from:

[1281] (1) an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 1 and a VL having the amino acid sequence of SEQ ID NO: 2, for example, an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 1 and a CH having the amino acid sequence of SEQ ID NO: 43 or 45, a VL having the amino acid sequence of SEQ ID NO: 2, and a CL having the amino acid sequence of SEQ ID NO: 44;

[1282] (2) an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence set forth in SEQ ID NO: 3 and a VL having the amino acid sequence set forth in SEQ ID NO: 4, for example, an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence set forth in SEQ ID NO: 3 and a CH having the amino acid sequence set forth in SEQ ID NO: 43 or 45, a VL having the amino acid sequence set forth in SEQ ID NO: 4, and a CL having the amino acid sequence set forth in SEQ ID NO: 44;

[1283] (3) an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 5 and a VL having the amino acid sequence of SEQ ID NO: 6, for example, an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 5 and a CH having the amino acid sequence of SEQ ID NO: 43 or 45, a VL having the amino acid sequence of SEQ ID NO: 6, and a CL having the amino acid sequence of SEQ ID NO: 44;

[1284] (4) an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 7 and a VL having the amino acid sequence of SEQ ID NO: 8, for example, an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence of SEQ ID NO: 7 and a CH having the amino acid sequence of SEQ ID NO: 43 or 45, a VL having the amino acid sequence of SEQ ID NO: 8, and a CL having the amino acid sequence of SEQ ID NO: 44;

[1285] (5) An antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence shown in SEQ ID NO: 9 and a VL having the amino acid sequence shown in SEQ ID NO: 10, for example, an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence shown in SEQ ID NO: 9 and a CH having the amino acid sequence shown in SEQ ID NO: 43 or 45, a VL having the amino acid sequence shown in SEQ ID NO: 10, and a CL having the amino acid sequence shown in SEQ ID NO: 44.

[1286] In another embodiment of the ADC, the antibody or antigen-binding fragment thereof comprises: (1) an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence shown in SEQ ID NO: 1 and a VL having the amino acid sequence shown in SEQ ID NO: 2, for example, an antibody or antigen-binding fragment thereof comprising a VH having the amino acid sequence shown in SEQ ID NO: 1 and a CH having the amino acid sequence shown in SEQ ID NO: 43 or 45 and a VL having the amino acid sequence shown in SEQ ID NO: 2 and a CL having the amino acid sequence shown in SEQ ID NO: 44;

[1287] In another embodiment of the ADC, the antibody or antigen-binding fragment thereof comprises: (2) an antibody or antigen-binding fragment thereof comprising a VH having an amino acid sequence as shown in SEQ ID NO: 3 and a VL having an amino acid sequence as shown in SEQ ID NO: 4, for example, an antibody or antigen-binding fragment thereof comprising a VH having an amino acid sequence as shown in SEQ ID NO: 3 and a CH having an amino acid sequence as shown in SEQ ID NO: 43 or 45 and a VL having an amino acid sequence as shown in SEQ ID NO: 4 and a CL having an amino acid sequence as shown in SEQ ID NO: 44;

[1288] In another embodiment of the ADC, the antibody or antigen-binding fragment thereof comprises: (3) an antibody or antigen-binding fragment thereof comprising a VH having an amino acid sequence as shown in SEQ ID NO: 5 and a VL having an amino acid sequence as shown in SEQ ID NO: 6, for example, an antibody or antigen-binding fragment thereof comprising a VH having an amino acid sequence as shown in SEQ ID NO: 5 and a CH having an amino acid sequence as shown in SEQ ID NO: 43 or 45 and a VL having an amino acid sequence as shown in SEQ ID NO: 6 and a CL having an amino acid sequence as shown in SEQ ID NO: 44;

[1289] In another embodiment of the ADC, the antibody or antigen-binding fragment thereof comprises: (4) an antibody or antigen-binding fragment thereof comprising a VH having an amino acid sequence as shown in SEQ ID NO: 7 and a VL having an amino acid sequence as shown in SEQ ID NO: 8, for example, an antibody or antigen-binding fragment thereof comprising a VH having an amino acid sequence as shown in SEQ ID NO: 7 and a CH having an amino acid sequence as shown in SEQ ID NO: 43 or 45 and a VL having an amino acid sequence as shown in SEQ ID NO: 8 and a CL having an amino acid sequence as shown in SEQ ID NO: 44;

[1290] In another embodiment of the ADC, the antibody or antigen-binding fragment thereof comprises: (5) an antibody or antigen-binding fragment thereof comprising a VH having an amino acid sequence as shown in SEQ ID NO: 9 and a VL having an amino acid sequence as shown in SEQ ID NO: 10, for example, an antibody or antigen-binding fragment thereof comprising a VH having an amino acid sequence as shown in SEQ ID NO: 9 and a CH having an amino acid sequence as shown in SEQ ID NO: 43 or 45 and a VL having an amino acid sequence as shown in SEQ ID NO: 10 and a CL having an amino acid sequence as shown in SEQ ID NO: 44.

[1291] Antibody preparation

[1292] The antibodies described herein can be prepared by various methods known in the art, for example, by genetic engineering and recombinant techniques. For example, DNA molecules encoding the heavy and light chains of the antibodies described herein are obtained by chemical synthesis or PCR amplification. The resulting DNA molecules are inserted into expression vectors and then transfected into host cells. The transfected host cells are then cultured under specific conditions and express the antibodies of the present invention.

[1293] Coupling

[1294] In another aspect, the present application provides a method for conjugating the drug-linker described herein to the antibody described herein to prepare the antibody drug conjugate (ADC) described herein.

[1295] In certain embodiments, an antibody described herein is conjugated to a drug-linker described herein via a lysine in the antibody.

[1296] In certain embodiments, the antibodies described herein are conjugated to the drug-linkers described herein via cysteine ​​residues in the antibodies. In certain embodiments, the cysteine ​​residues are derived from reduced intrachain disulfide bonds in the antibodies. In certain embodiments, the cysteine ​​residues are derived from reduced interchain disulfide bonds in the antibodies.

[1297] In some embodiments, the antibody is conjugated to the drug-linker via reduced interchain disulfide bonds in the antibody. For example, an IgG1 antibody is composed of four polypeptide chains, two heavy chains comprising VH, CH1, and Fc (e.g., hinge, CH2, and CH3) domains, and two light chains comprising VL and CL domains, connected by interchain cysteine ​​disulfide bonds (-SS-) (e.g., two heavy chain-light chain interchain disulfide bonds and two hinge heavy chain-heavy chain interchain disulfide bonds). In certain embodiments, when these disulfide bonds are cleaved under reducing conditions, eight (8) reactive cysteine ​​thiol moieties are generated. In certain embodiments, each of the eight reactive cysteine ​​thiol moieties is a site of attachment for a drug-linker, such that a maximum of eight (x=8) drug-linkers can be attached to the reduced antibody. In certain embodiments, any one of the four disulfide bonds is cleaved under reducing conditions, resulting in two (2) reactive cysteine ​​thiol moieties. In further embodiments, each of the two reactive cysteine ​​thiol moieties is a site of attachment for a drug-linker, such that two (x=2) drug-linkers can be attached to the reduced antibody. In certain embodiments, any two of the four disulfide bonds are cleaved under reducing conditions, resulting in four (4) reactive cysteine ​​thiol moieties. In further embodiments, each of the four reactive cysteine ​​thiol moieties is a site of attachment for a drug-linker, such that four (x=4) drug-linkers can be attached to the reduced antibody. In certain embodiments, any three of the four disulfide bonds are cleaved under reducing conditions, resulting in six (6) reactive cysteine ​​thiol moieties. In further embodiments, each of the six reactive cysteine ​​thiol moieties is a site of attachment for a drug-linker, such that six (x=6) drug-linkers can be attached to the reduced antibody.

[1298] In some embodiments, the interchain disulfide bond is located between two cysteine ​​residues, which break under reducing conditions to produce two reactive cysteine ​​sulfhydryl moieties. In a further embodiment, the interchain disulfide bond in the antibody is located between the heavy chain and the light chain, for example, between C220 of the heavy chain according to EU numbering and C214 of the kappa light chain, or between C220 of the heavy chain according to EU numbering and C214 of the lambda light chain according to Kabat numbering. In addition, or alternatively, the interchain disulfide bond in the antibody is located between the two heavy chains, for example, between C226 and / or C229 of the first heavy chain according to EU numbering and C226 and / or C229 of the second heavy chain. In some embodiments, the cysteine ​​residue is located in the hinge region of the antibody. In some embodiments, according to EU numbering, the cysteine ​​residue is located at any one or more of positions 220, 226, or 229 in the heavy chain (also referred to herein as C220, C226, or C229, respectively). In some embodiments, the cysteine ​​residue is located at position 214 of the light chain (also referred to herein as C214, e.g., at position 214 of a kappa light chain according to EU and / or Kabat numbering, or at position 214 of a lambda light chain according to Kabat numbering) according to EU and / or Kabat numbering. In one embodiment, the cysteine ​​residue is located at position 220, 226, and 229 of the heavy chain according to EU and / or Kabat numbering, or at position 214 of a light chain according to EU or Kabat numbering. In one embodiment, the cysteine ​​residue is located at position 220, 226, and 229 of the heavy chain according to EU and / or Kabat numbering, or at position 214 of a kappa light chain according to EU and Kabat numbering. In one embodiment, the cysteine ​​residue is located at position 220, 226, and 229 of the heavy chain according to EU and Kabat numbering, or at position 214 of a lambda light chain according to Kabat numbering. In one embodiment, the cysteine ​​residue is located at any one or more of the following positions:

[1299] (i) any one, any two, any three, or all four of positions 220, 226, and 229 in the first heavy chain, according to EU numbering;

[1300] (ii) any one, any two, any three, or all four of positions 220, 226, and 229 in the second heavy chain, according to EU numbering;

[1301] (iii) position 214 in the first light chain according to Kabat numbering; and / or

[1302] (iv) Position 214 in the second light chain according to Kabat numbering.

[1303] As used herein, C220, C226, and C229 refer to amino acid residues of immunoglobulins identified according to EU numbering (cysteine, Cys, C). As will be appreciated by those skilled in the art, such numbering correspondingly represents amino acid residues of polypeptides that align with the defined amino acid residues of immunoglobulins, as shown in www.imgt.org / IMGTScientificChart / Numbering / Hu_IGHGnber.html.

[1304] As used herein, the cysteine ​​residue at position 214 of the kappa light chain refers to the amino acid residues of immunoglobulins identified according to Kabat numbering (cysteine, Cys, C). As will be appreciated by those skilled in the art, such numbering correspondingly represents amino acid residues of polypeptides that align with the defined amino acid residues of immunoglobulins, as shown in www.imgt.org / IMGTScientificChart / Numbering / Hu_IGKCnber.html.

[1305] As used herein, the cysteine ​​residue at position 214 of the lambda light chain refers to the amino acid residues of immunoglobulins identified according to Kabat numbering (cysteine, Cys, C). As will be appreciated by those skilled in the art, such numbering correspondingly represents amino acid residues of polypeptides that align with the defined amino acid residues of immunoglobulins, as shown at www.imgt.org / IMGTScientificChart / Numbering / Hu_IGLCnber.html.

[1306] In certain embodiments, the antibodies described herein comprise four interchain disulfide bonds in the hinge region that can be reduced, thereby breaking and revealing reactive sulfhydryl moieties that can be conjugated to maleimide moieties on drug-linkers, e.g., maleimide moieties on drug-linkers described herein.

[1307] In certain embodiments, the antibodies described herein comprise a lysine residue, wherein the reactive amine side chain of the lysine residue can be conjugated to a drug linker (eg, a maleimide moiety on a drug linker described herein).

[1308] In one embodiment, the present invention provides a method for preparing the ADC described herein, comprising the following steps:

[1309] a) providing a solution comprising an antibody;

[1310] b) contacting the solution of a) with a reducing agent;

[1311] c) contacting the solution of b) with a solution comprising a drug-linker or a salt thereof as described herein to prepare the ADC.

[1312] In one embodiment, the reducing agent is tris(2-carboxyethyl)phosphine (TCEP).

[1313] Composition

[1314] In another aspect, the present application provides a composition of an antibody drug conjugate (ADC) as described herein. Such a composition may comprise a plurality of ADCs as described herein, wherein each ADC comprises a drug-linker as described herein, wherein x is independently 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10. In other words, each antibody molecule in the composition can be coupled to 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 drug-linkers. Thus, the composition is characterized in that the "drug to antibody" ratio (DAR) is in the range of about 1 to about 10. Methods for determining DAR are well known to those skilled in the art, including methods using reverse phase chromatography or HPLC-MS.

[1315] For example, in any embodiment, the ADC compositions described herein have a DAR of about 1 to about 10, or any subrange therebetween, e.g., about 1 to 2, about 1 to 3, about 1 to 4, about 1 to 5, about 1 to 6, about 1 to 7, about 1 to 8, about 1 to 9, about 1 to 10, about 2 to 3, about 2 to 4, about 2 to 5, about 2 to 6, about 2 to 7, about 2 to 8, about 2 to 9, about 2 to 10, about 3 to 4, about 3 to 5, about 3 to 6, about 3 to 7, about 3 to 8, about 3 to 9, about 3 to 10, about 4 to 5, about 4 to 6, about 4 to 7, about 4 to 8, about 4 to 9, about 4 to 10, about 5 to 6, about 5 to 7, about 5 to 8, about 5 to 9, about 5 to 10, about 6 to 7, about 6 to 8, about 6 to 9, about 6 to 10, about 7 to 8, about 7 to 9, about 7 to 10, about 8 to 9, about 8 to 10, or about 9 to 10.

