Biologically active substance conjugates, methods for their preparation and uses

Abstract

Biologically active substance conjugates, methods for their preparation and uses thereof. The present invention relates to a ligand-drug conjugate represented by formula XV, methods for its preparation and uses thereof in the prevention and / or treatment of diseases associated with abnormal cellular activity, including but not limited to, in the prevention and / or treatment of tumor diseases. JPEG2024508081000723.jpg19128

Description

[Technical Field]

[0001] This application claims priority to Chinese Patent Application No. 2021101781361 filed on February 9, 2021, priority to Chinese Patent Application No. 2021103408065 filed on March 30, 2021, priority to Chinese Patent Application No. 202110825932X filed on July 21, 2021, priority to Chinese Patent Application No. 2021108259226 filed on July 21, 2021 (sequence listing in txt format is also available). This application claims priority to Chinese Patent Application No. 2021108259226 filed on July 21, 2021 (filed together with Chinese Patent Application No. 2021108259226 filed on July 21, 2021), and Chinese Patent Application No. 2021108259067 filed on July 21, 2021 (a sequence listing in txt format is also filed together with Chinese Patent Application No. 2021108259067 filed on July 21, 2021), the contents of all of which are incorporated herein by reference in their entireties. The sequence listing contained in this application is a part of the present specification and is incorporated herein by reference in its entirety.

[0002] The present disclosure belongs to the field of pharmaceutical technology and relates to biologically active substance conjugates (ligand drug conjugates), compounds, drug linker conjugates, and methods for their preparation and their use in the prevention and / or treatment of diseases associated with abnormal cellular activity, including, but not limited to, the prevention and / or treatment of tumor diseases. [Background technology]

[0003] Chemotherapy using cytotoxic agents was once the standard treatment for cancer, but these highly lethal cytotoxic molecules can kill normal cells and cause severe toxic side effects. Tumor-targeted drugs have become a hot topic in the field of cancer research due to their simultaneous tumor specificity and antitumor activity. However, issues such as target selectivity of targeted drugs often result in high toxicity and side effects, limiting the therapeutic efficacy of such drugs in many cases. Biologics such as antibodies or antibody fragments are highly target-selective but often have limited or no therapeutic efficacy against solid tumors. ADCs are conjugates of antibodies and small molecule drugs, combining the targeting effect of antibodies with the activity of bioactive molecules. ADCs have become biological missiles with promising advantages in efficacy and safety. Antibodies induce ADCs to bind to target cells, which are then internalized by the cells, and the small molecule drugs are then released intracellularly by enzymatic hydrolysis under the action of specific enzymes to treat the disease.

[0004] ADC drug research has developed rapidly in recent years, and 14 ADCs have been approved and marketed: (1) Mylotarg (gemtuzumab ozogamicin, gemtuzumab (CD33)-calicheamicin), approved by the US Food and Drug Administration in 2000 for CD33-positive acute myeloid leukemia (AML), withdrawn from the market in 2010, and reapproved in 2017; (2) Adcetris (brentuximab vedotin, CD30 monoclonal antibody-MMAE), approved by the US Food and Drug Administration in 2011 for typical Hodgkin lymphoma and anaplastic large cell lymphoma; ), (3) Kadcyla (trastuzumab emtansine, trastuzumab monoclonal antibody (Her2)-maytansine TM1), approved by the U.S. Food and Drug Administration in 2013 for Her2-positive breast cancer; (4) Besponsa (inotuzumab ozogamicin, CD22 monoclonal antibody-calicheamicin), approved in 2017 for adult relapsed or refractory B-cell lymphocytic leukemia; (5) Lumoxiti (moxetumomab pasudotox), marketed in 2018 for relapsed or refractory hairy cell leukemia (HCL). (pasudotox)-tdfk, CD22-Pseudomonas exotoxin), (6) Polivy (polatuzumab vedotin-piiq, CD79b-MMAE) for relapsed or refractory diffuse large B-cell lymphoma (R / R DLBCL), marketed in 2019, (7) Padcev (enfortumab vedotin-ejfv, Nectin4-MMAE) for locally advanced or metastatic urothelial carcinoma, marketed in 2019, (8) Enhertu (famtrastuzumab deruxtecan-nxki, Her2-topoisomerase I inhibitor) for breast cancer, marketed in 2019, (9) Trodelvy (Trop2-topoisomerase I inhibitor ADC) for triple-negative breast cancer, approved by the U.S. Food and Drug Administration in April 2020, (10) BCMA ADC Belantamab mafodotin (GSK2857916), (11) CD19 ADC Zynlonta (loncastuximab tesirin), (12) EGFR ADC Akalux, (13) Her2 ADC Disitamab vedotin RC48, (14) Tissue factor ADCTivadak (tisotumab vedotin).

[0005] ADC drugs consist of three components: an antibody, a biologically active molecule (drug molecule), and a linker. The biologically active molecule is covalently attached to the antibody via the linker. In addition to targeting and biological activity, the coupling mode between the biologically active molecule and the antibody plays an important role for ADC drugs, determining the homogeneity and stability of the drug and significantly affecting the pharmacokinetic properties and toxicity of the ADC. Improving the stability of the linking moiety between the biologically active molecule and the antibody improves the stability of the ADC in the systemic circulation, reduces drug release in non-target tissues, and thus relatively increases the amount of drug delivered to and around target cells, increasing the amount of biologically active molecule released into and around target cells, further enhancing efficacy and reducing toxicity. Therefore, increasing the stability of the bond between the biologically active molecule and the antibody is one of the most important technical aspects in ADC drug research.

[0006] Currently, there are two main methods for coupling antibodies and linkers for the 10 commercially available ADCs: (1) Lysine is the most common linking site in antibodies, and its ε-amino group can react with an activated carboxyl group on a linker to form an amide bond. Currently, there is a technology that can achieve this type of site-specific coupling: activating the carboxyl group on the linker with an activating group and then forming an amide bond with a specific lysine ε-amino group in the antibody to complete the coupling. However, site-specific quantitative coupling via this amide bond is not widely applicable and can only be used under certain conditions. Meanwhile, hydrolysis is prone to occur under the action of enzymes in vivo, resulting in dissociation of the bioactive molecule from the antibody before it reaches the target cell, thereby increasing toxicity. (2) Cysteine ​​thiols (SH) in antibodies exist in the form of disulfide bonds. Cleavage of disulfide bonds in antibodies can provide multiple free thiol groups as coupling sites. For coupling with thiols of antibodies, one such method is the Michael addition reaction between free thiols in antibodies and maleimides, or the double Michael addition reaction between specific substrates and free thiols in antibodies to form structurally unique sulfur bridges. However, there are reports in the literature that more than 30% of ADCs obtained by the thiol-Michael addition method can undergo retro-Michael addition in the systemic circulation, resulting in premature toxin release and toxic reactions.

[0007] Antibody drug conjugates can be classified into four generations according to their development history. First-generation antibody-drug conjugates are based on mouse-derived antibodies and often have strong toxic side effects due to their complex immunogenicity. Second-generation antibody-drug conjugates use humanized or fully human antibodies based on the first generation. The drug potency of second-generation antibody-drug conjugates has been significantly improved. The first two generations employed relatively random coupling between the antibody and the drug, resulting in a multi-component mixture of the final antibody-drug conjugate product. This type of drug product is not only toxic, but also difficult to control in terms of quality. Third-generation antibody-drug conjugates are based on the use of humanized or fully human antibodies, and simultaneously apply site-specific quantitative conjugation of toxin-linkers and antibodies, thus producing homogeneous single-molecule antibody-drug conjugates. The toxicity and quality control of these antibody-drug conjugates are significantly improved compared to the second generation. Fourth-generation antibody-drug conjugates represent an innovation in the toxin-linker moiety from the third generation. These conjugates have shifted from the strategy of using highly toxic toxins in previous antibody-drug conjugates to select the relatively less toxic camptothecin molecule as the bioactive component, while simultaneously using a high toxin-to-antibody ratio (DAR), e.g., a DAR value of 8. This type of ADC can also achieve good therapeutic efficacy against tumors with low tumor surface antigen expression. Despite a slow start, fourth-generation antibody-drug conjugates have rapidly gained clinical recognition due to their remarkable therapeutic efficacy. Representative drugs, Enhertu and Trodelvy, have been approved for sale by the U.S. Food and Drug Administration. However, fourth-generation ADCs still have problems such as low stability or solubility, making existing technologies lack versatility. Further development of new toxin linkers remains necessary to resolve these issues.

[0008] Existing antibody-drug conjugates function through the binding of the antibody in the ADC to a tumor cell surface antigen, then internalize into endosomes via endocytosis, and subsequently migrate from the endosome to lysosomes, where the bioactive molecule (toxin or payload) is then cleaved from the ADC by the action of lysosomal enzymes. The released bioactive molecule migrates from the lysosome to the cytoplasm to kill tumor cells. Bioactive molecules that escape after killing tumor cells can further kill nearby tumor cells that do not express or have low expression of their peripheral antigen through the so-called bystander effect. During the entire process, any changes during treatment, such as antigen presentation level, weakened or lost endocytosis, or loss of endosomal or lysosomal function, can render the ADC therapeutically resistant or ineffective. Therefore, it would be extremely important in clinical treatment to find an ADC that is stable in the systemic circulation in vivo and, at the same time, can kill tumor cells by releasing a toxin through extracellular cleavage in the tumor microenvironment without the need for endocytosis. This type of ADC may overcome the mechanisms associated with the multiple drug resistance problem of traditional ADC products.

[0009] Although there is evidence that Trodelvy from Immunomedics and Gilead can be cleaved outside tumor cells without endocytosis, Trodelvy is highly unstable in plasma and aqueous solutions, with a plasma half-life of less than 24 hours, and this type of ADC may only be suitable for those composed of toxins with low biological activity to avoid off-target toxic side effects caused by premature cleavage of the ADC toxin.