[1316] In certain embodiments, the DAR of the ADC compositions described herein is about 3 to 9, e.g., about 3.0 to 3.5, about 3.0 to 4.0, about 3.0 to 4.5, about 3.0 to 5.0, about 3.0 to 6.0, about 3.5 to 4.0, about 3.5 to 4.5, about 3.5 to 5.0, about 3.5 to 5.5, about 3.5 to 6.0, about 3.5 to 6.5, about 4.0 to 4.5, about 4.0 to 5.0, about 4.0 to 5.5, about 4.0 to 6.0, about 4.0 to 6.5, about 4.0 to 7.0, about 4.0 to 8. .0, about 4.5 to 5.0, about 4.5 to 5.5, about 4.5 to 6.0, about 4.5 to 6.5, about 4.5 to 7.0, about 4.5 to 7.5 about 5.0 to 8.0, about 5.5 to 6.0, about 5.5 to 6.5, about 5.5 to 7.0, about 5.5 to 7.5, about 5.5 to 8.0, about 6.0 to 6.5, about 6.0 to 7.0, about 6.0 to 7.5, about 6.0 to 8.5, about 6.5 to 7.0, about 6.5 to 7.5, about 6.5 to 8.5, about 7.0 to 7.5.

[1317] Pharmaceutical composition

[1318] In another aspect, the present application provides a pharmaceutical composition comprising the antibody-drug conjugate according to any one of the above embodiments and optionally the drug-linker according to any one of the above embodiments, and one or more pharmaceutically acceptable excipients.

[1319] In one embodiment, the pharmaceutical composition provided herein contains an antibody drug conjugate (ADC) and one or more pharmaceutically acceptable excipients, wherein the ADC is:

[1320]

[1321] HA is an antibody or an antigen-binding fragment thereof, which comprises a heavy chain (HC) having the amino acid sequence shown in SEQ ID NO: 46 and a light chain (LC) having the amino acid sequence shown in SEQ ID NO: 47; and x is 8 to 9.

[1322] In one embodiment, the pharmaceutical composition provided herein contains an antibody drug conjugate (ADC) and one or more pharmaceutically acceptable excipients, wherein the ADC is:

[1323]

[1324] wherein HA is an antibody or an antigen-binding fragment thereof comprising a heavy chain (HC) having the amino acid sequence shown in SEQ ID NO: 46 and a light chain (LC) having the amino acid sequence shown in SEQ ID NO: 47; and x is 8.

[1325] In one embodiment, the pharmaceutical composition provided herein contains an antibody drug conjugate (ADC) and one or more pharmaceutically acceptable excipients, wherein the ADC is:

[1326]

[1327] HA is an antibody or an antigen-binding fragment thereof, which comprises a heavy chain (HC) having the amino acid sequence shown in SEQ ID NO: 46 and a light chain (LC) having the amino acid sequence shown in SEQ ID NO: 47; and x is 7 to 8.

[1328] In one embodiment, the pharmaceutical composition provided herein contains an antibody drug conjugate (ADC) and one or more pharmaceutically acceptable excipients, wherein the ADC is:

[1329]

[1330] wherein HA is an antibody or an antigen-binding fragment thereof comprising a heavy chain (HC) having the amino acid sequence shown in SEQ ID NO: 46 and a light chain (LC) having the amino acid sequence shown in SEQ ID NO: 47; and x is 8.

[1331] In one embodiment, the pharmaceutical composition provided herein contains an antibody drug conjugate (ADC) and one or more pharmaceutically acceptable excipients, wherein the ADC is:

[1332]

[1333] wherein HA is an antibody or an antigen-binding fragment thereof comprising a heavy chain (HC) having the amino acid sequence shown in SEQ ID NO: 46 and a light chain (LC) having the amino acid sequence shown in SEQ ID NO: 47; and x is 7 to 8.

[1334] In one embodiment, the pharmaceutical composition provided herein contains an antibody drug conjugate (ADC) and one or more pharmaceutically acceptable excipients, wherein the ADC is:

[1335]

[1336] wherein HA is an antibody or an antigen-binding fragment thereof comprising a heavy chain (HC) having the amino acid sequence shown in SEQ ID NO: 46 and a light chain (LC) having the amino acid sequence shown in SEQ ID NO: 47; and x is 8.

[1337] In one embodiment, the pharmaceutical composition provided herein contains an antibody drug conjugate (ADC) and one or more pharmaceutically acceptable excipients, wherein the ADC is:

[1338] In one embodiment, the pharmaceutical composition provided herein contains an antibody drug conjugate (ADC) and one or more pharmaceutically acceptable excipients, wherein the ADC is:

[1339]

[1340] wherein HA is an antibody or an antigen-binding fragment thereof comprising a heavy chain (HC) having the amino acid sequence shown in SEQ ID NO: 46 and a light chain (LC) having the amino acid sequence shown in SEQ ID NO: 47; and x is 8 to 9.

[1341] In one embodiment, the pharmaceutical composition provided herein contains an antibody drug conjugate (ADC) and one or more pharmaceutically acceptable excipients, wherein the ADC is:

[1342]

[1343] wherein HA is an antibody or an antigen-binding fragment thereof comprising a heavy chain (HC) having the amino acid sequence shown in SEQ ID NO: 46 and a light chain (LC) having the amino acid sequence shown in SEQ ID NO: 47; and x is 8.

[1344] In one embodiment, the pharmaceutical composition provided herein contains an antibody drug conjugate (ADC) and one or more pharmaceutically acceptable excipients, wherein the ADC is:

[1345]

[1346] wherein HA is an antibody or an antigen-binding fragment thereof comprising a heavy chain (HC) having the amino acid sequence shown in SEQ ID NO: 46 and a light chain (LC) having the amino acid sequence shown in SEQ ID NO: 47; and x is 8 to 9.

[1347] Pharmaceutically acceptable excipients include, for example, pharmaceutically acceptable carriers and / or excipients. Pharmaceutically acceptable excipients further include salts and solvates.

[1348] The ADC described herein is generally formulated in a unit injectable form together with a pharmaceutically acceptable parenteral vehicle for parenteral use, such as bolus injection, intravenous injection, intratumoral injection, etc. Optionally, the antibody drug conjugate having the desired purity is mixed with a pharmaceutically acceptable diluent, carrier, excipient or stabilizer in the form of a lyophilized agent or solution (Remington's Pharmaceutical Sciences (1980) 16 th The antibody drug conjugates described herein or pharmaceutical compositions containing the same can be administered by any route appropriate to the subject to be treated.

[1349] The ADCs and pharmaceutical compositions described herein can be formulated into any dosage form known in the medical field, such as tablets, pills, suspensions, emulsions, solutions, gels, capsules, powders, granules, elixirs, lozenges, suppositories, injections (including injections, sterile powders for injection, and concentrated solutions for injection), inhalants, sprays, and the like. The preferred dosage form depends on the intended route of administration and therapeutic use. The pharmaceutical compositions of the present invention should be sterile and stable under production and storage conditions. The preferred dosage form is an injection. Such an injection can be a sterile injectable solution. For example, a sterile injectable solution can be prepared by the following method: the necessary dose of the antibody of the present invention is incorporated into a suitable solvent, and other required ingredients (including but not limited to pH regulators, surfactants, adjuvants, ionic strength enhancers, etc., penetrants, preservatives, diluents, or any combination thereof) are optionally added, followed by filtration sterilization. In addition, for ease of storage and use, the sterile injectable solution can be prepared as a sterile lyophilized powder (e.g., by vacuum drying or freeze drying). Such a sterile lyophilized powder can be dispersed in a suitable carrier, such as sterile, pyrogen-free water, before use.

[1350] In addition, the ADC described herein can be present in a pharmaceutical composition in a unit dosage form for administration by any suitable method known in the art, including but not limited to oral, oral, sublingual, ophthalmic, topical, parenteral, rectal, intrathecal, intracytoplasmic, inguinal, intravesical, topical (e.g., powder, ointment or drops) or intranasal routes. However, for many therapeutic uses, the preferred route / mode of administration is parenteral administration (e.g., intravenous, subcutaneous, intraperitoneal, intramuscular). The skilled artisan will appreciate that the route and / or mode of administration will vary depending on the intended purpose. In some preferred embodiments, the ADC and pharmaceutical compositions described herein are administered by intravenous infusion or injection.

[1351] In certain embodiments, the pharmaceutical composition may further comprise other pharmaceutically active agents. In certain embodiments, the other pharmaceutically active agents are drugs with anti-tumor activity. In certain embodiments, the other pharmaceutically active agents are selected from EGFR inhibitors, HER2 inhibitors, HER3 inhibitors, HER4 inhibitors, IGFR-1 inhibitors, mTOR inhibitors, PI3 kinase inhibitors, PTK7 inhibitors, c-met or VEGF inhibitors, chemotherapeutic agents, or any combination thereof. In certain embodiments, the ADC described herein and the other pharmaceutically active agents are provided as separate components or as mixed components. Therefore, the ADC described herein and the other pharmaceutically active agents may be administered simultaneously, separately, or sequentially.

[1352] How to use

[1353] The antibody drug conjugates described herein, the drug-linkers described herein, or pharmaceutical compositions thereof can be used to treat various diseases or conditions, such as cancers with high PTK7 expression, including solid tumors or hematological malignancies, such as lung cancer, breast cancer, epidermal cancer (e.g., cutaneous squamous cell carcinoma, oral squamous cell carcinoma), ovarian cancer, or esophageal cancer (e.g., esophageal squamous cell carcinoma).

[1354] Therefore, the present application provides use of an antibody-drug conjugate (ADC), a drug-linker, or a pharmaceutical composition comprising the same as described in any of the aforementioned embodiments in the preparation of a drug for treating cancers with high PTK7 expression.

[1355] At the same time, the present application also provides a method for treating PTK7-overexpressing cancer, which comprises the step of administering to a subject in need thereof a therapeutically effective amount of an antibody-drug conjugate (ADC), a drug-linker, or a pharmaceutical composition comprising the same as described in any one of the aforementioned embodiments.

[1356] In certain embodiments, the antibody drug conjugate (ADC), drug-linker, or pharmaceutical composition is sufficient (e.g., in a subject):

[1357] (1) Inhibit cell (such as tumor cell) proliferation;

[1358] (2) inhibit tumor growth;

[1359] (3) induce and / or increase antibody-dependent cellular cytotoxicity activity;

[1360] (4) inhibiting PTK7-mediated signal transduction;

[1361] (5) preventing and / or treating PTK7-mediated diseases / disorders; or

[1362] (6) Any combination of (1) to (5) above.

[1363] In certain embodiments, the PTK7-mediated disease / disorder is a tumor, e.g., a tumor expressing PTK7. In certain embodiments, the tumor is selected from lung cancer, breast cancer, epidermal cancer (e.g., cutaneous squamous cell carcinoma, oral squamous cell carcinoma), ovarian cancer, or esophageal cancer (e.g., esophageal squamous cell carcinoma), or any combination thereof. Example

[1364] The present invention will be further described below by describing specific embodiments, but this is not intended to limit the present invention. Those skilled in the art can make various modifications or improvements based on the teachings of the present invention without departing from the basic idea and scope of the present invention.

[1365] The information of the sequences involved in the present invention is described in the following table:

[1366]

[1367]

[1368]

[1369]

[1370]

[1371]

[1372]

[1373]

[1374]

[1375] The abbreviations used in this document have the following meanings:

[1376]

[1377] The structures of the compounds described in the following examples were determined by nuclear magnetic resonance ( 1 H NMR) or mass spectrometry (MS).

[1378] Nuclear magnetic resonance (NMR) 1 H NMR measurements were performed using a Bruker 400 MHz nuclear magnetic resonance instrument; the deuterated reagent was hexadeuterated dimethyl sulfoxide (DMSO-d6); and the internal standard was tetramethylsilane (TMS).

[1379] The abbreviations used in the nuclear magnetic resonance (NMR) spectra in the examples are shown below.

[1380] s: singlet, d: doublet, t: triplet, q: quartet, m: multiplet, br: broad, J: coupling constant, Hz: hertz, DMSO-d6: deuterated dimethyl sulfoxide. δ values ​​are expressed in ppm.

[1381] Mass spectrometry (MS) was performed using an Agilent (ESI) mass spectrometer, model Agilent 6120B.

[1382] Example 1 N-((S)-10-benzyl-1-((1S,9S)-9-ethyl-5-fluoro-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)amino)-1,6,9,12,15-pentaoxo-3-oxa-5,8,11,14-tetraazahexadec-16-yl-6-(2,5-dioxo-2,5-dihydro-1-H-pyrrol-1-yl)hexanamide (A-01)

[1383]

[1384] Compound A-01-1 (0.40 g, 640.59 μmol, synthesis reference CN 111936169A) and isotecan mesylate (0.37 g, 704.65 μmol) were dissolved in DMF (8 mL). HATU (0.32 g, 832.77 μmol) and DIPEA (0.25 g, 1.92 mmol) were added and reacted at 25°C for 4 hours. DIPEA was removed under reduced pressure, and the mixture was freeze-dried to remove most of the DMF to obtain a crude product. The crude product was purified by preparative HPLC (conditions as follows) to obtain 273 mg of the title compound.

[1385] Column: Waters XBridge Prep C18 OBD 45 mm × 450 mm × 8.0 μm

[1386] Mobile phase A: acetonitrile; Mobile phase B: water (0.05% trifluoroacetic acid)

[1387] Time [min] Mobile phase A[%] Mobile phase B[%] Flow rate [mL / min] 0.00 30 70 70 5.00 30 70 70 60.00 70 30 70

[1388] The structural characterization data are as follows:

[1389] ESI-MS (m / z): 1034.4 [M+H] + .

[1390] Example 2 Synthesis of N-((1S,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)-2-hydroxyacetamide and N-((1R,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)-2-hydroxyacetamide (1-8-A and 1-8-B)

[1391]

[1392] Step 1: Synthesis of 1-chloro-3-bromo-2-methyl-5-nitrobenzene (1-8-2)

[1393] Compound 1-8-1 (5.00 g, 29.14 mmol) was dissolved in n-heptane (25 mL) at 25°C; concentrated sulfuric acid (25 mL) was added and the mixture was heated to 50°C. NBS (6.22 g, 34.97 mmol) was added portionwise at 50°C and the reaction was maintained at 50°C for 2 hours. The reaction was monitored by thin-layer chromatography (ethyl acetate:petroleum ether = 1:10). The reaction solution was cooled to room temperature and then added dropwise to ice water. The mixture was extracted with toluene. The organic phases were combined, washed with sodium sulfite solution, water, and saturated brine, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was purified by preparative HPLC (conditions as follows). The preparative solution was freeze-dried to obtain 4.88 g of the title compound.

[1394] Chromatographic column: C18 ODS 45mm×450mm×8.0μm

[1395] Mobile phase A: acetonitrile; Mobile phase B: water (0.05% formic acid)

[1396] Time [min] Mobile phase A[%] Mobile phase B[%] Flow rate [mL / min] 0 60 40 60 10 60 40 60 40 100 0 60

[1397] Step 2: Synthesis of 3-chloro-5-bromo-4-methylaniline (1-8-3)

[1398] Compound 1-8-2 (4.88 g, 19.48 mmol) was dissolved in ethyl acetate (100 mL) at 25°C, and platinum on carbon (2.00 g, 19.48 mmol, 5% content) was added. After hydrogen replacement, the reaction was continued at 60°C under hydrogen protection for 4 hours. The reaction was monitored by HPLC-MS / MS. The reaction solution was filtered and concentrated to obtain 3.68 g of the crude title compound, which was used directly in the next reaction without further purification.

[1399] Step 3: Synthesis of N-(3-chloro-5-bromo-4-methylphenyl)acetamide (1-8-4)

[1400] Compound 1-8-3 (3.63 g, 14.82 mmol) was dissolved in ethyl acetate (70 mL) at 20°C. Triethylamine (4.50 g, 44.45 mmol) and acetic anhydride (2.27 g, 22.23 mmol) were added and the mixture was maintained at 20°C for 20 h. The reaction was monitored by HPLC-MS / MS. Water was added to the reaction solution, followed by extraction with ethyl acetate. The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated under reduced pressure to obtain a crude product. The crude product was then slurried in a mixture of ethyl acetate and petroleum ether (1:5) to afford 2.86 g of the title compound.

[1401] Step 4: Synthesis of 4-(5-acetamido-3-chloro-2-methylphenyl)but-3-enoic acid (1-8-5)

[1402] Compound 1-8-4 (1.80 g, 6.86 mmol) was dissolved in THF (20 mL) and water (5 mL) at 20°C. Vinyl acetic acid (708.31 mg, 8.23 ​​mmol), DIPEA (1.95 g, 15.08 mmol), and tris(o-methylphenyl)phosphine (62.60 mg, 0.20 mmol) were added. The reaction system was purged with nitrogen and heated to 70°C for 5 h. The reaction was monitored by HPLC-MS / MS. 1N sodium hydroxide solution was added to the reaction mixture to adjust the pH to 8. Ethyl acetate was added for extraction. The aqueous phase was adjusted to pH 3 with 1N hydrochloric acid and then extracted with ethyl acetate. The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated under reduced pressure to obtain 0.82 g of the title compound, which was used directly in the next step.

[1403] Step 5: Synthesis of 4-(5-acetamido-3-chloro-2-methylphenyl)butyric acid (1-8-6)

[1404] Compound 1-8-5 (2.60 g, 9.71 mmol) was dissolved in THF (50 mL) at 20°C. Pd / C (0.52 g, 10%) was added. The system was placed under positive pressure using a hydrogen balloon and reacted at 40°C for 2 h. The reaction was monitored by HPLC-MS / MS. The reaction solution was filtered and the filtrate was concentrated to obtain 2.43 g of the title compound, which was used in the next step without further purification.

[1405] Step 6: Synthesis of N-(3-chloro-4-methyl-8-oxo-5,6,7,8-tetrahydronaphthalen-1-yl)acetamide (1-8-7)

[1406] Compound 1-8-6 (2.43 g, 9.01 mmol) was dissolved in trifluoroacetic acid (10 mL) and cooled to 5°C. Trifluoroacetic anhydride (3.78 g, 18.02 mmol, 2.50 mL) was added dropwise. The reaction was maintained at 5°C for 4 h. The reaction was monitored by HPLC-MS / MS. The reaction solution was added to water and adjusted to pH 9 with 10N sodium hydroxide. Ethyl acetate was added for extraction. The organic phases were combined, dried over anhydrous sodium sulfate, concentrated under reduced pressure, and purified on a silica gel column (ethyl acetate:petroleum ether = 0-20%) to obtain 1.53 g of the title compound.

[1407] Step 7: Synthesis of (Z)-N-(3-chloro-7-(hydroxyimino)-4-methyl-8-oxo-5,6,7,8-tetrahydronaphthalen-1-yl)acetamide (1-8-8)

[1408] Potassium tert-butoxide (1.50 g, 13.37 mmol) was dissolved in THF (16 mL) and tert-butanol (4 mL) at 5°C; a THF solution (16 mL) of compound 1-8-7 (1.53 g, 6.08 mmol) was added dropwise. After 10 minutes, amyl nitrite (1.14 g, 9.73 mmol) was added dropwise. The reaction was maintained at 5°C for 1 hour and monitored by HPLC-MS / MS. The reaction solution was adjusted to pH 5 with 1N hydrochloric acid and extracted with ethyl acetate. The combined organic phases were dried over anhydrous sodium sulfate and concentrated under reduced pressure. The crude product was slurried with methyl tert-butyl ether to obtain 1.20 g of the title compound.

[1409] Step 8: Synthesis of N-(7-amino-3-chloro-4-methyl-8-oxo-5,6,7,8-tetrahydronaphthalen-1-yl)acetamide (1-8-9)

[1410] Compound 1-8-8 (0.50 g, 1.78 mmol) was dissolved in methanol (8 mL) and 2N hydrochloric acid (8 mL) at 20°C. Pd / C (0.15 g, 10%) was added. The system was placed under positive pressure using a hydrogen balloon and reacted at 5°C for 2 h. The reaction was monitored by HPLC-MS / MS. The reaction solution was filtered and concentrated to obtain 0.52 g of the title compound, which was used in the next step without further purification.

[1411] Step 9: Synthesis of N,N'-(3-chloro-4-methyl-8-oxo-5,6,7,8-tetrahydronaphthalene-1,7-diyl)diethylamide (1-8-10)

[1412] Compound 1-8-9 (0.52 g, 1.70 mmol) was dissolved in pyridine (5 mL) at 20°C; acetic anhydride (2 mL) was added; the reaction was maintained at 20°C for 2 h, and the reaction was monitored by HPLC-MS / MS. The reaction solution was added to water and extracted with ethyl acetate. The organic phases were washed with water, combined, dried over anhydrous sodium sulfate, concentrated under reduced pressure, and purified on a silica gel column (ethyl acetate:petroleum ether = 0-30%) to obtain 0.22 g of the title compound.

[1413] Step 10: Synthesis of N-(8-amino-6-chloro-5-methyl-1-oxo-1,2,3,4-tetrahydronaphthalen-2-yl)acetamide (1-8-11)

[1414] Compound 1-8-10 (450.97 mg, 1.46 mmol) was dissolved in methanol (16 mL) at 20°C; 2N hydrochloric acid (16 mL) was added, and the mixture was heated to 60°C for 2 h. The reaction was monitored by HPLC-MS / MS. Saturated sodium bicarbonate solution was added to the cooled reaction mixture to adjust the pH to 8; ethyl acetate was added for extraction; the organic phases were combined, dried over anhydrous sodium sulfate, and concentrated under reduced pressure to obtain 230.00 mg of the title compound, which was used in the next step without further purification.

[1415] Step 11: Synthesis of N-((9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzopyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)acetamide (1-8-12)

[1416] Compound 1-8-11 (230.00 mg, 0.78 mmol) was dissolved in toluene (10 mL); (S)-4-ethyl-4-hydroxy-7,8-dihydro-1H-pyrano[3,4-f]indolizine-3,6,10(4H)-trione (230.00 mg, 0.87 mmol) and p-toluenesulfonic acid (26.73 mg, 0.16 mmol) were added, and the mixture was heated to 140°C for 5 h. The reaction was monitored by HPLC-MS / MS. The reaction solution was concentrated, and the crude product was purified on a silica gel column (methanol:dichloromethane = 0-10%) to obtain 150.00 mg of the title compound.

[1417] Step 12: Synthesis of (9S)-1-amino-5-chloro-9-ethyl-9-hydroxy-4-methyl-1,2,3,9,12,15-hexahydro-10H,13H-benzopyrano[3',4':6,7]indolizino[1,2-b]quinoline-10,13-dione (1-2)

[1418] Compound 1-8-12 (40.00 mg, 0.081 mmol) was added to concentrated hydrochloric acid (1 mL) and heated to 100°C for 5 h. The reaction was monitored by HPLC-MS / MS. The reaction mixture was filtered, and the filtrate was purified by preparative HPLC (conditions described below). The preparative solution was freeze-dried to obtain the title compound 1-2 hydrochloride. Compound 1-2 hydrochloride was separated under the following purification conditions to yield two isomers, designated 1-2-A (trifluoroacetate salt 5.00 mg, retention time 9.85 min) and 1-2-B (trifluoroacetate salt 7.00 mg, retention time 10.62 min) based on their retention times.

[1419] Chromatographic column: SunFire Prep C18 OBD 19mm×150mm×5.0μm

[1420] Mobile phase A: acetonitrile; Mobile phase B: water (0.05% trifluoroacetic acid)

[1421] Time [min] Mobile phase A[%] Mobile phase B[%] Flow rate [mL / min] 0 5 95 28 2 5 95 28 18 50 50 28

[1422] The structural characterization data are as follows:

[1423] 1-2-A:

[1424] 1 H NMR(400MHz,DMSO-d6)δ8.42(s,3H),8.27(s,1H),7.36(s,1H),6.59(s,1H),5.78-5.63(m,1H),5.50-5.36(m,3H) ,5.10-5.06(m,1H),3.20-3.04(m,2H),2.56(s,3H),2.26-2.13(m,2H),1.93-1.79(m,2H),0.88(t,J=7.2Hz,3H).

[1425] ESI-MS (m / z): 452.1 [M+H] + .

[1426] 1-2-B:

[1427] 1 H NMR(400MHz,DMSO-d6)δ8.42(s,3H),8.27(s,1H),7.36(s,1H),6.58(s,1H),5.78-5.63(m,1H),5.50-5.36(m,3H) ,5.10-5.06(m,1H),3.20-3.04(m,2H),2.55(s,3H),2.26-2.13(m,2H),1.93-1.79(m,2H),0.88(t,J=7.2Hz,3H).

[1428] ESI-MS (m / z): 452.0 [M+H] + .

[1429] Step 13: 2-((tert-butyldiphenylsilyl)oxy)-N-((1S,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)acetamide and 2-((tert-butyldiphenylsilyl)oxy)-N-((1S,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)acetamide Synthesis of (1-8-13-A and 1-8-13-B)-N-((1R,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)acetamide

[1430] At 25°C, the hydrochloride of compound 1-2 (40.00 mg, 81.91 μmol) was dissolved in N,N-dimethylformamide (1 mL); 2-((tert-butyldiphenylsilyl)oxy)acetic acid (30.91 mg, 98.29 μmol), HATU (62.25 mg, 163.81 μmol) and N,N-diisopropylethylamine (42.34 mg, 327.63 μmol) were added sequentially; the reaction was maintained at 25°C for 0.5 hour and the reaction was monitored by HPLC-MS / MS. After the reaction was completed, water was added to the reaction solution; the mixture was extracted with dichloromethane / methanol (v / v=10 / 1); the organic phases were combined, dried over anhydrous sodium sulfate, and concentrated under reduced pressure; the crude product was purified by preparative thin-layer chromatography (dichloromethane:methanol=20:1) to separate and obtain two isomers; the two isomers were named 1-8-13-A (15.00 mg, Rf value 0.3) and 1-8-13-B (12.00 mg, Rf value 0.35) based on their Rf values.

[1431] Step 14: Synthesis of N-((1S,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)-2-hydroxyacetamide and N-((1R,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-1-yl)-2-hydroxyacetamide (1-8-A and 1-8-B)

[1432] At 25°C, 1-8-13-A (15.00 mg) and 1-8-13-B (12.00 mg) were dissolved in tetrahydrofuran (1 mL) in two reaction flasks; a mixture of tetrabutylammonium fluoride (1 M tetrahydrofuran solution) and glacial acetic acid (v / v = 13 / 1) (50 μL) was added dropwise; the reaction was maintained at 25°C for 0.5 hours, and the reaction was monitored by HPLC-MS / MS. After completion of the reaction, the reaction solutions were purified by preparative HPLC, and the prepared solutions were lyophilized to obtain the corresponding title compounds 1-8-A (i.e., 1-10) (6.94 mg) and 1-8-B (i.e., 1-9) (4.00 mg).

[1433] Chromatographic column: SunFire Prep C18 OBD 19mm×150mm×5.0μm

[1434] Mobile phase A: acetonitrile; Mobile phase B: water (0.05% formic acid)

[1435] Time [min] Mobile phase A [%] Mobile phase B [%] Flow rate [mL / min]

[1436] Time [min] Mobile phase A[%] Mobile phase B[%] Flow rate [mL / min] 0 20 80 28 3 20 80 28 18 90 10 28

[1437] The structural characterization data of 1-8-A (i.e. 1-10) are as follows:

[1438] 1H NMR (400MHz, DMSO-d6) δ8.43(d,J=8.8Hz,1H),8.16(s,1H),7.31(s,1H),6.55(s,1H),5.65-5.36(m,4H),5.21(q,J=19.0H z,2H),3.95(d,J=5.7Hz,2H),3.26-3.11(m,2H),2.53(s,3H),2.30-2.08(m,2H),1.94-1.79(m,2H),0.87(t,J=7.3Hz,3H).

[1439] ESI-MS (m / z): 510.1 [M+H] + .

[1440] The structural characterization data of 1-8-B (i.e. 1-9) are as follows:

[1441] 1H NMR (400MHz, DMSO-d6) δ8.45(d,J=8.9Hz,1H),8.15(s,1H),7.31(s,1H),6.54(s,1H),5.64-5.35(m,4H),5.19(q,J=19.0H z,2H),3.97(d,J=5.2Hz,2H),3.27-3.10(m,2H),2.51(s,3H),2.27-2.10(m,2H),1.93-1.80(m,2H),0.88(t,J=7.3Hz,3H).

[1442] ESI-MS (m / z): 510.1 [M+H] + .