[0010] In addition, the functions of linkers in conventional ADCs are mainly lysosomal enzyme cleavage in cells or regulating water solubility. Therefore, how to use linkers to concentrate ADCs in tumor tissues while simultaneously maintaining their enzyme cleavability is also very important for reducing the toxic side effects of ADCs in normal tissues.

[0011] B7H3 is a type I transmembrane protein of the B7 family and has two isotypes in humans: 2Ig-B7H3 and 4Ig-B7H3. B7H3 is widely expressed in normal tissues, constitutively expressed in non-immune quiescent fibroblasts, endothelial cells, osteoblasts, and amniotic fluid stem cells, and inducibly expressed in activated T cells, NK cells, DCs, and macrophages, but no positive expression was detected in lymphoid organs (Chapoval, AI, et al., B7H3: A costimulatory molecule for T cell activation and IFN production. Nature Immunology, 2001.2(3):pp.269-274). Studies have shown that approximately 76.5% of patients with early-stage liver cancer have serum B7H3 levels approximately three times higher than normal individuals (60.79±19.45 vs. 20.52±8.46 ng / mL), but the correlation with disease progression is unclear (Zhao, L., et al., Early Detection of Hepatocellular Carcinoma in Patients with Hepatocirrhosis by Soluble B7H3. Journal of Gastrointestinal Surgery. 21(5):p.807-812).

[0012] Currently, therapeutic approaches targeting B7H3 are being studied in preclinical trials. For example, antibodies against mouse B7H3 enhance tumor-infiltrating CD8+ T cells and inhibit tumor growth (Mod. Pathol. 2010 Aug;23(8):1104-12). Additionally, WO 2008 / 066691 demonstrated that antibodies recognizing the B7H3 variant, B7H3a, have an in vivo antitumor effect against adenocarcinoma. In clinical studies, the combination of mouse B7H3 antibodies and radioactive iodine-131 significantly inhibited the growth of neuroblastoma in patients (J Neurooncol 97(3):409-18(2010)). Summary of the Invention

[0013] To improve the therapeutic efficacy of antibody drug conjugates (ADCs) or other ligand drug conjugates (PDCs), reduce drug toxicity and side effects, and expand the therapeutic window, the present disclosure provides a ligand drug conjugate of formula (XV) or a pharmaceutically acceptable salt or solvate thereof, which contains a biologically active molecule (drug molecule), a linker, and a targeting moiety linked to the linker via a reactive group (e.g., a sulfhydryl group) to form the ligand drug conjugate. The present disclosure also develops antibodies containing a variable light chain domain and / or a variable heavy chain (VH) domain that binds to a B7H3 molecule, which are derived from a human antibody.

[0014] Thus, in a first aspect of the present disclosure, the present disclosure provides a Ligand Drug Conjugate of Formula XV: [ka] During the ceremony, Tb is a ligand or targeting moiety that binds to the target; q is the drug-to-ligand coupling ratio, D is a biologically active molecular fragment; L1 is an extender unit, L2 is absent or a linking unit, and L3 is selected from an amino acid residue or a short peptide consisting of 2 to 10 amino acid residues; L4 is absent or present, and if present, L4 is [ka] wherein position 1 is attached to L3 and position 2 is attached to D, or a stereoisomer of the ligand drug conjugate, a prodrug thereof, a pharmaceutically acceptable salt thereof, or a pharmaceutically acceptable solvate thereof.

[0015] In addition, it should be noted that "position 1 of L1 is bound to Tb via an S atom" can be understood by those skilled in the art to mean that position 1 of L1 is bound to a sulfhydryl group contained in Tb after the disulfide bond is opened (e.g., reduction of the disulfide bond with a reducing agent, TCEP, can cleave the disulfide bond to generate a sulfhydryl -SH group). In other words, -S- between L1 and Tb is not an additional foreign sulfur atom.

[0016] for example, [ka] In the formula, -S- is not an additional foreign sulfur atom, but [ka] The -S- is formed by linking the sulfhydryl group contained in Tb to position 1 of L1 after opening the disulfide bond of the above.

[0017] The extender unit is a component of a ligand drug conjugate or drug linker conjugate or linker, and its function is to connect the ligand or targeting moiety that binds to the target to the rest of the ligand drug conjugate or linker. The extender unit can connect the Tb unit to either L2 (if present) or L3, and specific examples include: [ka] (wherein position 1 is attached to a ligand or targeting moiety that binds to a target, and position 2 is attached to L2 or L3).

[0018] In some embodiments, L1 is [ka] [ka] [ka] is selected from wherein each Z is independently selected from a direct bond, a carbon-carbon triple bond, a carbon-carbon double bond, a C6-10 aryl, a 5-10 membered heteroaryl, an amide, a sulfonamide, an imino, and a CF2; Rx and Ry are each independently selected from H and C1-4 alkyl; each m is independently selected from 0, 1, 2, 3, 4, 5, and 6; y1, y2, y3, and y4 are each independently selected from any integer of 0 to 20; Position 1 is bonded to Tb via the S atom, and position 2 is bonded to L2 or L3.

[0019] The linking unit is a component of the ligand drug conjugate or drug linker conjugate or linker, and its function is to attach the extender unit to an amino acid residue or a short peptide consisting of 2 to 10 amino acid residues. When a linking unit is present, it can attach L1 to L3. Specific examples include: [ka] (position 1 is attached to the extender unit and position 2 is attached to L3).

[0020] In some embodiments, L2 is absent or present, and when L2 is present, L2 is [ka] [ka] is selected from In the formula, y1, y2, y3, and y4 are each independently selected from any integer of 0 to 20, and the 1-position is bonded to L1, and the 2-position is bonded to L3.

[0021] In some embodiments, L1 is [ka] is.

[0022] In some embodiments, L1 is [ka] wherein each Z is independently selected from a direct bond, a carbon-carbon triple bond, a carbon-carbon double bond, a C6-10 aryl, a 5-10 membered heteroaryl, and an amide (preferably selected from a direct bond, a carbon-carbon triple bond, and a carbon-carbon double bond); Rx and Ry are each independently selected from H and a C1-4 alkyl; each m is independently selected from 0, 1, 2, 3, 4, 5, and 6; y1 is independently selected from any integer between 1 and 6 (e.g., 4, 5, 6); each y2 is independently selected from any integer between 0 and 15 (e.g., 6-15); each y3 is independently selected from 1, 2, and 3; and each y4 is independently selected from 0 and 1;

[0023] In some embodiments, L1 is [ka] wherein m is selected from 2, 3, and 4; y1 is selected from any integer from 1 to 6 (e.g., 4, 5, and 6); each y2 is independently selected from any integer from 0 to 10 (e.g., 6 to 10); and each y3 is independently selected from 1 and 2, wherein the 1-position is bonded to Tb via an S atom, and the 2-position is bonded to L2 or L3.

[0024] In some embodiments, L1 is [ka] wherein position 1 is bonded to Tb via the S atom and position 2 is bonded to L2 or L3.

[0025] In some embodiments, L1 is [ka] is selected from.

[0026] In some embodiments, L1 is [ka] wherein position 1 is bonded to Tb via the S atom and position 2 is bonded to L2 or L3.

[0027] In some embodiments, L1 is [ka] wherein position 1 is bonded to Tb via the S atom and position 2 is bonded to L2 or L3.

[0028] In some embodiments, L1 is [ka] wherein position 1 is bonded to Tb via the S atom and position 2 is bonded to L2 or L3.

[0029] In some embodiments, L2 is absent or present, and when L2 is present, L2 is [ka] wherein y1 is selected from any integer from 1 to 6 (e.g., 4, 5, 6), each y2 is independently selected from any integer from 0 to 10 (e.g., 6 to 10), each y3 is independently selected from 1 or 2, and each y4 is independently selected from 0 or 1, and position 1 is bonded to L1 and position 2 is bonded to L3.

[0030] In some embodiments, L2 is absent or present, and when L2 is present, L2 is [ka] wherein position 1 is attached to L1 and position 2 is attached to L3.

[0031] In some embodiments, L2 is absent or present, and when L2 is present, L2 is [ka] wherein position 1 is attached to L1 and position 2 is attached to L3.

[0032] In some embodiments, L2 is absent or present, and when L2 is present, L2 is [ka] wherein position 1 is attached to L1 and position 2 is attached to L3.

[0033] In some embodiments, L2 is absent.

[0034] In some embodiments, L2 is [ka] is selected from.

[0035] In some embodiments, L3 is selected from an amino acid residue or a short peptide consisting of 2 to 10 amino acid residues, wherein the amino acid residues are selected from natural amino acid residues, unnatural amino acid residues, or AA 1 or a stereoisomer thereof.

[0036] In some embodiments, L3 is selected from the group consisting of the amino acid residues Val, D-Val, Cit, Phe, Lys, Lys(Ac), Leu, Gly, Ala, Asn, Asp, Arg, and AA. 1 , or Val, Cit, Phe, Lys, D-Val, Leu, Gly, Ala, Asn, Asp, and AA 1The peptide is selected from short peptides consisting of 2 to 10 amino acid residues selected from the following:

[0037] In some embodiments, L3 is Val, Cit, Phe, Lys, D-Val, Leu, Gly, Ala, Asn, AA 1 , Val-Cit, Cit-Val, Cit-Ala, Val-Ala, Lys-Val, Val-Lys(Ac), Phe-Lys, Phe-Lys(Ac), Ala-Ala, Val-AA 1 , Ala-AA 1 , Gly-AA 1 , A.A. 1 -Gly, Ala-Ala-Ala, Ala-Ala-Asn, Ala-Ala-Asp, Val-AA 1 -Gly, Ala-AA 1 -Gly, Gly-AA 1 and Lys-Ala-Asn, Gly-Gly-Phe, D-Val-Leu-Lys, Gly-Gly-Arg, Ala-Ala-Asn, Gly-Gly-Phe, Val-Lys-Gly, Val-Lys-Gly-Gly, Val-Lys, and Lys-Ala-Asn.

[0038] In some embodiments, L3 is AA 1 , A.A. 1 -Gly, Val-Cit, Val-AA 1 -Gly, AA 1 -Ala-Asn, and Gly-Gly-Phe-Gly.

[0039] In some embodiments, L3 is AA 1 and Val-AA 1 -Gly.

[0040] In some embodiments, L3 is Val-AA 1 -Gly.

[0041] In some embodiments, L3 is [ka] [ka] is selected from In the formula, X - is a halide ion, a carboxylate ion, a sulfate ion, a hydrogen sulfate ion, and an OH - wherein position 1 is attached to L1 or L2 and position 2 is attached to L4 or D.

[0042] In some embodiments, L3 is [ka] wherein X is selected from - is a halide ion, a carboxylate ion, a sulfate ion, a hydrogen sulfate ion, and an OH - wherein position 1 is attached to L1 or L2 and position 2 is attached to L4 or D.

[0043] In some embodiments, L3 is [ka] wherein X is selected from - is a halide ion, a carboxylate ion, a sulfate ion, a hydrogen sulfate ion, and an OH - wherein position 1 is attached to L1 or L2 and position 2 is attached to L4 or D.

[0044] In some embodiments, L3 is [ka] wherein position 1 is attached to L1 or L2 and position 2 is attached to L4 or D.

[0045] In some embodiments, L3 is [ka] wherein position 1 is attached to L1 or L2 and position 2 is attached to L4 or D.

[0046] In some embodiments, AA 1 The structure of the amino acid residue represented by is as follows: [ka] During the ceremony, R a and R b are each independently H, [ka] wherein R a and R b Is it possible that both of them are H? Or R a and R b together with the carbon atoms to which they are both attached form a 4- to 10-membered heterocyclic ring, and the 4- to 10-membered heterocyclic ring is 0 is optionally replaced by r, r 1 are each independently selected from any integer of 0 to 20, R m1 , R n1 are each independently H, C1-6 alkyl, C3-6 cycloalkyl, or -COOR x1 is selected from R x1 is selected from C1-6 alkyl; Or R m1 and R n1 together with the nitrogen atom to which they are both attached form a 4- to 10-membered heterocyclic ring, and the 4- to 10-membered heterocyclic ring is 0’ is optionally replaced by R z is selected from C1-6 alkyl; R 0 , R 0’are each independently C1-6 alkyl, C3-6 cycloalkyl, -NR m2 R n2 and 4-10 membered heterocyclyl optionally substituted with C alkyl; R m2 , R n2 are each independently selected from H and C1-6 alkyl.

[0047] In some embodiments, R a and R b One of them is H and the other is [ka] is selected from.

[0048] In some embodiments, R a and R b One of them is H and the other is [ka] is selected from.

[0049] In some embodiments, R a and R b together with the carbon atom to which they are both attached, form R 0 Forms a 5- to 6-membered heterocyclic ring substituted with

[0050] In some embodiments, R a and R b together with the carbon atom to which they are both attached, form R 0 The piperidine or piperazine ring is formed by the following procedure.

[0051] In some embodiments, R a and R b together with the carbon atom to which they are both attached, form R 0 The resulting piperidine ring is substituted with

[0052] In some embodiments, R a and R b together with the carbon atom to which they are both attached, [ka] and the carbon atom numbered 1 is R a and R b is the carbon atom to which both are bonded.

[0053] In some embodiments, R a and R b together with the carbon atom to which they are both attached, [ka] and the carbon atom numbered 1 is R a and R b is the carbon atom to which both are bonded.

[0054] In some embodiments, r and r 1 are each independently selected from 0, 1, 2, 3, 4 and 5.

[0055] In some embodiments, r and r 1 are each independently selected from 0 and 4.

[0056] In some embodiments, r and r 1 One of them is 0 and the other is 4.

[0057] In some embodiments, R m1 and R n1 are each independently selected from H, methyl, ethyl, n-propyl, n-butyl, —COOCH, —COOCHCH, —COOCHCH, —COOCH(CH), —COOC(CH), and —COOCHCHCHCH.

[0058] In some embodiments, R m1 and Rn1 are each independently selected from H, C1-6 alkyl, C3-6 cycloalkyl, and tert-butoxycarbonyl.

[0059] In some embodiments, R m1 and R n1 are each independently selected from H and C1-6 alkyl.

[0060] In some embodiments, R m1 and R n1 are each independently selected from H, methyl, ethyl, and n-propyl.

[0061] In some embodiments, r and r 1 For r = 4, r 1 is 0 and R m1 and R n1 are each independently selected from H and C alkyl (e.g., H and CH), r is 0, and r 1 If is 4, R m1 and R n1 are each independently selected from C1-6 alkyl (eg, methyl, ethyl, n-propyl), preferably selected from C2-6 alkyl (eg, ethyl, n-propyl).

[0062] In some embodiments, R m1 and R n1 together with the nitrogen atom to which they are both attached, form R 0’ to form an optionally substituted 5- to 6-membered heterocycle.

[0063] In some embodiments, R m1 and R n1 together with the nitrogen atom to which they are both attached, form R 0’ to form an optionally substituted piperidine or piperazine ring.

[0064] In some embodiments, R m1 and R n1together with the nitrogen atom to which they are both attached, [ka] and the nitrogen atom numbered 1 is R m1 and R n1 are both bonded nitrogen atoms.

[0065] In some embodiments, R z is methyl.

[0066] In some embodiments, R 0 , R 0’ are each independently C1-6 alkyl, -NR m2 R n2 and 5-6 membered heterocyclyl optionally substituted with C1-6 alkyl.

[0067] In some embodiments, R 0 is selected from C1-6 alkyl and 5- to 6-membered heterocyclyl substituted with C1-6 alkyl, said 5- to 6-membered heterocyclyl being selected from piperidinyl and piperazinyl.

[0068] In some embodiments, R 0 is selected from methyl, ethyl, and 5- to 6-membered heterocyclyl substituted with methyl, wherein the 5- to 6-membered heterocyclyl is piperidinyl.

[0069] In some embodiments, R 0 is selected from methyl and 5- to 6-membered heterocyclyl substituted with methyl, and the 5- to 6-membered heterocyclyl is piperidinyl.

[0070] In some embodiments, R 0 is methyl, ethyl and [ka] is selected from.

[0071] In some embodiments, R 0 is methyl and [ka] is selected from.

[0072] In some embodiments, R 0’ is C1-6 alkyl and -NR m2 R n2 is selected from.

[0073] In some embodiments, R 0’ is methyl and -NR m2 R n2 is selected from.

[0074] In some embodiments, R m2 and R n2 is methyl.

[0075] In some embodiments, AA 1 The amino acid residue represented by [ka] is selected from.

[0076] In some embodiments, AA 1 The amino acid residue represented by [ka] is selected from.

[0077] In some embodiments, AA 1 The amino acid residue represented by [ka] is selected from.

[0078] In some embodiments, AA 1The amino acid residue represented by [ka] is selected from.

[0079] In some embodiments, L4 is absent or present, and when L4 is present, L4 is [ka] wherein position 1 is attached to L3 and position 2 is attached to D.

[0080] In some embodiments, L4 is absent or present, and when L4 is present, L4 is [ka] and position 1 is attached to L3 and position 2 is attached to D.

[0081] In some embodiments, L4 is absent.

[0082] In some embodiments, L4 is [ka] wherein position 1 is attached to L3 and position 2 is attached to D.

[0083] In some embodiments, L4 is [ka] wherein position 1 is attached to L3 and position 2 is attached to D.

[0084] In some embodiments, the structure: [ka] is selected from the following structural fragments: [Table 1]

[0085] In some embodiments, Tb is an antibody or an antigen-binding fragment thereof.

[0086] In some embodiments, antibodies or antigen-binding fragments thereof and monoclonal antibodies or antigen-binding fragments thereof include Fab, Fab', F(ab')2, Fd, Fv (e.g., scFv), dAb, complementarity-determining region fragments, non-human antibodies, humanized antibodies, chimeric antibodies, fully human antibodies, probodies, monoclonal antibodies, bispecific antibodies, or multispecific antibodies.

[0087] In some embodiments, Tb is an endocytic or non-endocytic antibody or antigen-binding fragment thereof.

[0088] In some embodiments, Tb is an endocytic antibody or antigen-binding fragment thereof.

[0089] In some embodiments, Tb is an antibody or antigen-binding fragment thereof that has activity of binding to free antigens in tumor tissue and / or tumor cell surface antigens.

[0090] In some embodiments, Tb is an antibody or antigen-binding fragment thereof that has tumor cell endocytosis activity and that has activity binding to free antigens in tumor tissue or tumor cell surface antigens.

[0091] In some embodiments, Tb is an antibody or antigen-binding fragment thereof that has tumor cell endocytosis activity and that has the activity of binding to free antigens in tumor tissue and tumor cell surface antigens.

[0092] In some embodiments, Tb is an antibody or antigen-binding fragment thereof that has tumor cell endocytosis activity but does not have the activity of binding to free antigens in tumor tissue.

[0093] In some embodiments, Tb is an antibody or antigen-binding fragment thereof that has tumor cell endocytosis activity and does not have tumor cell surface antigen binding activity.

[0094] In some embodiments, Tb is an antibody or antigen-binding fragment thereof that has no or weak endocytotic activity.

[0095] In some embodiments, Tb is an antibody or antigen-binding fragment thereof that does not have the activity of binding to free antigens in tumor tissue and / or tumor cell surface antigens.

[0096] In some embodiments, Tb is an antibody or antigen-binding fragment thereof that has no or weak tumor cell endocytosis activity and has the activity of binding to a free antigen in tumor tissue or a tumor cell surface antigen.

[0097] In some embodiments, Tb is an antibody or antigen-binding fragment thereof that has no or weak tumor cell endocytosis activity and has the activity of binding to free antigens in tumor tissue and tumor cell surface antigens.

[0098] In some embodiments, Tb is an antibody or antigen-binding fragment thereof that has no or weak tumor cell endocytosis activity and no activity to bind to tumor cell surface antigens. In some embodiments, Tb is an antibody or antigen-binding fragment thereof that has no or weak tumor cell endocytosis activity and no activity to bind to free antigens in tumor tissues.