[1443] Example 3 N-((10S)-10-benzyl-1-(((1S,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4';6,7]indolizino[1,2-b]quinolin-1-yl)amino)-1,6,9,12,15-pentaoxo-3-oxa-5,8,11,14-tetraazahexadec-16-yl)-6-(2-(methylsulfonyl)pyrimidin-5-yl)-hex-5-ynamide and N-((10S)-10 -Benzyl-1-(((1R,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4';6,7]indolizino[1,2-b]quinolin-1-yl)amino)-1,6,9,12,15-pentaoxo-3-oxa-5,8,11,14-tetraazahexadec-16-yl)-6-(2-(methylsulfonyl)pyrimidin-5-yl)-hex-5-ynamide (A-07-A (A-05) and A-07-B (A-06))

[1444]

[1445] Step 1: Synthesis of (S)-10-benzyl-23-(2-(methylsulfonyl)pyrimidin-5-yl)-6,9,12,15,18-pentaoxo-3-oxa-5,8,11,14,17-pentaazatricosane-22-ynecarboxylic acid (A-07-3)

[1446] Compound A-07-2 (30.00 mg, 0.07 mmol) was dissolved in DMF (0.2 mL) at 25°C. 2,5-Dioxopyrrolidin-1-yl-6-(2-(methylsulfonyl)pyrimidin-5-yl)hexyl-5-ynoate (A-07-1, 28.00 mg, 0.08 mmol) was added and reacted at 30°C for 1 h. The reaction was monitored by HPLC-MS / MS. The reaction solution was directly purified by preparative HPLC (conditions as follows) and the preparative solution was freeze-dried to obtain 20.00 mg of the title compound.

[1447] Chromatographic column: SunFire Prep C18 OBD 19mm×150mm×5.0μm

[1448] Mobile phase A: acetonitrile; Mobile phase B: water (0.05% formic acid)

[1449]

[1450] The structural characterization data are as follows:

[1451] ESI-MS (m / z): 691.0 [M+H2O] + .

[1452] Step 2: N-((10S)-10-benzyl-1-(((1S,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4';6,7]indolizino[1,2-b]quinolin-1-yl)amino)-1,6,9,12,15-pentaoxo-3-oxa-5,8,11,14-tetraazahexadec-16-yl)-6-(2-(methylsulfonyl)pyrimidin-5-yl)-hexane-5-amide and N-((10 Synthesis of S)-10-benzyl-1-(((1R,9S)-5-chloro-9-ethyl-9-hydroxy-4-methyl-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4';6,7]indolizino[1,2-b]quinolin-1-yl)amino)-1,6,9,12,15-pentaoxo-3-oxa-5,8,11,14-tetraazahexadec-16-yl)-6-(2-(methylsulfonyl)pyrimidin-5-yl)-hexan-5-amide (A-07-A and A-07-B)

[1453] At 25°C, the hydrochloride salt of 1-2 (30.00 mg, 61.43 μmol) was dissolved in N,N-dimethylformamide (1 mL). A-07-3 (49.66 mg, 73.72 μmol), HATU (35.01 mg, 92.14 μmol), and N,N-diisopropylethylamine (23.82 mg, 184.29 μmol) were added sequentially. The reaction was maintained at 25°C for 0.5 hours and monitored by HPLC-MS / MS. After completion, the reaction solution was purified by preparative HPLC (conditions listed below) and freeze-dried to obtain the title compound A-07. A-07 was separated under the following purification conditions to yield two isomers, designated A-07-A (i.e., A-05) (11.04 mg, retention time 7.5 min) and A-07-B (i.e., A-06) (19.42 mg, retention time 8.0 min) based on their retention times.

[1454] Chromatographic column: SunFire Prep C18 OBD 19mm×150mm×5.0μm

[1455] Mobile phase A: acetonitrile; Mobile phase B: water (0.05% formic acid)

[1456] Time [min] Mobile phase A [%] Mobile phase B[%] Flow rate [mL / min] 0 30 70 28 3 30 70 28 18 90 10 28

[1457] The structural characterization data are as follows:

[1458] A-07-A (i.e. A-05):

[1459] ESI-MS (m / z): 1107.3 [M+H] + .

[1460] 1H NMR (400MHz, DMSO) δ9.10 (s, 2H), 8.66-8.63 (m, 1H), 8.51 (d, J = 8.8Hz, 1H), 8.34-8.31 (m, 1H), 8.21-8.19 (m, 1H), 8.17-8.09 (m, 2H), 8.08-8.04(m,1H),7.30(s,1H),7.26-7.15(m,5H),6.55(s,1H),5.56-5.55(m,1H),5.48-5.35(m,2H),5.25-5.10(m,2H),4.64(d,J=6 .4Hz,2H),4.45-4.44(m,1H),4.06-3.98(m,2H),3.77-3.52(m,6H),3.41(s,3H),3.25-3.12(m,2H),3.03-3.00(m,1H),2.83-2.72(m, 1H),2.58-2.56(m,2H),2.48(s,3H),2.33-2.30(m,2H),2.21-2.13(m,2H),1.91-1.76(m,4H),0.87(t,J=7.2Hz,3H).A-07-B (i.e. A-06):

[1461] ESI-MS (m / z): 1107.3 [M+H] + .

[1462] Example 4 (S)-7-ethyl-7-hydroxy-14-(2-(isopropylamino)ethyl)-10,13-dihydro-11H-[1,3]dioxolano[4,5-g]pyrano[3',4':6,7]indolizino[1,2-b]quinoline-8,11(7H)-dione (2-2)

[1463]

[1464] Step 1: Synthesis of 3-(isopropylamino)-1-(6-nitrobenzo[d][1,3]dioxin-5-yl)propan-1-one (2-2-2)

[1465] Compound 2-2-1 (1.90 g, 8.08 mmol) was slowly added to nitric acid (8 mL) at 0°C and the temperature was slowly raised to 25°C for 1 hour. The reaction solution was directly purified by reverse phase column chromatography (acetonitrile / 0.5% formic acid aqueous solution) to obtain 1.50 g of the formate salt of the title compound.

[1466] The structural characterization data are as follows:

[1467] ESI-MS (m / z): 281.1 [M+H] + .

[1468] Step 2: Synthesis of 1-(6-aminobenzo[d][1,3]dioxin-5-yl)-3-(isopropylamino)propan-1-one (2-2-3)

[1469] The formate salt of compound 2-2-2 (1.25 g, 4.46 mmol) was added to tetrahydrofuran (20 mL); 10% palladium on carbon (125.00 mg) was added, and the atmosphere was replaced with hydrogen three times. The reaction was then allowed to react at 25°C for 16 hours. The reaction solution was filtered through celite, and the filtrate was concentrated under reduced pressure to dryness to obtain 895.00 mg of the crude title compound, which was used directly in the next reaction without purification.

[1470] The structural characterization data are as follows:

[1471] ESI-MS (m / z): 251.1 [M+H] + .

[1472] Step 3: Synthesis of ((S)-7-ethyl-7-hydroxy-14-(2-(isopropylamino)ethyl)-10,13-dihydro-11H-[1,3]dioxolano[4,5-g]pyrano[3',4':6,7]indolizino[1,2-b]quinoline-8,11(7H)-dione (2-2)

[1473] Compound 2-2-3 (23.00 mg, 0.09 mmol) and (4S)-4-ethyl-4-hydroxy-7,8-dihydro-1H-pyrano[3,4-f]indolizine-3,6,10(4H)-trione (21.00 mg, 0.09 mmol) were dissolved in toluene (4 mL), p-toluenesulfonic acid (1.40 mg, 0.009 mmol) was added, and the mixture was reacted at 140°C for 12 hours. The solvent was evaporated under reduced pressure to obtain a crude product of the title compound. The crude product was purified by HPLC (mobile phase A: acetonitrile, mobile phase B: 0.05% formic acid in water). Three drops of 3M hydrochloric acid were added to the prepared solution, followed by freeze-drying to obtain 8.70 mg of the hydrochloride salt of the title compound.

[1474] The structural characterization data are as follows:

[1475] ESI-MS (m / z): 478.2 [M+H] + .

[1476] 1H-NMR (400MHz, DMSO-d6): δ9.38(brs,2H),7.85(s,1H),7.52(s,1H),7.25(s,1H),6.30(d,J=2.0Hz,2H),5.43(s,2H),5.30(s, 2H),3.57-3.45(m,2H),3.40-3.25(m,1H),3.22-3.06(m,2H),1.95-1.77(m,2H),1.27(d,J=6.4Hz,6H),0.87(t,J=7.6Hz,3H).

[1477] Example 5 Synthesis of 4-((S)-2-(4-aminobutyl)-35-(4-((6-(2-(methylsulfonyl)pyrimidin-5-yl)hex-5-ynamido)methyl)-1H-1,2,3-triazol-1-yl)-4,8-dioxo-6,12,15,18,21,24,27,30,33-nonaoxa-3,9-diazapentatriacontamido)benzyl((S)-4-ethyl-11-(2-(N-isopropylmethylsulfonamido)ethyl)-3,14-dioxo-3,4,12,14-tetrahydro-1H-pyrano[3',4':6,7]indolizino[1,2-b]quinolin-4-yl)carbonate (B-01)

[1478]

[1479] Step 1: Synthesis of (S)-4-ethyl-11-(2-(N-isopropylmethylsulfonylamino)ethyl)-3,14-dioxo-3,4,12,14-tetrahydro-1H-pyrano[3',4':6,7]indolizino[1,2-b]quinolin-4-yl(4-((S)-2-(4-(((4-methoxyphenyl)diphenylmethyl)amino)butyl)-35-(4-((6-(2-(methylsulfonyl)pyrimidin-5-yl)hex-5-ynamido)methyl)-1H-1,2,3-triazol-1-yl)-4,8-dioxo-6,12,15,18,21,24,27,30,33-nonaoxy-3,9-diazapentatriacontamido)benzyl)carbonate (B-01-2)

[1480] At room temperature, compound B-01-1 (413.40 mg, 0.251 mmol, its synthesis reference patent CN111295389B) was dissolved in dimethyl sulfoxide and water (2.0 mL: 0.5 mL); cuprous bromide (72.95 mg, 0.503 mmol) and 6-(2-(methylsulfonyl)pyrimidin-5-yl)-N-(prop-2-yn-1-yl)-hex-5-ynamide (95.10 mg, 0.302 mmol) were added, the reaction was stirred for 1 hour and then filtered; the filtrate was purified by preparative high performance liquid chromatography (conditions as follows) to obtain 30.00 mg of the title compound.

[1481] Chromatographic column: SunFire Prep C18 OBD 19mm×150mm×5.0μm

[1482] Mobile phase A: acetonitrile; mobile phase B: water

[1483] Time [min] Mobile phase A[%] Mobile phase B[%] Flow rate [mL / min] 0.00 40 60 28 8.00 40 60 28 50.00 90 10 28

[1484] The structural characterization data are as follows:

[1485] ESI-MS(m / z):815.9[(M-273) / 2+H] + .

[1486] Step 2: Synthesis of 4-((S)-2-(4-aminobutyl)-35-(4-((6-(2-(methylsulfonyl)pyrimidin-5-yl)hex-5-ynamido)methyl)-1H-1,2,3-triazol-1-yl)-4,8-dioxo-6,12,15,18,21,24,27,30,33-nonaoxa-3,9-diazapentatriacontamido)benzyl((S)-4-ethyl-11-(2-(N-isopropylmethylsulfonamido)ethyl)-3,14-dioxo-3,4,12,14-tetrahydro-1H-pyrano[3',4':6,7]indolizino[1,2-b]quinolin-4-yl)carbonate (B-01)

[1487] Dissolve compound B-01-3 (30.00 mg, 0.02 mmol) in dichloromethane (1.0 mL); add trifluoroacetic acid (0.2 mL) to the reaction mixture and react at room temperature for 30 min. The reaction mixture is concentrated under reduced pressure and purified by preparative HPLC (conditions as follows) to obtain 20.00 mg of the trifluoroacetate salt of the title compound.

[1488] Chromatographic column: SunFire Prep C18 OBD 19mm×150mm×5.0μm

[1489] Mobile phase A: acetonitrile; Mobile phase B: water (0.05% trifluoroacetic acid)

[1490] Time [min] Mobile phase A[%] Mobile phase B[%] Flow rate [mL / min] 0.00 15 85 28 8.00 15 85 28 50.00 60 40 28

[1491] The structural characterization data are as follows: ESI-MS (m / z): 508.2 [M+H] + .

[1492] Example 6 (2S,3S,4S,5R,6S)-6-(4-((((2-((S)-7-ethyl-7-hydroxy-8,11-dioxo-7,8,11,13-tetrahydro-10H-[1,3]dioxolano[4,5-g]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-14-yl)ethyl)(isopropyl)carbamoyl)oxy)methyl)-2-(2-(2-(2-(6-(2-(methylsulfonyl)pyrimidin-5-yl)hex-5-ynamido)ethoxy)ethoxy)acetamido)phenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid (B-03)

[1493]

[1494] Step 1: Synthesis of (2S,3R,4S,5S,6S)-2-(4-(hydroxymethyl)-2-nitrophenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triacetate (B-03-3)

[1495] Dissolve (2R,3R,4S,5S,6S)-2-bromo-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triacetic acid triester (B-03-1, 12.32 g, 31.02 mmol) and 4-hydroxy-3-nitrobenzyl alcohol (B-03-2, 5.00 g, 29.56 mmol) in acetonitrile (200 mL). Add silver oxide (27.40 g, 118.25 mmol) with stirring. After nitrogen substitution, allow the mixture to react at room temperature in the dark for 12 hours. Monitor the reaction using HPLC-MS / MS. Filter the reaction mixture through celite, concentrate the filtrate under reduced pressure, and purify it via silica gel column chromatography (petroleum ether:ethyl acetate = 1:3) to obtain 12.80 g of the title compound.

[1496] The structural characterization data are as follows:

[1497] ESI-MS (m / z): 503 [M+18] + .

[1498] Step 2: Synthesis of (2S,3R,4S,5S,6S)-2-(2-amino-4-(hydroxymethyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triacetate (B-03-4)

[1499] Compound B-03-3 (2.20 g, 4.53 mmol) was dissolved in ethyl acetate and tetrahydrofuran (50 mL each); PtO2 (0.20 g) was added. The reaction system was then purged with a hydrogen balloon three times and allowed to react under a hydrogen atmosphere for 2 hours. The reaction was monitored by HPLC-MS / MS. The reaction mixture was filtered, the filter cake was rinsed with ethyl acetate, and the filtrate was evaporated to dryness under reduced pressure to obtain 2.02 g of the crude title compound, which was used directly in the next reaction.