[0099] In some embodiments, Tb is an antibody or antigen-binding fragment thereof that does not have tumor cell endocytosis activity and does not have a corresponding antigen in the human body.

[0100] In some embodiments, Tb is an antibody that does not have tumor cell-associated antigen binding.

[0101] In some embodiments, the antibody or antigen-binding fragment thereof is selected from IgG isotype antibodies.

[0102] In some embodiments, the antibody or antigen-binding fragment thereof is selected from isotype IgG1, isotype IgG2, isotype IgG3, or isotype IgG4. In some embodiments, the antibody or antigen-binding fragment thereof is an anti-chicken lysozyme human IgG1 isotype antibody.

[0103] In some embodiments, Tb is an antibody or antigen-binding fragment thereof that has activity of binding to a tumor cell surface non-endocytic antigen (eg, ALCAM / CD166).

[0104] In some embodiments, Tb is an antibody having tumor cell-associated antigen binding.

[0105] In some embodiments, Tb is an antibody or antigen-binding fragment thereof that has activity of binding to a free antigen in tumor tissue or a tumor cell surface antigen.

[0106] In some embodiments, Tb is an antibody or antigen-binding fragment thereof that has the activity of binding to free antigens in tumor tissue and tumor cell surface antigens.

[0107] In some embodiments, Tb is an antibody or antigen-binding fragment thereof that has B7H3-2Ig and / or B7H-4Ig.

[0108] In some embodiments, Tb is an antibody or antigen-binding fragment thereof that has higher binding activity to B7H3-4Ig than to B7H3-2Ig.

[0109] In some embodiments of the present disclosure, Tb is a ligand or targeting moiety that binds to a target.

[0110] In some embodiments, the target of Tb is selected from targets whose expression in tumor cells is higher than that in normal cells.

[0111] In some embodiments, the target of Tb is selected from targets that are highly expressed in tumor cells and low expressed in normal cells.

[0112] In some embodiments, the target of Tb is selected from B7H3, CD20, CD19, CD30, GPNMB, Her2, Trop-2, EGFR, Her3, GD-2, CD79b, BCMA, and the like.

[0113] In some embodiments, Tb is an antibody or an antigen-binding fragment thereof.

[0114] In some embodiments, Tb is a non-human antibody, a humanized antibody, a chimeric antibody, or a fully human antibody.

[0115] In some embodiments, Tb is a monoclonal antibody, a bispecific antibody, or a multispecific antibody.

[0116] In some embodiments, Tb is a monoclonal antibody or an antigen-binding fragment thereof.

[0117] In some embodiments, Tb is an anti-B7H3 antibody or an antigen-binding fragment thereof, an anti-Trop-2 antibody or an antigen-binding fragment thereof, an anti-Her2 antibody or an antigen-binding fragment thereof, an anti-Her3 antibody or an antigen-binding fragment thereof, or an anti-EGFR antibody or an antigen-binding fragment thereof.

[0118] In some embodiments, Tb is an anti-B7H3 antibody or an antigen-binding fragment thereof, such as antibody 1D1, antibody 1D1-01, antibody 2E3, antibody 2E3-02, enoblituzumab, mirzotamab, omburtamab, or an antigen-binding fragment thereof.

[0119] The sequence of 1D1 is as set forth in SEQ ID NO: 1. The VH sequence of 1D1-01 is as set forth in SEQ ID NO: 3, and the VL sequence is as set forth in SEQ ID NO: 13. The heavy chain sequence of 1D1-01 is as set forth in SEQ ID NO: 45, and the light chain sequence is as set forth in SEQ ID NO: 46. The sequence of 2E3 is as set forth in SEQ ID NO: 2. The VH sequence of 2E3-02 is as set forth in SEQ ID NO: 23, and the VL sequence is as set forth in SEQ ID NO: 33. The heavy chain sequence of 2E3-02 is as set forth in SEQ ID NO: 47, and the light chain sequence is as set forth in SEQ ID NO: 48.

[0120] In some embodiments, Tb is an anti-B7H3 monoclonal antibody or an antigen-binding fragment thereof.

[0121] In some embodiments, Tb is an anti-Trop-2 antibody or an antigen-binding fragment thereof, such as datopotamab, sacituzumab, or an antigen-binding fragment thereof.

[0122] In some embodiments, Tb is an anti-Trop-2 monoclonal antibody or an antigen-binding fragment thereof.

[0123] In some embodiments, Tb is an anti-Her2 antibody or an antigen-binding fragment thereof, such as anbenitamab, coprelotamab, dicitamab, gancotamab, margetuximab, pertuzumab, timigutuzumab, zanidatamab, trastuzumab, pertuzumab, or an antigen-binding fragment thereof.

[0124] In some embodiments, Tb is an anti-Her2 monoclonal antibody or an antigen-binding fragment thereof, such as trastuzumab, pertuzumab, or an antigen-binding fragment thereof.

[0125] In some embodiments, Tb is an anti-Her3 antibody or an antigen-binding fragment thereof, such as barecetamab, durigotuzumab, elgemtumab, istiratumab, lanretuzumab, patritumab, seribantumab, zenocutuzumab, antibody 202-2-1, or an antigen-binding fragment thereof.

[0126] In some embodiments, Tb is an anti-Her3 monoclonal antibody or an antigen-binding fragment thereof.

[0127] In some embodiments, Tb is an anti-EGFR antibody or an antigen-binding fragment thereof, such as demupitamab, depatuxizumab, futuximab, imatuzumab, laprituximab, losatuxizumab, matuzumab, modotuximab, necitumumab, nimotuzumab, panitumumab, pimurutamab, serclutamab, tomuzotuximab, zalutumumab, cetuximab, or an antigen-binding fragment thereof.

[0128] In some embodiments, Tb is an anti-EGFR monoclonal antibody or an antigen-binding fragment thereof.

[0129] In some embodiments, the antibody has antigen-binding activity but does not have endocytic activity, ie, an antibody has antigen-binding activity such as B7H3, CD20, CD19, CD30, GPNMB, Her2, Trop-2, EGFR, Her3, GD-2, CD79b, and BCMA, but does not have endocytic activity.

[0130] In some embodiments, the antibody has binding activity to the antigen B7H3 but does not have endocytic activity, such as INV721 and I7-01 disclosed in WO2021168379A1. More specifically, the anti-B7H3 antibody has a VH of SEQ ID NO:2 and a VL of SEQ ID NO:1, as described in WO2021168379A1.

[0131] In some embodiments, the antibody has binding activity to the antigen GD2 but does not have endocytic activity, such as INV721 and GD2-5 disclosed in WO2021168379A1. More specifically, the anti-GD2 antibody has a VH of SEQ ID NO:4 and a VL of SEQ ID NO:3, as described in WO2021168379A1.

[0132] In some embodiments, the antibody has activity of binding to the antigen HER3 and does not have endocytic activity, such as the anti-HER3 antibody 21F06 set forth in SEQ ID NO: 22 in US Pat. No. 10,808,032 B2.

[0133] In some embodiments, the antibody has binding activity to the antigen CD20 but does not have endocytic activity. It has been shown that so-called "type II" CD20-specific antibodies are difficult to internalize by CD20-positive target cells, whereas other so-called "type I" CD20-specific antibodies have been found to be internalized and degraded to some extent, depending on the activation and inhibition of FcγR expression levels on the target cells they interact with. In some embodiments, the antibody has binding activity to the antigen CD20 but does not have endocytic activity is a "type II" CD20-specific antibody, such as obinutuzumab.

[0134] In some embodiments, the antibody is an antibody that has binding activity to a non-endocytic antigen (e.g., ALCAM / CD166). In some embodiments, the antibody is an antibody comprising a VH of SEQ ID NO: 73 and a VL of SEQ ID NO: 74, a VH of SEQ ID NO: 75 and a VL of SEQ ID NO: 76, a VH of SEQ ID NO: 77 and a VL of SEQ ID NO: 78, or a VH of SEQ ID NO: 79 and a VL of SEQ ID NO: 88, as described in EP 3911682A1.

[0135] In some embodiments, Tb is a targeting moiety, such as a ligand, a protein, a polypeptide, a non-protein reagent (eg, a sugar, RNA or DNA), an antibody mimic, or the like.

[0136] In some embodiments, q is selected from any number between 0.1 and 16.0. In a preferred embodiment, q is selected from any integer between 0.1 and 16.0.

[0137] In some embodiments, q is selected from any number between 0.1 and 8.0. In a preferred embodiment, q is selected from any integer between 0.1 and 8.0.

[0138] In some embodiments, q is selected from any number between 2 and 8.

[0139] In some embodiments, q is selected from any number between 3 and 8.

[0140] In some embodiments, q is selected from any number between 4 and 8.

[0141] In some embodiments, q is selected from any number between 6 and 8.

[0142] In some embodiments, q is selected from any integer from 2 to 8.

[0143] In some embodiments, q is selected from any integer from 3 to 8.

[0144] In some embodiments, q is selected from any integer from 4 to 8.

[0145] In some embodiments, q is selected from any integer from 6 to 8.

[0146] In some embodiments, q is selected from 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, and 12.

[0147] In some embodiments, q is selected from 2, 4, 6, and 8.

[0148] In the present disclosure, a biologically active molecular fragment refers to a moiety (fragment or group) in an antibody-drug conjugate (ADC, or referred to as an antibody conjugate drug) that can form a biologically active drug (e.g., a small molecule cytotoxic drug comprising the group after loss of an atom or group of atoms) or a derivative thereof (e.g., a precursor thereof) after cleavage / degradation / enzymatic cleavage of the linker in tumor tissue or tumor cells, as is well known in the art. For the avoidance of ambiguity, "drug" not only means a "drug product" approved by drug regulatory authorities, but also includes any molecule with potential therapeutic biological activity in clinical practice or research and development as well as academic research.

[0149] In some embodiments, D is a molecular fragment that has anti-tumor biological activity.