[1500] The structural characterization data are as follows:

[1501] ESI-MS (m / z): 456.1 [M+1] + .

[1502] Step 3: Synthesis of (2S,3R,4S,5S,6S)-2-(2-(1-(9H-fluoren-9-yl)-3-oxo-2,7,10-trioxa-4-azadodecane-12-amino)-4-(hydroxymethyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triacetate (B-03-5)

[1503] Compound B-03-4 (456.00 mg, 1.00 mmol) and [2-[2-(Fmoc-amino)ethoxy]ethoxy]acetic acid (385.91 mg, 1.00 mmol) were dissolved in dichloromethane (10 mL). 2-Ethoxy-1-ethoxycarbonyl-1,2-dihydroquinoline (495.22 mg, 2.00 mmol) was added with stirring. The mixture was stirred for 2 hours. The reaction was monitored by HPLC-MS / MS. The reaction solution was concentrated under reduced pressure and purified by silica gel column chromatography (methanol:dichloromethane = 1:20) to obtain 507.00 mg of the title compound.

[1504] The structural characterization data are as follows:

[1505] ESI-MS (m / z): 823.3 [M+1] + .

[1506] Step 4: Synthesis of (2S,3R,4S,5S,6S)-2-(2-(1-(9H-fluoren-9-yl)-3,11-dioxo-2,7,10-trioxa-4,12-diazadodecyl)-4-((((4-nitrophenoxy)carbonyl)oxy)methyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triacetate (B-03-6)

[1507] Compound B-03-5 (507.00 mg, 616.18 μmol) and diisopropylethylamine (238.91 mg, 1.85 mmol) were dissolved in dichloromethane (20 mL). p-Nitrophenyl chloroformate (372.60 mg, 1.85 mmol) was then dissolved in dichloromethane (1 mL) and slowly added dropwise to the reaction mixture. After addition, the mixture was allowed to react at room temperature for 15 h. The reaction was monitored by HPLC-MS / MS. The reaction mixture was concentrated under reduced pressure and purified by silica gel column chromatography (methanol:dichloromethane = 1:20) to obtain 496.00 mg of the title compound.

[1508] The structural characterization data are as follows:

[1509] ESI-MS (m / z): 988.5 [M+1] + .

[1510] Step 5: Synthesis of (2S,3R,4S,5S,6S)-2-(2-(1-(9H-fluoren-9-yl)-3-oxo-2,7,10-trioxa-4-azadodecanamido-12-yl)-4-((((2-((S)-7-ethyl-7-hydroxy-8,11-dioxo-7,8,11,13-tetrahydro-10H-[1,3]dioxolano[4,5-g]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-14-yl)ethyl)(isopropyl)carbamoyl)oxy)methyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-triacetate (B-03-7)

[1511] Compound B-03-6 (165.51 mg, 0.17 mmol), compound 2-2 (40.00 mg, 0.084 mmol), and 1-hydroxybenzotriazole (33.96 mg, 0.25 mmol) were dissolved in DMF (4 mL); diisopropylethylamine (32.48 mg, 0.25 mmol) was added dropwise and the mixture was stirred for 12 h. The reaction was monitored by HPLC-MS / MS. Water and ethyl acetate were added, stirred, and allowed to stand. The organic phase was washed with saturated brine, dried, and concentrated under reduced pressure to obtain 100.00 mg of a crude product of the title compound, which was directly used in the next step.

[1512] The structural characterization data are as follows:

[1513] ESI-MS (m / z): 1326.2 [M+1] + .

[1514] Step 6: (2S,3S,4S,5R,6S)-6-(2-(2-(2-(2-aminoethoxy)ethoxy)acetamido)-4-((((2-((S)-7-ethyl-7-hydroxy-8,11-dioxo-7,8,11,13-tetrahydro-10H-[1,3]dioxolano[4,5-g]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-14-yl)ethyl)(isopropyl)carbamoyl)oxy)methyl)phenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid (B-03-8)

[1515] Dissolve compound B-03-7 (100.00 mg, 0.08 mmol) in MeOH (5 mL); add 1 drop of dichloromethane; add a 1 mL solution of lithium hydroxide monohydrate (15.82 mg, 0.377 mmol) dropwise, and stir for 2 hours. Monitor the reaction by HPLC-MS / MS; adjust the pH of the reaction solution to 4 by adding 3N aqueous hydrochloric acid; concentrate under reduced pressure and purify by preparative HPLC (conditions as follows); and freeze-dry the prepared solution to obtain 27.00 mg of the title compound.

[1516] Chromatographic column: SunFire Prep C18 OBD 19mm×150mm×5.0μm

[1517] Mobile phase A: acetonitrile; Mobile phase B: water (0.05% formic acid)

[1518] Time [min] Mobile phase A[%] Mobile phase B[%] Flow rate [mL / min] 0.00 20 80 28 2.00 20 80 28 18.00 80 20 28

[1519] The structural characterization data are as follows:

[1520] ESI-MS (m / z): 964.2 [M+1] + .

[1521] Step 7: (2S,3S,4S,5R,6S)-6-(4-((((2-((S)-7-ethyl-7-hydroxy-8,11-dioxo-7,8,11,13-tetrahydro-10H-[1,3]dioxolano[4,5-g]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-14-yl)ethyl)(isopropyl)carbamoyl)oxy)methyl)-2-(2-(2-(2-(6-(2-(methylsulfonyl)pyrimidin-5-yl)hex-5-ynamido)ethoxy)ethoxy)acetamido)phenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid (B-03)

[1522] Compound B-03-8 (27.00 mg, 0.03 mmol) and 2,5-dioxopyrrolidin-1-yl 6-(2-(methylsulfonyl)pyrimidin-5-yl)hex-5-ynoate (A-07-1, 11.26 mg, 0.03 mmol) were dissolved in DMF (1 mL). Diisopropylethylamine (3.62 mg, 0.03 mmol) was added dropwise with stirring. The mixture was allowed to react at room temperature for 4 hours. The reaction was monitored by HPLC-MS / MS. The reaction solution was purified by preparative HPLC (conditions as follows) and the prepared solution was freeze-dried to obtain 11.70 mg of the title compound.

[1523] Chromatographic column: SunFire Prep C18 OBD 19mm×150mm×5.0μm

[1524] Mobile phase A: acetonitrile; Mobile phase B: water (0.05% formic acid)

[1525]

[1526] The structural characterization data are as follows:

[1527] ESI-MS (m / z): 1214.4 [M+1] + .

[1528] Example 7 Synthesis of N-((1S,9S)-4-chloro-9-ethyl-5-fluoro-9-hydroxy-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4:6,7]indolizino[1,2-b]quinolin-1-yl)-2-hydroxyacetamide and N-((1R,9S)-4-chloro-9-ethyl-5-fluoro-9-hydroxy-10,13-dioxo-2,3,9,10,13,15-hexahydro-1H,12H-benzo[de]pyrano[3',4:6,7]indolizino[1,2-b]quinolin-1-yl)-2-hydroxyacetamide (1-11-A and 1-11-B)

[1529]

[1530] Step 1: Synthesis of 3-bromo-4-chloro-5-fluoroaniline (1-5-02)

[1531] Compound 1-5-01 (2.00 g, 10.53 mmol) was dissolved in N,N-dimethylformamide (30 mL); N-chlorosuccinimide (1.69 g, 12.63 mmol) was then slowly added. The mixture was allowed to react at room temperature for 16 hours. The reaction was monitored by HPLC-MS / MS. The reaction solution was concentrated under reduced pressure to obtain a crude product, which was then purified by flash chromatography on a silica gel column (ethyl acetate:petroleum ether = 0-25%) to afford 0.95 g of the title compound.

[1532] The structural characterization data are as follows:

[1533] 1 H NMR (400MHz, DMSO-d6) δ6.77 (dd, J=2.5, 1.4Hz, 1H), 6.51 (dd, J=11.7, 2.5Hz, 1H), 5.84 (s, 2H).

[1534] Step 2: Synthesis of N-(3-bromo-4-chloro-5-fluorophenyl)acetamide (1-5-03)

[1535] Compound 1-5-02 (0.95 g, 4.23 mmol) was dissolved in ethyl acetate (20 mL). Acetic anhydride (648.13 mg, 6.35 mmol) was added under nitrogen. After addition, the temperature was raised to 50°C and the reaction mixture was allowed to react for 15 hours. The reaction was monitored by HPLC-MS / MS. The reaction mixture was quenched with methanol (5 mL) and evaporated to dryness under reduced pressure to obtain a crude product. The crude product was purified by flash chromatography on a silica gel column (ethyl acetate:petroleum ether = 0-40%) to obtain 1.01 g of the title compound.

[1536] The structural characterization data are as follows:

[1537] ESI-MS (m / z): 265.9 [M+H] + .

[1538] Step 3: Synthesis of (E)-4-(5-acetylamino-2-chloro-3-fluorophenyl)-3-butenoic acid (1-5-04)

[1539] Compound 1-5-03 and 3-butenoic acid (387.65 mg, 4.50 mmol) were dissolved in a mixture of 1,4-dioxane (24 mL) and water (8 mL). N,N-diisopropylethylamine (1.45 g, 11.26 mmol), tris(o-methylphenyl)phosphine (114.21 mg, 375.24 μmol), and palladium acetate (42.12 mg, 187.62 μmol) were then added. The reaction system was purged with nitrogen three times and heated to 100°C under a nitrogen atmosphere for 16 hours. The reaction was monitored by HPLC-MS / MS. After cooling the reaction solution to room temperature, 1N aqueous sodium hydroxide solution (60 mL) and ethyl acetate (50 mL) were added and the layers were shaken. After separating the lower aqueous phase, the pH was adjusted to about 3 with a 4 mol / L hydrochloric acid aqueous solution; then extracted with ethyl acetate; the combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, and filtered; the filtrate was evaporated to dryness under reduced pressure to obtain 1.00 g of a crude product of the title compound.

[1540] The structural characterization data are as follows:

[1541] ESI-MS (m / z): 272.0 [M+H] + .

[1542] Step 4: Synthesis of 4-(5-acetylamino-2-chloro-3-fluorophenyl)butyric acid (1-5-05)

[1543] Dissolve the crude product of compound 1-5-04 (1.00 g, 3.68 mmol) in tetrahydrofuran (15 mL); then add 10% palladium on carbon (0.10 g). After addition, replace the reaction system with a hydrogen balloon three times and react under a hydrogen atmosphere for 4 hours. Monitor the reaction by HPLC-MS / MS. Filter the reaction solution, and concentrate the filtrate under reduced pressure to dryness to obtain 1.00 g of the crude title compound.

[1544] The structural characterization data are as follows:

[1545] ESI-MS (m / z): 274.0 [M+H] + .

[1546] Step 5: Synthesis of N-(4-chloro-3-fluoro-8-oxo-5,6,7,8-tetrahydronaphthalen-1-yl)acetamide (1-5-06)

[1547] Dissolve the crude product of compound 1-5-05 (1.00 g, 3.65 mmol) in trifluoroacetic acid (5 mL). After cooling to 5°C, slowly add trifluoroacetic anhydride (3.84 g, 18.27 mmol, 2.54 mL). Maintain the reaction at 5°C for 2 hours. Detect the reaction by HPLC-MS / MS. Pour the reaction solution slowly into water and extract with ethyl acetate. Combine the organic phases, wash with saturated brine, dry over anhydrous sodium sulfate, and filter. Evaporate the filtrate to dryness under reduced pressure to obtain the crude product, which is then purified by flash chromatography on a silica gel column to yield 0.43 g of the title compound.

[1548] The structural characterization data are as follows:

[1549] ESI-MS (m / z): 256.1 [M+H] + .

[1550] Step 6: Synthesis of N-(4-chloro-3-fluoro-7-(hydroxyimino)-8-oxo-5,6,7,8-tetrahydronaphthalen-1-yl)acetamide (1-5-07)

[1551] Tetrahydrofuran (16 mL) and tert-butanol (4 mL) were added to the reaction mixture. After cooling to 5°C in an ice bath, potassium tert-butoxide (415.18 mg, 3.70 mmol) was added. Compound 1-5-06 (0.43 mg, 1.68 mmol) was then dissolved in tetrahydrofuran (1 mL) and added dropwise to the reaction mixture. After 10 minutes, isoamyl nitrite (315.24 mg, 2.69 mmol) was added. After addition, the mixture was maintained at 5°C for 1 hour. The reaction was monitored by HPLC-MS / MS. The reaction mixture was quenched with saturated aqueous ammonium chloride and extracted with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, and filtered. The filtrate was concentrated under reduced pressure to obtain 455.00 mg of the crude title compound.

[1552] The structural characterization data are as follows:

[1553] ESI-MS (m / z): 285.0 [M+H] + .

[1554] Step 7: Synthesis of N-(7-amino-4-chloro-3-fluoro-8-oxo-5,6,7,8-tetrahydronaphthalen-1-yl)acetamide (1-5-08)

[1555] Dissolve the crude product of compound 1-5-07 (0.40 g, 1.41 mmol) in methanol (10 mL); then add 3 mol / L aqueous hydrochloric acid (1 mL) and 10% palladium on carbon (40.00 mg); after addition, replace the reaction system with hydrogen three times; react at room temperature under a hydrogen atmosphere for 1 hour, and monitor the reaction by HPLC-MS / MS. Filter the reaction solution, and concentrate the filtrate under reduced pressure to dryness to obtain 0.43 g of the crude hydrochloride salt of the title compound.

[1556] The structural characterization data are as follows:

[1557] ESI-MS (m / z): 271.0 [M+H] + .