[0150] In some embodiments, D is a molecular fragment having anti-tumor biological activity, and the biologically active molecule is selected from a cytotoxic agent or a derivative thereof, for example, a DNA topoisomerase inhibitor (e.g., a camptothecin-type biologically active molecule such as camptothecin, DXD, camptothecin with modified substituents, or DXD with modified substituents) or a tubulin inhibitor (e.g., an MMAF-type tubulin inhibitor, an MMAE-type tubulin inhibitor).

[0151] In some embodiments, the Ligand Drug Conjugate has a structure represented by Formula I: [ka] During the ceremony, R1, R2 are each independently selected from H, halogen, —OH, optionally substituted C1-6 alkyl, and optionally substituted C1-6 alkoxy; or R1 and R2, together with the carbon atoms to which they are attached, form a 5- to 7-membered carbocyclic or heterocyclic ring containing one or more O, S, N, carbonyl, sulfoxide or sulfone groups, or any combination thereof; R3 is selected from H, halogen, —OH, —NH2, optionally substituted C1-6 alkyl and optionally substituted C1-6 alkoxy; or R3 and X together with the carbon atoms to which they are attached form a 5- to 7-membered carbocyclic or heterocyclic ring containing one or more O, S, N, carbonyl, sulfoxide or sulfone groups or any combination thereof; or R3 and R2, together with the carbon atoms to which they are attached, form a 5- to 7-membered carbocyclic or heterocyclic ring containing one or more O, S, N, carbonyl, sulfoxide or sulfone groups, or any combination thereof; W is absent or present, and when W is present, W is -O-, -S-, -NR4-, [ka] wherein position 1 is bonded to X and position 2 is bonded to L4 or L3; X is a direct bond, an optionally substituted -O-(CH2) n3 -, -NR4-(CH2) n3 -, -S-(CH2) n3 -, carbonyl-(CH2) n3 -, -SO2-(CH2) n3 -, [ka] -(CH2) n1 -, C3-6 cycloalkyl, C6-10 aryl, 5-10 membered heteroaryl and 4-10 membered heterocyclyl, wherein position 1 is bonded to the parent ring and position 2 is bonded to W or L4, and the substituents are selected from one or more C1-4 alkyl groups, C3-6 cycloalkyl groups, or multiple C1-4 alkyl groups together with the carbon atoms to which they are both attached form a C3-6 cycloalkyl group; Each M independently represents a direct bond and -CR 5a R 5b - selected from R4, R5, R 5a , R 5b , R6, R7 are each independently selected from H, optionally substituted C1-4 alkyl, optionally substituted C1-4 alkoxy, and optionally substituted C3-6 cycloalkyl; n, n', n1, n2, and n3 are each independently selected from any integer of 0 to 6; L4 is absent or present, and if present, L4 is [ka] wherein position 1 is bonded to L3 and position 2 is bonded to W or X.

[0152] Tb, L1, L2, L3 and q are as defined above and in any embodiment specifically described herein; In some embodiments, R1 and R2 are each independently selected from H, halogen, and C1-4 alkyl.

[0153] In some embodiments, R1 and R2 together with the carbon atoms to which they are attached form a 5-6 membered heterocycle containing 1, 2, or 3 O, S, or N, or any combination thereof.

[0154] In some embodiments, R1 is selected from H and halogen, and R2 is selected from H and C1-4 alkyl.

[0155] In some embodiments, R1 and R2, together with the carbon atom to which they are attached, [ka] where the dotted line indicates where the heterocycle is fused to the benzene ring.

[0156] In some embodiments, R1 is H or F and R2 is H or methyl.

[0157] In some embodiments, R1 is F and R2 is methyl, or R1 and R2 together with the carbon atom to which they are attached are: [ka] Form.

[0158] In some embodiments, R1 is F and R2 is methyl.

[0159] In some embodiments, R1 and R2, together with the carbon atom to which they are attached, [ka] Form.

[0160] In some embodiments, R3 is selected from H and C1-4 alkyl.

[0161] In some embodiments, R3 and X together with the carbon atoms to which they are attached form a 5- to 6-membered carbocyclic ring.

[0162] In some embodiments, R3 is H, or R3 and X together with the carbon atom to which they are attached are [ka] where the dotted lines indicate where the carbocyclic ring is fused to the benzene and pyridine rings.

[0163] In some embodiments, R3 is H.

[0164] In some embodiments, W is absent or present, and when W is present, W is -O-, -S-, -NR4-, [ka] wherein position 1 is bonded to X and position 2 is bonded to L4 or L3.

[0165] In some embodiments, W is absent or present, and when W is present, W is -O-, -S-, -NR4-, [ka] wherein position 1 is bonded to X and position 2 is bonded to L4 or L3.

[0166] In some embodiments, W is —O—, —NR4—, and [ka] wherein position 1 is bonded to X and position 2 is bonded to L4 or L3.

[0167] In some embodiments, W is selected from -O- and -NR4-, position 1 is bonded to X, and position 2 is bonded to L4 or L3.

[0168] In some embodiments, X is optionally substituted -(CH) n1 -, [ka] is selected from C6-10 aryl, 5-10 membered heteroaryl and 4-10 membered heterocyclyl, wherein position 1 is bonded to the parent ring and position 2 is bonded to W or L4, and the substituents are selected from one or two C1-4 alkyl groups, or two C1-4 alkyl groups together with the carbon atom to which they are both attached form a C3-6 cycloalkyl group; In some embodiments, X is optionally substituted [ka] wherein position 1 is attached to the parent ring and position 2 is attached to W or L4, and the substituents are selected from one or two C1-4 alkyl groups (e.g., methyl), or two C1-4 alkyl groups (e.g., methyl) taken together with the carbon atom to which they are both attached form a C3-6 cycloalkyl group (e.g., cyclopropyl).

[0169] In some embodiments, X is [ka] wherein position 1 is attached to the parent ring and position 2 is attached to W or L4.

[0170] In some embodiments, X is [ka] wherein position 1 is attached to the parent ring and position 2 is attached to W.

[0171] In some embodiments, when W is absent, X is [ka] wherein position 1 is attached to the parent ring and position 2 is attached to L4; and when W is present, X is selected from: [ka] wherein position 1 is attached to the parent ring and position 2 is attached to W.

[0172] In some embodiments, W is —O—, —NR4—, and [ka] wherein position 1 is bonded to X and position 2 is bonded to L4 or L3, and X is selected from: [ka] wherein position 1 is attached to the parent ring and position 2 is attached to W.

[0173] In some embodiments, R4, R5 are each independently selected from H, C1-4 alkyl, and C3-6 cycloalkyl.

[0174] In some embodiments, each R4 is independently selected from H, C1-4 alkyl, and C3-6 cycloalkyl; and R5 is H.

[0175] In some embodiments, each R4 is independently selected from H, methyl, ethyl, n-propyl, isopropyl, t-butyl, and cyclopropyl; and R5 is H.

[0176] In some embodiments, R 5a , R 5b are each independently selected from H and C1-4 alkyl.

[0177] In some embodiments, R 5a , R 5b are each independently selected from H and methyl.

[0178] In some embodiments, each R7 is independently selected from H and C1-4 alkyl.

[0179] In some embodiments, R7 is H.

[0180] In some embodiments, n is selected from 1, 2, and 3.

[0181] In some embodiments, n is 1.

[0182] In some embodiments, n1 is selected from 1, 2, 3, and 4.

[0183] In some embodiments, n2 is 1.

[0184] In some embodiments, n3 is 0.

[0185] In some embodiments, L3 is [ka] wherein X is selected from - is a halide ion, a carboxylate ion, a sulfate ion, a hydrogen sulfate ion, and an OH - wherein position 1 is attached to L1 or L2 and position 2 is attached to L4 or W.

[0186] In some embodiments, L3 is [ka] wherein X is selected from - is a halide ion, a carboxylate ion, a sulfate ion, a hydrogen sulfate ion, and an OH - wherein position 1 is attached to L1 or L2 and position 2 is attached to L4 or W.

[0187] In some embodiments, L3 is [ka] wherein X is selected from - is a halide ion, a carboxylate ion, a sulfate ion, a hydrogen sulfate ion, and an OH -wherein position 1 is attached to L1 or L2 and position 2 is attached to L4 or W.

[0188] In some embodiments, L3 is [ka] wherein position 1 is attached to L1 or L2 and position 2 is attached to L4 or W.

[0189] In some embodiments, L3 is [ka] wherein position 1 is attached to L1 or L2 and position 2 is attached to L4 or W.

[0190] In some embodiments, L4 is absent or present, and when L4 is present, L4 is [ka] wherein position 1 is bonded to L3 and position 2 is bonded to W or X.

[0191] In some embodiments, L4 is absent or present, and when L4 is present, L4 is [ka] wherein position 1 is bonded to L3 and position 2 is bonded to W or X.

[0192] In some embodiments, L4 is absent.

[0193] In some embodiments, L4 is [ka] wherein position 1 is bonded to L3 and position 2 is bonded to W or X.

[0194] In some embodiments, L4 is [ka] wherein position 1 is bonded to L3 and position 2 is bonded to W or X.

[0195] As described above, W is either absent or present, and therefore, when W is absent, position 1 of L4 is bonded to L3 and position 2 is bonded to X, and when W is present, position 1 of L4 is bonded to L3 and position 2 is bonded to W. The following bonding relationships for L4 can be understood with reference to the above.

[0196] In some embodiments, the structure: [ka] is selected from the following structural fragments: [Table 2-1] [Table 2-2] [Table 2-3] In the formula, position 1 is bonded to Tb and position 2 is bonded to W.

[0197] In some embodiments, D is [ka] and position 1 is linked to L3 or L4, e.g. [ka] is.

[0198] In some embodiments, the structure: [ka] is selected from the following structural fragments: [ka] [ka] [ka] wherein position 1 is bonded to L4, and if L4 is absent, position 1 is bonded to L3.

[0199] In some embodiments, W is absent or present, and when W is present, W is -O-, -S-, -NR4-, [ka] is selected from, for example, absent, -O-, -NR4- or [ka] wherein R4 and R5 are each independently selected from H and C1-4 alkyl, and n is independently selected from 0, 1, 2, 3, and 4; X is [ka] is selected from wherein R1 is selected from H and halogen and R2 is selected from H and C1-4 alkyl, or R1 and R2 together with the carbon atom to which they are attached are [ka] where the dotted line indicates where the heterocycle is fused to the benzene ring; R3 is H and C1-4 alkyl, or R3 and X together with the carbon atoms to which they are attached form a 5- to 6-membered carbocyclic ring; Preferably, W is absent or present, and when W is present, W is -O-, -NR4- (e.g., -NH-, -N(CH3)-, -N(C2H5)-), [ka] wherein R4 is independently selected from H, methyl, ethyl, isopropyl, n-propyl, t-butyl, and cyclopropyl; If W is not present, then X is [ka] where position 1 is attached to the parent ring and when W is present, X is selected from: [ka] is selected from In AA1, [ka] r is selected from 0, 1, 2, 3, 4 and 5; R a and R b One of them is H and the other is [ka] or R a and R b together with the carbon atom to which they are both attached, form R 0 forming a 5- to 6-membered heterocyclic ring substituted with R1 is selected from H and halogen and R2 is selected from H and C1-4 alkyl, or R1 and R2 together with the carbon atom to which they are attached are [ka] Forming R3 is H and C1-4 alkyl, or R3 and X together with the carbon atom to which they are attached are [ka] Forming More preferably, W is selected from -O- and -NR4-, where position 1 is bonded to X and position 2 is bonded to L4 or L3; X is [ka] wherein position 1 is attached to the parent ring and position 2 is attached to W; [ka] For example, [ka] and R1 is F, R2 is methyl, or R1 and R2 together with the carbon atom to which they are attached are [ka] Forming R3 is H.

[0200] In some embodiments, the Ligand Drug Conjugate has the structure represented by Formula I-1: [ka] and wherein Tb, L1, L2, L3, L4, X, R1, R2, R3, R4, and q are as defined above and in any embodiment specifically described herein.

[0201] In some embodiments, the Ligand Drug Conjugate has the structure of Formula I-1A or Formula I-1B: [ka] wherein Tb, L2, L3, L4, X, R1, R2, R3, R4, and q are as defined above and in any embodiment specifically described herein.

[0202] In some embodiments, the Ligand Drug Conjugate has a structure represented by Formula I-2: [ka] wherein Tb, L1, L2, L3, L4, X, R1, R2, R3, and q are as defined above and in any embodiment specifically described herein.

[0203] In some embodiments, the Ligand Drug Conjugate has the structure of Formula I-2A or Formula I-2B: [ka] and wherein Tb, L2, L3, L4, X, R1, R2, R3, and q are as defined above and in any embodiment specifically described herein.

[0204] In some embodiments, the Ligand Drug Conjugate has a structure represented by Formula I-3: [ka] wherein Tb, L1, L2, L3, L4, X, R1, R2, R3, R4, R5, n and q are as defined above and in any embodiment specifically described herein.

[0205] In some embodiments, the Ligand Drug Conjugate has the structure of Formula I-3A or Formula I-3B: [ka] and wherein Tb, L2, L3, L4, X, R1, R2, R3, R4 and q are as defined above and in any embodiment specifically described herein.

[0206] In some embodiments, the Ligand Drug Conjugate has the structure represented by Formula IA: [ka] and In the formula, Tb, X, R1, R2, R3, R a , R b and q are as defined above and in any embodiment specifically described herein.

[0207] In some embodiments, the Ligand Drug Conjugate has the structure represented by Formula IB: [ka] and In the formula, Tb, X, R1, R2, R3, R a , R b and q are as defined above and in any embodiment specifically described herein.

[0208] In some embodiments, the Ligand Drug Conjugate is selected from the following: [Table 3-1] [Table 3-2] [Table 3-3] [Table 3-4] [Table 3-5] [Table 3-6] [Table 3-7] [Table 3-8] [Table 3-9] [Table 3-10] [Table 3-11] [Table 3-12] [Table 3-13] [Table 3-14] [Table 3-15] [Table 3-16] [Table 3-17] [Table 3-18] [Table 3-19] The Tb antibody or its antigen-binding fragment can be prepared by various methods known in the art, for example, by recombinant genetic engineering. For example, DNA molecules encoding the heavy and light chain genes of the antibody of the present disclosure can be obtained by chemical synthesis or PCR amplification. The obtained DNA molecules are inserted into an expression vector and then transfected into host cells. The transfected host cells are then cultured under specific conditions to express the antibody of the present disclosure.

[0209] In some embodiments of the present disclosure, the targeting moiety is an anti-B7H3 antibody or an antigen-binding fragment thereof. In some embodiments, the anti-B7H3 antibody includes all prior art anti-B7H3 antibodies, such as enoblituzumab, mirzotamab, and omburtamab, as well as those disclosed in CN112521512, WO2021027674, WO2021021543, WO2021006619, Chinese Patent No. 111662384 ... National Patent No. 111454357, International Publication No. 2020151384, International Publication No. 2020140094, International Publication No. 2020103100, International Publication No. 2020102779, International Publication No. 2020063673, International Publication No. 2020047257, International Publication No. 2020041626, Chinese Patent No. 110684790, Chinese Patent No. 10642948, International Publication No. 2019225787, International Publication No. 2019226017, U.S. Patent No. 20190338030, Chinese Patent No. 110305213, International Publication No. 2018209346, International Publication No. 2018177393, U.S. Patent No. 9150656, International Publication No. 2016106004, International Publication No. 20160443 83, WO 2016033225, WO 2015181267, U.S. Patent No. 20120294796, WO 2011109400, Chinese Patent No. 101104639, WO 2004093894, WO 2002010187, and WO 2001018021. In some embodiments, the anti-B7H3 antibody or antigen-binding fragment thereof is selected from antibody M30-H1-L4 in Chinese Patent No. 103687945B and antibody mAb-C-DUBA in Chinese Patent No. 109069633A.

[0210] In some embodiments, the anti-B7H3 antibody or antigen-binding fragment thereof is an antibody or antigen-binding fragment thereof that comprises the complementarity-determining regions (CDRs) shown below, wherein the CDRs are numbered according to the IMGT numbering system: (a) an HCDR1 consisting of the sequence of SEQ ID NO: 10, an HCDR2 consisting of the sequence of SEQ ID NO: 11, an HCDR3 consisting of the sequence of SEQ ID NO: 12, and / or LCDR1 consisting of the sequence of SEQ ID NO: 20, LCDR2 consisting of the sequence of GTF, and LCDR3 consisting of the sequence of SEQ ID NO: 22; (b) an HCDR1 consisting of the sequence of SEQ ID NO: 10, an HCDR2 consisting of the sequence of SEQ ID NO: 11, an HCDR3 consisting of the sequence of SEQ ID NO: 12, and / or LCDR1 consisting of the sequence of SEQ ID NO: 40, LCDR2 consisting of the sequence of GAS, and LCDR3 consisting of the sequence of SEQ ID NO: 42; (c) an HCDR1 consisting of the sequence of SEQ ID NO: 30, an HCDR2 consisting of the sequence of SEQ ID NO: 31, an HCDR3 consisting of the sequence of SEQ ID NO: 32, and / or LCDR1 consisting of the sequence of SEQ ID NO: 40, LCDR2 consisting of the sequence of GAS, and LCDR3 consisting of the sequence of SEQ ID NO: 42; or (d) an HCDR1 consisting of the sequence of SEQ ID NO: 30, an HCDR2 consisting of the sequence of SEQ ID NO: 31, an HCDR3 consisting of the sequence of SEQ ID NO: 32, and / or It is defined by LCDR1 consisting of the sequence of SEQ ID NO:20, LCDR2 consisting of the sequence of GTF, and LCDR3 consisting of the sequence of SEQ ID NO:22.

[0211] In some embodiments, the anti-B7H3 antibody or antigen-binding fragment thereof as defined by the IMGT numbering system is: (a) a VH comprising a heavy chain HCDR1, HCDR2, and HCDR3, and (a) a VL comprising a light chain LCDR1, LCDR2, and LCDR3; (b) a VH comprising a heavy chain HCDR1, HCDR2, and HCDR3 of (b), and (b) a light chain VL comprising LCDR1, LCDR2, and LCDR3 of (b); (c) a VH comprising a heavy chain HCDR1, HCDR2, and HCDR3, and (c) a VL comprising a light chain LCDR1, LCDR2, and LCDR3; or (d) VH comprising the heavy chain HCDR1, HCDR2, and HCDR3 of (d), and (d) VL comprising the light chain LCDR1, LCDR2, and LCDR3 of (d).

[0212] In some embodiments, the anti-B7H3 antibody or antigen-binding fragment thereof comprises the complementarity determining regions (CDRs) shown below, wherein the CDRs are numbered according to the Chothia numbering system: (a) an HCDR1 consisting of the sequence of SEQ ID NO: 4, an HCDR2 consisting of the sequence of SEQ ID NO: 5, an HCDR3 consisting of the sequence of SEQ ID NO: 6, and / or LCDR1 consisting of the sequence of SEQ ID NO: 14, LCDR2 consisting of the sequence of SEQ ID NO: 15, and LCDR3 consisting of the sequence of SEQ ID NO: 16; (b) an HCDR1 consisting of the sequence of SEQ ID NO: 24, an HCDR2 consisting of the sequence of SEQ ID NO: 25, an HCDR3 consisting of the sequence of SEQ ID NO: 26, and / or LCDR1 consisting of the sequence of SEQ ID NO: 34, LCDR2 consisting of the sequence of SEQ ID NO: 35, and LCDR3 consisting of the sequence of SEQ ID NO: 36; (c) HCDR1 consisting of the sequence of SEQ ID NO: 4, HCDR2 consisting of the sequence of SEQ ID NO: 5, HCDR3 consisting of the sequence of SEQ ID NO: 6, and / or LCDR1 consisting of the sequence of SEQ ID NO: 34, LCDR2 consisting of the sequence of SEQ ID NO: 35, and LCDR3 consisting of the sequence of SEQ ID NO: 36; or (d) an HCDR1 consisting of the sequence of SEQ ID NO: 24, an HCDR2 consisting of the sequence of SEQ ID NO: 25, an HCDR3 consisting of the sequence of SEQ ID NO: 26, and / or It is defined by LCDR1 consisting of the sequence of SEQ ID NO:14, LCDR2 consisting of the sequence of SEQ ID NO:15, and LCDR3 consisting of the sequence of SEQ ID NO:16.