[1558] Step 8: Synthesis of (9H-fluoren-9-yl)methyl (8-acetamido-5-chloro-6-fluoro-1-oxo-1,2,3,4-tetrahydronaphthalen-2-yl)carbamate (1-5-09)

[1559] The crude hydrochloride of compound 1-5-08 (0.43 g, 1.19 mmol) was dissolved in 1,4-dioxane (15 mL). Sodium bicarbonate (400.35 mg, 4.77 mmol), water (5 mL), and 9-fluorenylmethyl-N-succinimidyl carbonate (481.81 mg, 1.43 mmol) were then added. The mixture was stirred at room temperature for 2 hours. The reaction was monitored by HPLC-MS / MS. The reaction mixture was poured into water and extracted with ethyl acetate. The combined organic phases were washed with saturated brine and dried over anhydrous sodium sulfate. The organic phases were filtered, and the filtrate was concentrated under reduced pressure to obtain the crude product. The crude product was purified on a C18 reverse-phase column (acetonitrile: 0.05% formic acid in water = 20% to 100%) to obtain 301.00 mg of the title compound.

[1560] The structural characterization data are as follows:

[1561] ESI-MS (m / z): 493.2 [M+H] + .

[1562] Step 9: Synthesis of (9H-fluoren-9-yl)methyl (8-amino-5-chloro-6-fluoro-1-oxo-1,2,3,4-tetrahydronaphthalen-2-yl)carbamate (1-5-10)

[1563] Compound 1-5-09 (300.00 mg, 608.61 μmol) was dissolved in dioxane (5 mL); 12 mol / L concentrated hydrochloric acid (1 mL) was added. After addition, the mixture was heated to 60°C and reacted for 2 hours. The reaction was monitored by HPLC-MS / MS. The reaction solution was poured into water and extracted with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, and filtered. The filtrate was concentrated under reduced pressure to obtain the crude product. The crude product was purified on...

Claims

1. An antibody drug conjugate having the formula Ab-[MLED] x The structure shown, wherein: Ab is an antibody or an antigen-binding fragment thereof that specifically binds to human PTK7; M is a linker site connected to the antibody or antigen-binding fragment thereof; L is a connector connecting M and E; E is a structural fragment connecting L and D; D is a cytotoxic drug fragment; and x is selected from 1 to 10.

2. The antibody-drug conjugate according to claim 1, wherein the antibody or antigen-binding fragment thereof comprises: (1) the following heavy chain variable region (VH) and / or light chain variable region (VL), wherein the CDRs are defined according to the Chothia numbering system: (1a) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with a sequence of SEQ ID NO: 11 or a variant thereof, CDR-H2 with a sequence of SEQ ID NO: 12 or a variant thereof, and CDR-H3 with a sequence of SEQ ID NO: 13 or a variant thereof; and / or, a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with a sequence of SEQ ID NO: 14 or a variant thereof, CDR-L2 with a sequence of SEQ ID NO: 15 or a variant thereof, and CDR-L3 with a sequence of SEQ ID NO: 16 or a variant thereof; or, (1b) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with a sequence of SEQ ID NO: 27 or a variant thereof, CDR-H2 with a sequence of SEQ ID NO: 28 or a variant thereof, and CDR-H3 with a sequence of SEQ ID NO: 29 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with a sequence of SEQ ID NO: 30 or a variant thereof, CDR-L2 with a sequence of SEQ ID NO: 31 or a variant thereof, and CDR-L3 with a sequence of SEQ ID NO: 32 or a variant thereof; in, The variant described in any one of (1a) and (1b) has at least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity compared to the sequence from which it is derived, or the variant has one or more amino acid substitutions, deletions or additions compared to the sequence from which it is derived; or, (2) the following heavy chain variable region (VH) and / or light chain variable region (VL), wherein the CDRs are defined according to the Kabat numbering system: (2a) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with a sequence of SEQ ID NO: 17 or a variant thereof, CDR-H2 with a sequence of SEQ ID NO: 18 or 19 or a variant thereof, and CDR-H3 with a sequence of SEQ ID NO: 13 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with a sequence of SEQ ID NO: 14 or a variant thereof, CDR-L2 with a sequence of SEQ ID NO: 15 or a variant thereof, and CDR-L3 with a sequence of SEQ ID NO: 16 or a variant thereof; or, (2b) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with a sequence of SEQ ID NO: 33 or a variant thereof, CDR-H2 with a sequence of SEQ ID NO: 34 or 35 or a variant thereof, and CDR-H3 with a sequence of SEQ ID NO: 29 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with a sequence of SEQ ID NO: 30 or a variant thereof, CDR-L2 with a sequence of SEQ ID NO: 31 or a variant thereof, and CDR-L3 with a sequence of SEQ ID NO: 32 or a variant thereof; wherein the variant described in any one of (2a) and (2b) has at least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity compared to the sequence from which it is derived, or the variant has one or more amino acid substitutions, deletions or additions compared to the sequence from which it is derived; or, (3) the following heavy chain variable region (VH) and / or light chain variable region (VL), wherein the CDRs are defined according to the IMGT numbering system: (3a) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with a sequence of SEQ ID NO: 20 or a variant thereof, CDR-H2 with a sequence of SEQ ID NO: 21 or a variant thereof, and CDR-H3 with a sequence of SEQ ID NO: 22 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with a sequence of SEQ ID NO: 23 or a variant thereof, CDR-L2 with a sequence of SEQ ID NO: 24 or a variant thereof, and CDR-L3 with a sequence of SEQ ID NO: 16 or a variant thereof; or, (3b) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 36 or a variant thereof, CDR-H2 of SEQ ID NO: 37 or a variant thereof, and CDR-H3 of SEQ ID NO: 38 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 39 or a variant thereof, CDR-L2 of SEQ ID NO: 40 or a variant thereof, and CDR-L3 of SEQ ID NO: 32 or a variant thereof; wherein the variant described in any one of (3a) and (3b) has at least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with the sequence from which it is derived, or the variant has one or more amino acid substitutions, deletions or additions compared to the sequence from which it is derived; or, (4) the following heavy chain variable region (VH) and / or light chain variable region (VL), wherein the CDRs are defined according to the AbM numbering system: (4a) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 25 or a variant thereof, CDR-H2 of SEQ ID NO: 26 or a variant thereof, and CDR-H3 of SEQ ID NO: 13 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 14 or a variant thereof, CDR-L2 of SEQ ID NO: 15 or a variant thereof, and CDR-L3 of SEQ ID NO: 16 or a variant thereof; or, (4b) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 41 or a variant thereof, CDR-H2 of SEQ ID NO: 42 or a variant thereof, and CDR-H3 of SEQ ID NO: 29 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 30 or a variant thereof, CDR-L2 of SEQ ID NO: 31 or a variant thereof, and CDR-L3 of SEQ ID NO: 32 or a variant thereof; wherein the variant described in any one of (4a) and (4b) has at least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity compared to the sequence from which it is derived, or the variant has one or more amino acid substitutions, deletions or additions compared to the sequence from which it is derived.

3. The antibody-drug conjugate according to claim 1 or 2, wherein the antibody or antigen-binding fragment thereof comprises: (a) VH shown in SEQ ID NO: 1 or a variant thereof, and / or VL shown in SEQ ID NO: 2 or a variant thereof; (b) VH shown in SEQ ID NO: 3 or a variant thereof, and / or VL shown in SEQ ID NO: 4 or a variant thereof; (c) VH shown in SEQ ID NO: 5 or a variant thereof, and / or VL shown in SEQ ID NO: 6 or a variant thereof; (d) VH shown in SEQ ID NO: 7 or a variant thereof, and / or VL shown in SEQ ID NO: 8 or a variant thereof; or (e) VH shown in SEQ ID NO: 9 or a variant thereof, and / or VL shown in SEQ ID NO: 10 or a variant thereof; in, The variant has at least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity compared to the sequence from which it is derived, or the variant has one or several amino acid substitutions, deletions or additions compared to the sequence from which it is derived.

4. The antibody-drug conjugate according to any one of claims 1 to 3, wherein the antibody or antigen-binding fragment thereof further comprises: (a) a heavy chain constant region (CH) of a human immunoglobulin or a variant thereof, wherein the variant has one or more amino acid substitutions, deletions or additions compared to the wild-type sequence from which it is derived; and (b) a light chain constant region (CL) of a human immunoglobulin or a variant thereof, wherein the variant has one or more amino acid substitutions, deletions or additions compared to the wild-type sequence from which it is derived.

5. The antibody-drug conjugate according to any one of claims 1 to 4, wherein the antibody or antigen-binding fragment thereof comprises: (1) a heavy chain comprising a VH sequence of SEQ ID NO: 1 and a heavy chain constant region (CH) of SEQ ID NO: 43, and a light chain comprising a VL sequence of SEQ ID NO: 2 and a light chain constant region (CL) of SEQ ID NO: 44; (2) a heavy chain comprising a VH sequence of SEQ ID NO: 3 and a heavy chain constant region (CH) of SEQ ID NO: 43 or 45, and a light chain comprising a VL sequence of SEQ ID NO: 4 and a light chain constant region (CL) of SEQ ID NO: 44; (3) a heavy chain comprising a VH sequence of SEQ ID NO: 5 and a heavy chain constant region (CH) of SEQ ID NO: 43, and a light chain comprising a VL sequence of SEQ ID NO: 6 and a light chain constant region (CL) of SEQ ID NO: 44; (4) a heavy chain comprising a VH sequence of SEQ ID NO: 7 and a heavy chain constant region (CH) of SEQ ID NO: 43, and a light chain comprising a VL sequence of SEQ ID NO: 8 and a light chain constant region (CL) of SEQ ID NO: 44; or (5) A heavy chain comprising the VH of SEQ ID NO: 9 and the heavy chain constant region (CH) of SEQ ID NO: 43, and a light chain comprising the VL of SEQ ID NO: 10 and the light chain constant region (CL) of SEQ ID NO:

44.

6. The antibody drug conjugate according to any one of claims 1 to 5, wherein M is Ring A is a 5-6 membered alicyclic heterocyclic ring or a 5-20 membered aromatic ring system, wherein the alicyclic heterocyclic ring and the aromatic ring system are optionally substituted by one or more selected from oxy (=O), halogen, cyano, amino, carboxyl, thiol and C 1-6 alkyl; and M1 is selected from a single bond, C 1-20 Alkylene, C 2-20 Alkenylene and C 2-20 Alkyne group.

7. The antibody drug conjugate according to any one of claims 1 to 5, wherein M is Wherein ring A is a 5-membered alicyclic heterocycle, a 6-membered heteroaromatic ring, or a polycyclic ring formed by connecting one or more 6-membered aromatic heterocycles to a benzene ring via a single bond, wherein the alicyclic heterocycle is optionally substituted with one or more selected from oxy (=O), halogen and C 1-4 alkyl; and M1 is selected from a single bond, C 3-10 Alkylene, C 3-10 Alkenylene and C 3-10 Alkyne group.

8. The antibody drug conjugate according to any one of claims 1 to 5, wherein M is wherein ring A is selected from and M1 is selected from a single bond, C 5-8 Alkylene, C 5-8 Alkenylene and C 5-8 Alkyne group.

9. The antibody drug conjugate according to any one of claims 1 to 5, wherein M is selected from 10. The antibody drug conjugate according to any one of claims 1 to 5, wherein M is 11. The antibody drug conjugate according to any one of claims 1 to 10, wherein L is selected from a structure comprising one or more of the following: C 1-6 Alkylene, -N(R')-, Carbonyl, -O-, Val, Cit, Phe, Lys, Lys(COCH2CH2(OCH2CH2) s OCH3), D-Val, Leu, Gly, Ala, Asn, Val-Cit, Val-Ala, Val-Lys, Val-Lys(Ac), Phe-Lys, Phe-Lys(Ac), D-Val-Leu- Lys, Gly-Gly-Arg, Ala-Ala-Asn, Ala-Ala-Ala, Val-Lys-Ala, Val-Lys-Gly, Gly-Gly-Gly, Gly-Gly-Phe-Gly (SEQ ID NO:48), Gly-Phe-Leu-Gly (SEQ ID NO:49), Gly-Gly-Val-Ala (SEQ ID NO:50), Gly-Gly-Gly-Gly-Gly (SEQ ID NO:51), Where R' represents hydrogen, C 1-6 Alkyl or alkyl containing -(CH2CH2O)r-; r is selected from an integer of 1-10; s is selected from an integer of 1-20.

12. The antibody drug conjugate according to any one of claims 1 to 10, wherein L is selected from a structure comprising one or more of the following: C 1-6 Alkylene, -NH-, Val, Cit, Phe, Lys, Lys(COCH2CH2(OCH2CH2) s OCH3), Gly, Val-Cit, Gly-Gly-Phe-Gly (SEQ ID NO: 48), And s is selected from an integer of 1-20.

13. The antibody drug conjugate according to any one of claims 1 to 10, wherein L is selected from the following structures: s is an integer selected from 1-20.

14. The antibody drug conjugate according to any one of claims 1 to 10, wherein L is selected from the following structures:

15. The antibody drug conjugate according to any one of claims 1 to 10, wherein L is selected from the following structures:

16. The antibody drug conjugate according to any one of claims 1 to 15, wherein E is a single bond, -NH-CH2-, -NH-CH2-O-CH2-CO-, 17. The antibody drug conjugate according to any one of claims 1 to 15, wherein E is a single bond, -NH-CH2-, -NH-CH2-O-CH2-CO-, 18. The antibody drug conjugate according to any one of claims 1 to 10, wherein E is -NH-CH2-, 19. The antibody drug conjugate according to any one of claims 1 to 10, wherein E is -NH-CH2- or 20. The antibody drug conjugate according to any one of claims 1 to 19, wherein Select from the following structures:

21. The antibody drug conjugate according to any one of claims 1 to 20, wherein the cytotoxic drug is selected from tubulin inhibitors, DNA intercalators, DNA topoisomerase inhibitors and RNA polymerase inhibitors or pharmaceutically acceptable salts, esters or analogs thereof.

22. The antibody-drug conjugate according to claim 21, wherein the tubulin inhibitor is an auristatin compound or a maytansine compound.

23. The antibody drug conjugate of claim 21, wherein the DNA intercalator is pyrrolobenzodiazepine (PBD). The antibody-drug conjugate of claim 21 , wherein the DNA topoisomerase inhibitor is a topoisomerase I inhibitor or a topoisomerase II inhibitor.

25. The antibody drug conjugate of claim 24, wherein the topoisomerase I inhibitor is camptothecin, hydroxycamptothecin, 9-aminocamptothecin, SN-38, irinotecan, topotecan, belotecan or rubitecan.