[0213] In some embodiments, the anti-B7H3 antibody or antigen-binding fragment thereof as defined by the Chothia numbering system is: (a) a VH comprising a heavy chain HCDR1, HCDR2, and HCDR3, and (a) a VL comprising a light chain LCDR1, LCDR2, and LCDR3; (b) a VH comprising a heavy chain HCDR1, HCDR2, and HCDR3 of (b), and (b) a light chain VL comprising LCDR1, LCDR2, and LCDR3 of (b); (c) a VH comprising a heavy chain HCDR1, HCDR2, and HCDR3, and (c) a VL comprising a light chain LCDR1, LCDR2, and LCDR3; or (d) VH comprising the heavy chain HCDR1, HCDR2, and HCDR3 of (d), and (d) VL comprising the light chain LCDR1, LCDR2, and LCDR3 of (d).

[0214] In some embodiments, the anti-B7H3 antibody or antigen-binding fragment thereof comprises the complementarity determining regions (CDRs) shown below, where the CDRs are numbered according to the Kabat numbering system: (a) an HCDR1 consisting of the sequence of SEQ ID NO: 7, an HCDR2 consisting of the sequence of SEQ ID NO: 8, an HCDR3 consisting of the sequence of SEQ ID NO: 9, and / or LCDR1 consisting of the sequence of SEQ ID NO: 17, LCDR2 consisting of the sequence of SEQ ID NO: 18, and LCDR3 consisting of the sequence of SEQ ID NO: 19; (b) an HCDR1 consisting of the sequence of SEQ ID NO: 27, an HCDR2 consisting of the sequence of SEQ ID NO: 28, an HCDR3 consisting of the sequence of SEQ ID NO: 29, and / or LCDR1 consisting of the sequence of SEQ ID NO: 37, LCDR2 consisting of the sequence of SEQ ID NO: 38, and LCDR3 consisting of the sequence of SEQ ID NO: 39; (c) HCDR1 consisting of the sequence of SEQ ID NO: 7, HCDR2 consisting of the sequence of SEQ ID NO: 8, HCDR3 consisting of the sequence of SEQ ID NO: 9, and / or LCDR1 consisting of the sequence of SEQ ID NO: 37, LCDR2 consisting of the sequence of SEQ ID NO: 38, and LCDR3 consisting of the sequence of SEQ ID NO: 39; or (d) an HCDR1 consisting of the sequence of SEQ ID NO: 27, an HCDR2 consisting of the sequence of SEQ ID NO: 28, an HCDR3 consisting of the sequence of SEQ ID NO: 29, and / or It is defined by LCDR1 consisting of the sequence of SEQ ID NO:17, LCDR2 consisting of the sequence of SEQ ID NO:18, and LCDR3 consisting of the sequence of SEQ ID NO:19.

[0215] In some embodiments, the anti-B7H3 antibody or antigen-binding fragment thereof as defined by the Kabat numbering system is: (a) a VH comprising a heavy chain HCDR1, HCDR2, and HCDR3, and (a) a VL comprising a light chain LCDR1, LCDR2, and LCDR3; (b) a VH comprising a heavy chain HCDR1, HCDR2, and HCDR3 of (b), and (b) a light chain VL comprising LCDR1, LCDR2, and LCDR3 of (b); (c) a VH comprising a heavy chain HCDR1, HCDR2, and HCDR3, and (c) a VL comprising a light chain LCDR1, LCDR2, and LCDR3; or (d) VH comprising the heavy chain HCDR1, HCDR2, and HCDR3 of (d), and (d) VL comprising the light chain LCDR1, LCDR2, and LCDR3 of (d).

[0216] In some embodiments, the anti-B7H3 antibody or antigen-binding fragment thereof is (a) Below: (i) a sequence set forth in SEQ ID NO: 3 or 23; (ii) a sequence having one or more amino acid substitutions, deletions, or additions, or any combination thereof (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acid substitutions, deletions, or additions, or any combination thereof), compared to the sequence set forth in SEQ ID NO: 3 or 23; or (iii) a heavy chain variable region (VH) comprising an amino acid sequence selected from the group consisting of a sequence having at least 70%, at least 75%, 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%, or at least 99% sequence identity compared to the sequence set forth in SEQ ID NO: 3 or 23; and / or (b) Below: (iv) a sequence set forth in SEQ ID NO: 13 or 33; (v) a sequence having one or more amino acid substitutions, deletions, or additions, or any combination thereof, compared to the sequence set forth in SEQ ID NO: 13 or 33 (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acid substitutions, deletions, or additions, or any combination thereof); or (vi) a light chain variable region (VL) comprising an amino acid sequence selected from a sequence having at least 70%, at least 75%, 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%, or at least 99% sequence identity to the sequence set forth in SEQ ID NO: 13 or 33.

[0217] In some embodiments, the anti-B7H3 antibody or antigen-binding fragment thereof comprises a VH set forth in SEQ ID NO:3 and / or a VL set forth in SEQ ID NO:13.

[0218] In some embodiments, the anti-B7H3 antibody or antigen-binding fragment thereof comprises a VH set forth in SEQ ID NO:23 and / or a VL set forth in SEQ ID NO:33.

[0219] In some embodiments, the anti-B7H3 antibody or antigen-binding fragment thereof comprises a VH set forth in SEQ ID NO:3 and / or a VL set forth in SEQ ID NO:33.

[0220] In some embodiments, the anti-B7H3 antibody or antigen-binding fragment thereof comprises a VH set forth in SEQ ID NO:23 and / or a VL set forth in SEQ ID NO:13.

[0221] In some embodiments, the anti-B7H3 antibody or antigen-binding fragment thereof is (a) VH set forth in SEQ ID NO: 3 and VL set forth in SEQ ID NO: 13; (b) VH set forth in SEQ ID NO: 23 and VL set forth in SEQ ID NO: 33; (c) VH set forth in SEQ ID NO: 3 and VL set forth in SEQ ID NO: 13; (d) VH set forth in SEQ ID NO: 23 and VL set forth in SEQ ID NO: 13; (e) a heavy chain variable region (VH) and a light chain variable region (VL) that independently have 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%, or at least 99% sequence identity compared to the VH and VL defined in any one of (a) to (f); or (f) A heavy chain variable region (VH) and a light chain variable region (VL) that, compared to the VH and VL defined in any one of (a) to (d), independently have one or more amino acid substitutions, deletions, or additions, or any combination thereof (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acid substitutions, deletions, or additions, or any combination thereof). Preferably, the substitutions are conservative substitutions.

[0222] In some embodiments, the heavy chain of the anti-B7H3 antibody comprises a human immunoglobulin heavy chain constant region (CH) or a variant thereof, wherein the variant has up to 50 conservative amino acid substitutions (e.g., up to 45, up to 40, up to 35, up to 30, up to 25, up to 20, up to 15, up to 10, or up to 5 conservative amino acid substitutions, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 conservative amino acid substitutions) compared to the wild-type sequence from which it is derived. In some embodiments, the light chain of the anti-B7H3 antibody comprises a human immunoglobulin light chain constant region (CL) or a variant thereof, wherein the variant has up to 50 conservative amino acid substitutions (e.g., up to 45, up to 40, up to 35, up to 30, up to 25, up to 20, up to 15, up to 10, or up to 5 conservative amino acid substitutions, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 conservative amino acid substitutions) compared to the wild-type sequence from which it is derived.

[0223] In some embodiments, the constant region is altered, e.g., mutated, to modify the properties of the anti-B7H3 antibody molecule (e.g., to alter one or more of the following properties: Fc receptor binding, antibody glycosylation, the number of cysteine ​​residues, effector cell function, or complement function). Functional alterations can be made by substituting at least one amino acid residue in the constant region of the antibody with a different residue, e.g., to alter the affinity of the antibody for an effector ligand (e.g., FcR or complement C1q), thereby altering (e.g., decreasing) the effector function. The Fc region of an antibody mediates several important effector functions, such as ADCC, phagocytosis (ADCP), and CDC.

[0224] In some embodiments, an anti-B7H3 antibody or antigen-binding fragment thereof has a heavy chain constant region (CH) selected from, for example, the heavy chain constant regions of IgG1, IgG2, IgG3, IgG4, IgM, IgA1, IgA2, IgD, and IgE, particularly, for example, the heavy chain constant regions of IgG1, IgG2, IgG3, and IgG4, more particularly, the heavy chain constant region of IgG1 (e.g., human IgG1). In some embodiments, the heavy chain constant region of human IgG1 is as set forth in SEQ ID NO: 43. In some embodiments, an antibody or antigen-binding fragment thereof of the present disclosure has a light chain constant region selected from, for example, a kappa or lambda light chain constant region, preferably a kappa light chain constant region (e.g., a human kappa light chain constant region). In some embodiments, the light chain constant region has the sequence set forth in SEQ ID NO: 44.

[0225] In some embodiments, the anti-B7H3 antibody or antigen-binding fragment thereof comprises a CH set forth in SEQ ID NO:43 or a variant thereof, wherein the variant has up to 20 conservative amino acid substitutions (e.g., up to 20, up to 15, up to 10, or up to 5 conservative amino acid substitutions, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 conservative amino acid substitutions) compared to SEQ ID NO:43, or 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%, or at least 99% sequence identity compared to SEQ ID NO:43.