26. The antibody drug conjugate of claim 24, wherein the topoisomerase II inhibitor is doxorubicin, PNU-159682, duocarmycin, daunorubicin, mitoxantrone, podophyllotoxin or etoposide.

27. The antibody-drug conjugate of claim 21, wherein the RNA polymerase inhibitor is α-amanitin or a pharmaceutically acceptable salt, ester or analog thereof.

28. The antibody drug conjugate according to claim 21, wherein the cytotoxic drug is 29. The antibody-drug conjugate according to claim 21, wherein the cytotoxic drug is selected from the compounds represented by Formula I and Formula II: R1 and R2 are each independently selected from C 1-6 Alkyl and halogen; R3 is selected from H and -CO-CH2OH; R4 and R5 are each independently selected from H, halogen and hydroxyl; or R4 and R5 are connected to the connected carbon atom to form a 5-6 membered oxygen-containing heterocyclic ring; R6 is selected from hydrogen and -C 1-4 Alkylene-NR a R b ; R7 is selected from hydrogen, C 1-6 Alkyl and -C 1-4 Alkylene-NR a R b ;as well as R a and R b Each occurrence is independently selected from H, C 1-6 Alkyl, -SO2-C 1-6 Alkyl and -CO-C 1-6 alkyl.

30. The antibody drug conjugate according to claim 21, wherein the cytotoxic drug is selected from the following compounds: The corresponding fragment of the cytotoxic drug obtained after the cytotoxic drug is connected to the linker is D shown in the general formula.

31. The antibody drug conjugate according to claim 30, wherein D is a monovalent structure obtained by losing one H from -OH, -NH2 or a secondary amine group on the cytotoxic drug.

32. The antibody drug conjugate according to any one of claims 1 to 31, selected from ADC A-01 to ADCA-26, ADC B-01 to ADC B-06 and ADC C01 shown below: ADC A-01 ADC A-02 ADC A-03 ADC A-04 ADC A-05 ADC A-06 ADC A-07 ADC A-08 ADC A-09 ADC A-10 ADC A-11 ADC A-12 ADC A-13 ADC A-14 ADC A-15 ADC A-16 ADC A-17 ADC A-18 ADC A-19 ADC A-20 ADC A-21 ADC A-22 ADC A-23 ADC A-24 ADC A-25 ADC A-26 ADC B-01 ADC B-02 ADC B-03 ADC B-04 ADC B-05 ADC B-06 and ADC C-01 in, The HA in each antibody drug conjugate represents an antibody or antigen-binding fragment thereof comprising a VH as shown in SEQ ID NO: 1, 3, 5, 7 or 9 and a VL as shown in SEQ ID NO: 2, 4, 6, 8 or 10, and It indicates the connection formed by the thiol group in the antibody or antigen-binding fragment thereof and the linker.

33. The antibody drug conjugate according to any one of claims 1-32, wherein the antibody drug conjugate comprises the following formula: in, The HA in each antibody drug conjugate is selected from: (1) an antibody or an antigen-binding fragment thereof comprising a VH sequence as shown in SEQ ID NO: 1 and a VL sequence as shown in SEQ ID NO: 2, (2) an antibody or an antigen-binding fragment thereof comprising a VH sequence as shown in SEQ ID NO: 3 and a VL sequence as shown in SEQ ID NO: 4, (3) an antibody or antigen-binding fragment thereof comprising a VH sequence as shown in SEQ ID NO: 5 and a VL sequence as shown in SEQ ID NO: 6, (4) an antibody or an antigen-binding fragment thereof comprising a VH sequence as shown in SEQ ID NO: 7 and a VL sequence as shown in SEQ ID NO: 8, and (5) an antibody or antigen-binding fragment thereof comprising a VH sequence as shown in SEQ ID NO: 9 and a VL sequence as shown in SEQ ID NO: 10, represents the connection formed by the thiol group in the antibody or antigen-binding fragment thereof and the linker, wherein HA is linked via the sulfhydryl groups in the antibody or antigen-binding fragment thereof to form an antibody-drug conjugate.

34. The antibody drug conjugate according to any one of claims 1-32, wherein the antibody drug conjugate comprises the following formula: in, The HA in each antibody drug conjugate is selected from: (1) an antibody or antigen-binding fragment thereof comprising a VH having a sequence as shown in SEQ ID NO: 1, a CH having a sequence as shown in SEQ ID NO: 43 or 45, a VL having a sequence as shown in SEQ ID NO: 2, and a CL having a sequence as shown in SEQ ID NO: 44; (2) an antibody or antigen-binding fragment thereof comprising a VH having a sequence as shown in SEQ ID NO: 3, a CH having a sequence as shown in SEQ ID NO: 43 or 45, a VL having a sequence as shown in SEQ ID NO: 4, and a CL having a sequence as shown in SEQ ID NO: 44; (3) an antibody or antigen-binding fragment thereof comprising a VH having a sequence as shown in SEQ ID NO: 5, a CH having a sequence as shown in SEQ ID NO: 43 or 45, a VL having a sequence as shown in SEQ ID NO: 6, and a CL having a sequence as shown in SEQ ID NO: 44; (4) an antibody or antigen-binding fragment thereof comprising a VH having a sequence as shown in SEQ ID NO: 7, a CH having a sequence as shown in SEQ ID NO: 43 or 45, a VL having a sequence as shown in SEQ ID NO: 8, and a CL having a sequence as shown in SEQ ID NO: 44; and (5) an antibody or antigen-binding fragment thereof comprising a VH having a sequence as shown in SEQ ID NO: 9, a CH having a sequence as shown in SEQ ID NO: 43 or 45, a VL having a sequence as shown in SEQ ID NO: 10, and a CL having a sequence as shown in SEQ ID NO: 44; represents the connection formed by the thiol group in the antibody or antigen-binding fragment thereof and the linker; wherein HA is linked via one or more thiol groups in the antibody or antigen-binding fragment thereof to form an antibody-drug conjugate; And x is 3-9.

35. The antibody drug conjugate of any one of claims 1-34, wherein the antibody or antigen-binding fragment thereof comprises VH as shown in SEQ ID NO: 3 and CH as shown in SEQ ID NO: 43 or 45, VL as shown in SEQ ID NO: 4 and CL as shown in SEQ ID NO:

44.

36. The antibody drug conjugate according to any one of claims 1 to 33, wherein the antibody drug conjugate is: in, The HA in the antibody-drug conjugate is an antibody or an antigen-binding fragment thereof comprising a VH having a sequence as shown in SEQ ID NO: 3, a heavy chain constant region (CH) having a sequence as shown in SEQ ID NO: 43 or 45, a VL having a sequence as shown in SEQ ID NO: 4, and a light chain constant region (CL) having a sequence as shown in SEQ ID NO: 44; represents a connection formed between the thiol group in the antibody or antigen-binding fragment thereof and the linker; HA is linked via the sulfhydryl group in the antibody or antigen-binding fragment thereof to form an antibody-drug conjugate; And x is 7-9.

37. The antibody drug conjugate according to any one of claims 1 to 33, wherein the antibody drug conjugate is: in, HA is an antibody or an antigen-binding fragment thereof comprising a heavy chain (HC) having a sequence as shown in SEQ ID NO: 46 and a light chain (LC) having a sequence as shown in SEQ ID NO: 47; represents a connection formed between the thiol group in the antibody or antigen-binding fragment thereof and the linker; HA is linked via the thiol group of cysteine ​​in the antibody or antigen-binding fragment thereof to form an antibody-drug conjugate; And x is 7-8.

38. The antibody drug conjugate according to any one of claims 1 to 33, wherein the antibody drug conjugate is: in, HA is an antibody comprising a heavy chain (HC) having a sequence as shown in SEQ ID NO: 52, 53 or 54 and a light chain (LC) consisting of a sequence as shown in SEQ ID NO: 47; represents a connection formed between the thiol group in the antibody or antigen-binding fragment thereof and the linker; HA is linked via the thiol group of cysteine ​​in the antibody or antigen-binding fragment thereof to form an antibody-drug conjugate; And x is 7-8.

39. The antibody drug conjugate according to any one of claims 1 to 33, wherein the antibody drug conjugate is: in, HA is an antibody composed of two heavy chains and two light chains, each heavy chain (HC) consists of the sequence shown in SEQ ID NO: 52, and each light chain (LC) consists of the sequence shown in SEQ ID NO: 47; and represents a connection formed between the thiol group in the antibody or antigen-binding fragment thereof and the linker; HA is linked via the thiol group of cysteine ​​in the antibody or antigen-binding fragment thereof to form an antibody-drug conjugate; And x is 7-8.

40. The antibody drug conjugate according to any one of claims 1-33, wherein the antibody drug conjugate is: in, HA is an antibody or an antigen-binding fragment thereof comprising a heavy chain (HC) having a sequence as shown in SEQ ID NO: 46 and a light chain (LC) having a sequence as shown in SEQ ID NO: 47; It indicates the connection between the thiol group in the antibody or antigen-binding fragment thereof and the linker; HA is linked through the thiol group of cysteine ​​in the antibody or its antigen-binding fragment to form an antibody-drug conjugate; And x is 8.

41. The antibody drug conjugate according to any one of claims 1 to 33, wherein the antibody drug conjugate is: in, The HA in the antibody-drug conjugate is an antibody or an antigen-binding fragment thereof comprising a VH having a sequence as shown in SEQ ID NO: 3, a heavy chain constant region (CH) having a sequence as shown in SEQ ID NO: 43 or 45, a VL having a sequence as shown in SEQ ID NO: 4, and a light chain constant region (CL) having a sequence as shown in SEQ ID NO: 44; represents a connection formed between the thiol group in the antibody or antigen-binding fragment thereof and the linker; HA is linked via the sulfhydryl group in the antibody or antigen-binding fragment thereof to form an antibody-drug conjugate; And x is 7-9.

42. The antibody drug conjugate according to any one of claims 1-33, wherein the antibody drug conjugate is: in, HA is an antibody or an antigen-binding fragment thereof comprising a heavy chain (HC) having a sequence as shown in SEQ ID NO: 46 and a light chain (LC) having a sequence as shown in SEQ ID NO: 47; represents a connection formed between the thiol group in the antibody or antigen-binding fragment thereof and the linker; HA is linked via the sulfhydryl group in the antibody or antigen-binding fragment thereof to form an antibody-drug conjugate; And x is 7-8.

43. The antibody drug conjugate according to any one of claims 1-33, wherein the antibody drug conjugate is: in, HA is an antibody or an antigen-binding fragment thereof comprising a heavy chain (HC) having a sequence as shown in SEQ ID NO: 46 and a light chain (LC) having a sequence as shown in SEQ ID NO: 47; represents a connection formed between the thiol group in the antibody or antigen-binding fragment thereof and the linker; HA is linked via the sulfhydryl group in the antibody or antigen-binding fragment thereof to form an antibody-drug conjugate; And x is 8.

44. The antibody drug conjugate according to any one of claims 1-33, wherein the antibody drug conjugate is: in, The HA in the antibody-drug conjugate is an antibody or an antigen-binding fragment thereof comprising a VH having a sequence as shown in SEQ ID NO: 3, a heavy chain constant region (CH) having a sequence as shown in SEQ ID NO: 43 or 45, a VL having a sequence as shown in SEQ ID NO: 4, and a light chain constant region (CL) having a sequence as shown in SEQ ID NO: 44; represents a connection formed between the thiol group in the antibody or antigen-binding fragment thereof and the linker; HA is linked via the sulfhydryl group in the antibody or antigen-binding fragment thereof to form an antibody-drug conjugate; And x is 7-9.

45. The antibody drug conjugate according to any one of claims 1-33, wherein the antibody drug conjugate is: in, HA is an antibody or an antigen-binding fragment thereof comprising a heavy chain (HC) having a sequence as shown in SEQ ID NO: 46 and a light chain (LC) having a sequence as shown in SEQ ID NO: 47; represents a connection formed between the thiol group in the antibody or antigen-binding fragment thereof and the linker; HA is linked via the sulfhydryl group in the antibody or antigen-binding fragment thereof to form an antibody-drug conjugate; And x is 7-8.

46. ​​The antibody drug conjugate according to any one of claims 1-33, wherein the antibody drug conjugate is: in, HA is an antibody or an antigen-binding fragment thereof comprising a heavy chain (HC) having a sequence as shown in SEQ ID NO: 46 and a light chain (LC) having a sequence as shown in SEQ ID NO: 47; represents the connection formed by the thiol group in the antibody or antigen-binding fragment thereof and the linker; wherein HA is linked via the sulfhydryl group in the antibody or antigen-binding fragment thereof to form an antibody-drug conjugate; And x is 8.

47. A composition comprising one or more antibody drug conjugates according to any one of claims 1-33, wherein the DAR (drug antibody conjugate ratio) of the composition is 1-2, 1-3, 1-4, 1-5, 1-6, 1-7, 1-8, 1-9, 1-10, 2-3, 2-4, 2-5, 2-6, 2-7, 2-8, 2-9, 2-10, 3-4, 3-5, 3-6, 3-7, 3-8, 3-9, 3-10, 4-5, 4-6, 4-7, 4-8, 4-9, 4-10, 5-6, 5-7, 5-8, 5-9, 5-10, 6-7, 6-8, 6-9, 6-10, 7-8, 7-9, 7-10, 8-9, 8-10, or 9-10.

48. The antibody drug conjugate of any one of claims 1-47, wherein (i) the C-terminus of the heavy chain lacks a lysine residue; (ii) the N-terminus of the heavy chain is glutamine, glutamic acid, pyroglutamate or pyroglutamic acid; or, (iii) the C-terminus of the heavy chain lacks a lysine residue and the N-terminus of the heavy chain is glutamine, glutamic acid, pyroglutamate or pyroglutamate.

49. The antibody drug conjugate of any one of claims 1-46 and 48, wherein the antibody or antigen-binding fragment thereof comprises a heavy chain comprising a VH having a sequence selected from SEQ ID NOs: 1, 3, 5, 7 and 9.

50. The antibody drug conjugate of any one of claims 1-46, 48 and 49, wherein the antibody or antigen-binding fragment thereof comprises a heavy chain comprising a VH having a sequence selected from SEQ ID NOs: 1, 3, 5, 7 and 9, wherein the N-terminal glutamine or glutamic acid of the antibody variable region has been cyclized to pyroglutamate or pyroglutamic acid.

51. The antibody drug conjugate of any one of claims 1-46 and 48-50, wherein the antibody or antigen-binding fragment thereof comprises a light chain comprising a VL having a sequence as shown in SEQ ID NO: 8, wherein the N-terminal glutamic acid in the variable region of the antibody light chain has been cyclized to pyroglutamate or pyroglutamic acid.

52. The antibody drug conjugate of any one of claims 1-46 and 48-51, wherein the antibody or antigen-binding fragment thereof comprises: (a) a heavy chain having the sequence shown in SEQ ID NO: 52 and a light chain having the sequence shown in SEQ ID NO: 47; (b) a heavy chain having the sequence shown in SEQ ID NO: 53 and a light chain having the sequence shown in SEQ ID NO: 47; or (c) a heavy chain having the sequence shown in SEQ ID NO:54 and a light chain having the sequence shown in SEQ ID NO:

47.

53. The antibody drug conjugate according to any one of claims 1-46 and 48-52, wherein the antibody or antigen binding fragment thereof can be obtained by expressing a nucleic acid molecule encoding the heavy chain and light chain of the antibody or antigen binding fragment thereof in a host cell, wherein the antibody or antigen binding fragment thereof comprises: (1) a heavy chain comprising the VH sequence shown in SEQ ID NO: 1 and the heavy chain constant region (CH) shown in SEQ ID NO: 43, and a light chain comprising the VL sequence shown in SEQ ID NO: 2 and the light chain constant region (CL) shown in SEQ ID NO: 44; (2) a heavy chain comprising the VH sequence shown in SEQ ID NO: 3 and the heavy chain constant region (CH) shown in SEQ ID NO: 43 or 45, and a light chain comprising the VL sequence shown in SEQ ID NO: 4 and the light chain constant region (CL) shown in SEQ ID NO: 44; (3) a heavy chain comprising the VH sequence shown in SEQ ID NO:5 and the heavy chain constant region (CH) shown in SEQ ID NO:43, and a light chain comprising the VL sequence shown in SEQ ID NO:6 and the light chain constant region (CL) shown in SEQ ID NO:44; (4) a heavy chain comprising the VH sequence shown in SEQ ID NO:7 and the heavy chain constant region (CH) shown in SEQ ID NO:43, and a light chain comprising the VL sequence shown in SEQ ID NO:8 and the light chain constant region (CL) shown in SEQ ID NO:44; (5) a heavy chain comprising the VH sequence shown in SEQ ID NO:9 and the heavy chain constant region (CH) shown in SEQ ID NO:43, and a light chain comprising the VL sequence shown in SEQ ID NO:10 and the light chain constant region (CL) shown in SEQ ID NO:44; (6) a heavy chain (HC) having an amino acid sequence as shown in SEQ ID NO: 46 and a light chain (LC) having an amino acid sequence as shown in SEQ ID NO: 47; (7) VH with the amino acid sequence shown in SEQ ID NO: 1 and VL with the amino acid sequence shown in SEQ ID NO: 2; (8) VH with the amino acid sequence shown in SEQ ID NO: 3 and VL with the amino acid sequence shown in SEQ ID NO: 4; (9) VH with the amino acid sequence shown in SEQ ID NO: 5 and VL with the amino acid sequence shown in SEQ ID NO: 6; (10) a VH having an amino acid sequence as shown in SEQ ID NO: 7 and a VL having an amino acid sequence as shown in SEQ ID NO: 8; or (11) VH having the amino acid sequence shown in SEQ ID NO: 9 and VL having the amino acid sequence shown in SEQ ID NO:

10.

54. An antibody drug conjugate comprising an antibody that binds to PTK7, the antibody being conjugated to a drug linker having a structure as shown in formula MLED via one or more cysteine ​​or lysine residues or non-natural amino acid substitutions of the antibody amino acids, wherein M is in, Ring A is a 5-6 membered alicyclic heterocyclic ring or a 5-20 membered aromatic ring system, wherein the alicyclic heterocyclic ring and the aromatic ring system are optionally substituted by one or more selected from oxy (=O), halogen, cyano, amino, carboxyl, thiol and C 1-6 Alkyl radical substitution; M1 is selected from a single bond, C 1-20 Alkylene, C 2-20 Alkenylene and C 2-20 Alkyne; L is a connector connecting M and E; E is a structural fragment connecting L and D; and D is a cytotoxic drug fragment.

55. The antibody drug conjugate of claim 54, wherein Selected from:

56. The antibody drug conjugate of claim 54 or 55, wherein the cytotoxic drug is selected from the following compounds: The corresponding fragment of the cytotoxic drug obtained after the cytotoxic drug is connected to the linker is D shown in the general formula.

57. The antibody drug conjugate of claim 54 or 55, wherein the antibody that binds to PTK7 is selected from the group consisting of: (1) the following heavy chain variable region (VH) and / or light chain variable region (VL), wherein the CDRs are defined according to the Chothia numbering system: (1a) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with a sequence of SEQ ID NO: 11 or a variant thereof, CDR-H2 with a sequence of SEQ ID NO: 12 or a variant thereof, and CDR-H3 with a sequence of SEQ ID NO: 13 or a variant thereof; and / or, a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with a sequence of SEQ ID NO: 14 or a variant thereof, CDR-L2 with a sequence of SEQ ID NO: 15 or a variant thereof, and CDR-L3 with a sequence of SEQ ID NO: 16 or a variant thereof; or, (1b) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with a sequence of SEQ ID NO: 27 or a variant thereof, CDR-H2 with a sequence of SEQ ID NO: 28 or a variant thereof, and CDR-H3 with a sequence of SEQ ID NO: 29 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with a sequence of SEQ ID NO: 30 or a variant thereof, CDR-L2 with a sequence of SEQ ID NO: 31 or a variant thereof, and CDR-L3 with a sequence of SEQ ID NO: 32 or a variant thereof; in, The variant described in any one of (1a) and (1b) has at least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity compared to the sequence from which it is derived, or the variant has one or more amino acid substitutions, deletions or additions compared to the sequence from which it is derived; or, (2) the following heavy chain variable region (VH) and / or light chain variable region (VL), wherein the CDRs are defined according to the Kabat numbering system: (2a) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with a sequence of SEQ ID NO: 17 or a variant thereof, CDR-H2 with a sequence of SEQ ID NO: 18 or 19 or a variant thereof, and CDR-H3 with a sequence of SEQ ID NO: 13 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with a sequence of SEQ ID NO: 14 or a variant thereof, CDR-L2 with a sequence of SEQ ID NO: 15 or a variant thereof, and CDR-L3 with a sequence of SEQ ID NO: 16 or a variant thereof; or, (2b) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with a sequence of SEQ ID NO: 33 or a variant thereof, CDR-H2 with a sequence of SEQ ID NO: 34 or 35 or a variant thereof, and CDR-H3 with a sequence of SEQ ID NO: 29 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with a sequence of SEQ ID NO: 30 or a variant thereof, CDR-L2 with a sequence of SEQ ID NO: 31 or a variant thereof, and CDR-L3 with a sequence of SEQ ID NO: 32 or a variant thereof; wherein the variant described in any one of (2a) and (2b) has at least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity compared to the sequence from which it is derived, or the variant has one or more amino acid substitutions, deletions or additions compared to the sequence from which it is derived; or, (3) the following heavy chain variable region (VH) and / or light chain variable region (VL), wherein the CDRs are defined according to the IMGT numbering system: (3a) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 with a sequence of SEQ ID NO: 20 or a variant thereof, CDR-H2 with a sequence of SEQ ID NO: 21 or a variant thereof, and CDR-H3 with a sequence of SEQ ID NO: 22 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 with a sequence of SEQ ID NO: 23 or a variant thereof, CDR-L2 with a sequence of SEQ ID NO: 24 or a variant thereof, and CDR-L3 with a sequence of SEQ ID NO: 16 or a variant thereof; or, (3b) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 36 or a variant thereof, CDR-H2 of SEQ ID NO: 37 or a variant thereof, and CDR-H3 of SEQ ID NO: 38 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 39 or a variant thereof, CDR-L2 of SEQ ID NO: 40 or a variant thereof, and CDR-L3 of SEQ ID NO: 32 or a variant thereof; wherein the variant described in any one of (3a) and (3b) has at least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity with the sequence from which it is derived, or the variant has one or more amino acid substitutions, deletions or additions compared to the sequence from which it is derived; and, (4) the following heavy chain variable region (VH) and / or light chain variable region (VL), wherein the CDRs are defined according to the AbM numbering system: (4a) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 25 or a variant thereof, CDR-H2 of SEQ ID NO: 26 or a variant thereof, and CDR-H3 of SEQ ID NO: 13 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 14 or a variant thereof, CDR-L2 of SEQ ID NO: 15 or a variant thereof, and CDR-L3 of SEQ ID NO: 16 or a variant thereof; or, (4b) a heavy chain variable region (VH) comprising the following three CDRs: CDR-H1 of SEQ ID NO: 41 or a variant thereof, CDR-H2 of SEQ ID NO: 42 or a variant thereof, and CDR-H3 of SEQ ID NO: 29 or a variant thereof; and / or a light chain variable region (VL) comprising the following three CDRs: CDR-L1 of SEQ ID NO: 30 or a variant thereof, CDR-L2 of SEQ ID NO: 31 or a variant thereof, and CDR-L3 of SEQ ID NO: 32 or a variant thereof; wherein the variant described in any one of (4a) and (4b) has at least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity compared to the sequence from which it is derived, or the variant has one or more amino acid substitutions, deletions or additions compared to the sequence from which it is derived.

58. The antibody drug conjugate of claim 57, wherein the antibody that binds to PTK7 is selected from the group consisting of: (a) VH shown in SEQ ID NO: 1 or a variant thereof, and / or VL shown in SEQ ID NO: 2 or a variant thereof; (b) VH shown in SEQ ID NO: 3 or a variant thereof, and / or VL shown in SEQ ID NO: 4 or a variant thereof; (c) VH shown in SEQ ID NO: 5 or a variant thereof, and / or VL shown in SEQ ID NO: 6 or a variant thereof; (d) VH shown in SEQ ID NO: 7 or a variant thereof, and / or VL shown in SEQ ID NO: 8 or a variant thereof; and (e) VH shown in SEQ ID NO: 9 or a variant thereof, and / or VL shown in SEQ ID NO: 10 or a variant thereof; in, The variant has at least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity compared to the sequence from which it is derived, or the variant has one or several amino acid substitutions, deletions or additions compared to the sequence from which it is derived.

59. The antibody drug conjugate according to claim 54 or 55, wherein the antibody that binds to PTK7 is selected from the following groups: (1) a heavy chain comprising a VH sequence of SEQ ID NO: 1 and a heavy chain constant region (CH) of SEQ ID NO: 43, and a light chain comprising a VL sequence of SEQ ID NO: 2 and a light chain constant region (CL) of SEQ ID NO: 44; (2) a heavy chain comprising a VH sequence of SEQ ID NO: 3 and a heavy chain constant region (CH) of SEQ ID NO: 43 or 45, and a light chain comprising a VL sequence of SEQ ID NO: 4 and a light chain constant region (CL) of SEQ ID NO: 44; (3) a heavy chain comprising a VH sequence of SEQ ID NO: 5 and a heavy chain constant region (CH) of SEQ ID NO: 43, and a light chain comprising a VL sequence of SEQ ID NO: 6 and a light chain constant region (CL) of SEQ ID NO: 44; (4) a heavy chain comprising a VH sequence of SEQ ID NO: 7 and a heavy chain constant region (CH) of SEQ ID NO: 43, and a light chain comprising a VL sequence of SEQ ID NO: 8 and a light chain constant region (CL) of SEQ ID NO: 44; and (5) A heavy chain comprising the VH of SEQ ID NO: 9 and the heavy chain constant region (CH) of SEQ ID NO: 43, and a light chain comprising the VL of SEQ ID NO: 10 and the light chain constant region (CL) of SEQ ID NO:

44.

60. The antibody-drug conjugate according to claim 54 or 55, wherein the antibody that binds to PTK7 comprises: a heavy chain having the amino acid sequence shown in SEQ ID NO: 46 and a light chain having the amino acid sequence shown in SEQ ID NO:

47.

61. A pharmaceutical composition comprising the antibody drug conjugate according to any one of claims 1-60, and one or more pharmaceutically acceptable excipients.

62. A method for treating a cancer that over-expresses PTK7 in a subject, comprising administering to the subject an effective amount of the antibody drug conjugate according to any one of claims 1-60 or the pharmaceutical composition according to claim 61.

63. The method of claim 62, wherein the cancer comprises a solid tumor or a hematological malignancy.

64. The method of claim 63, wherein the cancer is lung cancer, breast cancer, epidermal cancer, ovarian cancer, or esophageal cancer.

65. Use of the antibody drug conjugate according to any one of claims 1 to 60 or the pharmaceutical composition according to claim 61 in the preparation of a medicament for treating cancers that overexpress PTK7.

66. The use of claim 65, wherein the cancer comprises a solid tumor or a hematological malignancy. The use according to claim 65 , wherein the cancer with high PTK7 expression is lung cancer, breast cancer, epidermal cancer, ovarian cancer or esophageal cancer.

68. Use of the antibody drug conjugate according to any one of claims 1 to 60 or the pharmaceutical composition according to claim 61 in treating cancers that overexpress PTK7.

69. The use of claim 68, wherein the cancer comprises a solid tumor or a hematological malignancy.

70. The use according to claim 69, wherein the cancer with high PTK7 expression is lung cancer, breast cancer, epidermal cancer, ovarian cancer or esophageal cancer.

71. The antibody drug conjugate according to any one of claims 1-60, or the pharmaceutical composition according to claim 61, for use in treating cancers that highly express PTK7.

72. The antibody drug conjugate of claim 71, wherein the cancer comprises a solid tumor or a hematological malignancy.

73. The antibody drug conjugate of claim 72, wherein the cancer is lung cancer, breast cancer, epidermal cancer, ovarian cancer, or esophageal cancer.

Images