[0226] In some embodiments, the anti-B7H3 antibody or antigen-binding fragment thereof comprises a light chain constant region or variant thereof. In some embodiments, the light chain constant region comprises a kappa light chain constant region. In some embodiments, the light chain constant region comprises the light chain constant region (CL) set forth in SEQ ID NO:44 or a variant thereof, wherein the variant has up to 20 amino acid conservative substitutions (e.g., up to 20, up to 15, up to 10, or up to 5 amino acid conservative substitutions, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acid conservative substitutions) compared to SEQ ID NO:44, or 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%, or at least 99% sequence identity compared to SEQ ID NO:44. In some embodiments, the anti-B7H3 antibody or antigen-binding fragment thereof comprises a heavy chain constant region (CH) set forth in SEQ ID NO:43 and a light chain constant region (CL) set forth in SEQ ID NO:44.

[0227] In some embodiments, the anti-B7H3 antibody or antigen-binding fragment thereof is (a) Below: (i) a sequence comprising the VH sequence set forth in SEQ ID NO: 3 and the CH sequence set forth in SEQ ID NO: 43; (ii) a sequence having one or more amino acid substitutions, deletions or additions, or any combination thereof, compared to the sequence set forth in (i) (e.g., up to 50, up to 45, up to 40, up to 35, up to 30, up to 25, up to 20, up to 15, up to 10, or up to 5 amino acid substitutions, deletions or additions, or any combination thereof, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acid substitutions, deletions or additions, or any combination thereof); or (iii) a heavy chain comprising an amino acid sequence selected from the group consisting of a sequence having 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%, or at least 99% sequence identity compared to the sequence set forth in (i); and (b) Below: (iv) a sequence comprising the VL sequence set forth in SEQ ID NO: 13 and the CL sequence set forth in SEQ ID NO: 44; (v) a sequence having one or more amino acid substitutions, deletions or additions, or any combination thereof, compared to the sequence set forth in (iv) (e.g., up to 50, up to 45, up to 40, up to 35, up to 30, up to 25, up to 20, up to 15, up to 10, or up to 5 amino acid substitutions, deletions or additions, or any combination thereof, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acid substitutions, deletions or additions, or any combination thereof); or (vi) a light chain comprising an amino acid sequence selected from a sequence having 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%, or at least 99% sequence identity compared to the sequence set forth in (iv).

[0228] In some embodiments, the substitution described in (ii) or (v) is a conservative substitution.

[0229] In some embodiments, the anti-B7H3 antibody or antigen-binding fragment thereof is (a) Below: (i) a sequence comprising the VH sequence set forth in SEQ ID NO: 23 and the CH sequence set forth in SEQ ID NO: 43; (ii) a sequence having one or more amino acid substitutions, deletions or additions, or any combination thereof, compared to the sequence set forth in (i) (e.g., up to 50, up to 45, up to 40, up to 35, up to 30, up to 25, up to 20, up to 15, up to 10, or up to 5 amino acid substitutions, deletions or additions, or any combination thereof, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acid substitutions, deletions or additions, or any combination thereof); or (iii) a heavy chain comprising an amino acid sequence selected from the group consisting of a sequence having 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%, or at least 99% sequence identity compared to the sequence set forth in (i); and (b) Below: (iv) a sequence comprising the VL sequence set forth in SEQ ID NO: 33 and the CL sequence set forth in SEQ ID NO: 44; (v) a sequence having one or more amino acid substitutions, deletions or additions, or any combination thereof, compared to the sequence set forth in (iv) (e.g., up to 50, up to 45, up to 40, up to 35, up to 30, up to 25, up to 20, up to 15, up to 10, or up to 5 amino acid substitutions, deletions or additions, or any combination thereof, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acid substitutions, deletions or additions, or any combination thereof); or (vi) a light chain comprising an amino acid sequence selected from a sequence having 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%, or at least 99% sequence identity compared to the sequence set forth in (iv).

[0230] In some embodiments, the substitution described in (ii) or (v) is a conservative substitution.

[0231] In some embodiments, the anti-B7H3 antibody or antigen-binding fragment thereof comprises a heavy chain and a light chain, The heavy chain is (i) the sequence set forth in SEQ ID NO: 45; (ii) a sequence having one or more amino acid substitutions, deletions or additions, or any combination thereof, compared to the sequence set forth in (i) (e.g., up to 50, up to 45, up to 40, up to 35, up to 30, up to 25, up to 20, up to 15, up to 10, or up to 5 amino acid substitutions, deletions or additions, or any combination thereof, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acid substitutions, deletions or additions, or any combination thereof); or (iii) a sequence having 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%, or at least 99% sequence identity compared to the sequence set forth in (i); and The light chain is (iv) the sequence set forth in SEQ ID NO: 46; (v) a sequence having one or more amino acid substitutions, deletions or additions, or any combination thereof, compared to the sequence set forth in (iv) (e.g., up to 50, up to 45, up to 40, up to 35, up to 30, up to 25, up to 20, up to 15, up to 10, or up to 5 amino acid substitutions, deletions or additions, or any combination thereof, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acid substitutions, deletions or additions, or any combination thereof); or (vi) A sequence having 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%, or at least 99% sequence identity compared to the sequence set forth in (iv).

[0232] Preferably, the substitutions described in (ii) or (v) are conservative substitutions.

[0233] In some embodiments, the anti-B7H3 antibody or antigen-binding fragment thereof comprises a heavy chain and a light chain, The heavy chain is (i) the sequence set forth in SEQ ID NO: 47; (ii) a sequence having one or more amino acid substitutions, deletions or additions, or any combination thereof, compared to the sequence set forth in (i) (e.g., up to 50, up to 45, up to 40, up to 35, up to 30, up to 25, up to 20, up to 15, up to 10, or up to 5 amino acid substitutions, deletions or additions, or any combination thereof, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acid substitutions, deletions or additions, or any combination thereof); or (iii) a sequence having 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%, or at least 99% sequence identity compared to the sequence set forth in (i); and The light chain is (iv) the sequence set forth in SEQ ID NO: 48; (v) a sequence having one or more amino acid substitutions, deletions or additions, or any combination thereof, compared to the sequence set forth in (iv) (e.g., up to 50, up to 45, up to 40, up to 35, up to 30, up to 25, up to 20, up to 15, up to 10, or up to 5 amino acid substitutions, deletions or additions, or any combination thereof, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acid substitutions, deletions or additions, or any combination thereof); or (vi) A sequence having 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%, or at least 99% sequence identity compared to the sequence set forth in (iv).

[0234] Preferably, the substitutions described in (ii) or (v) are conservative substitutions.

[0235] In some embodiments, the anti-B7H3 antibody or antigen-binding fragment thereof is a chimeric antibody, a humanized antibody, or a fully human antibody.

[0236] In some embodiments, the anti-B7H3 antibody or antigen-binding fragment thereof is selected from an scFv, a Fab, a Fab', a (Fab')2, an Fv fragment, a disulfide-linked Fv (dsFv), and a diabody.

[0237] In some embodiments, the anti-B7H3 antibody or antigen-binding fragment thereof is an scFv. In certain embodiments, the scFv of the present disclosure comprises: (a) VH set forth in SEQ ID NO: 3 and VL set forth in SEQ ID NO: 13; (b) VH set forth in SEQ ID NO: 23 and VL set forth in SEQ ID NO: 33; (c) VH set forth in SEQ ID NO: 3 and VL set forth in SEQ ID NO: 33; (d) VH set forth in SEQ ID NO: 23 and VL set forth in SEQ ID NO: 13; (e) a heavy chain variable region (VH) and a light chain variable region (VL), wherein the heavy chain variable region (VH) and the light chain variable region (VL) independently have 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%, or at least 99% sequence identity compared to the VH and VL in any one of groups (a) to (d), respectively; or (f) A heavy chain variable region (VH) and a light chain variable region (VL) that independently have one or more amino acid substitutions, deletions, or additions, or any combination thereof (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acid substitutions, deletions, or additions, or any combination thereof), compared to the VH and VL of any one of groups (a) to (d). Preferably, the substitutions are conservative substitutions.

[0238] In some embodiments, the antibodies of the present disclosure are scFvs. In certain embodiments, the scFvs of the present disclosure are (i) a sequence set forth in SEQ ID NO: 1 or 2; (ii) a sequence having one or more amino acid substitutions, deletions or additions, or any combination thereof, compared to the sequence set forth in (i) (e.g., up to 50, up to 45, up to 40, up to 35, up to 30, up to 25, up to 20, up to 15, up to 10, or up to 5 amino acid substitutions, deletions or additions, or any combination thereof, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acid substitutions, deletions or additions, or any combination thereof); or (iii) a sequence having 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%, or at least 99% sequence identity compared to the sequence set forth in (i); Preferably, the substitutions described in (ii) are conservative substitutions.

[0239] In some embodiments, the anti-B7H3 antibody or antigen-binding fragment thereof is an scFv. In certain embodiments, an scFv of the present disclosure comprises the sequence set forth in SEQ ID NO: 1 or 2.

[0240] In some embodiments, the anti-B7H3 antibody or antigen-binding fragment thereof is selected from antibody 1D1, antibody 1D1-01, antibody 2E3, and antibody 2E3-02.

[0241] In some embodiments of the present disclosure, the targeting moiety is trastuzumab or pertuzumab. Trastuzumab is an anti-Her2 monoclonal antibody, the amino acid sequence of which is known to those skilled in the art, and an exemplary sequence can be found, for example, in Chinese Patent No. 103319599, and the terminal Lys residue is easily deleted without affecting its biological activity (see Dick, L. Wet et al., Biotechnol. Bioeng., 100:1132-1143).

[0242] Exemplary heavy and light chain sequences of trastuzumab can refer to, for example, IMGT / mAb-DB ID 97. Exemplary heavy and light chain sequences of pertuzumab can refer to SEQ ID NO: 16 and SEQ ID NO: 15 disclosed in U.S. Pat. No. 7,560,111, and IMGT / mAb-DB ID 80.

[0243] In some embodiments of the present disclosure, the targeting moiety is an anti-Her3 antibody or an antigen-binding fragment thereof. In some embodiments, the anti-Her3 antibody includes all anti-Her3 antibodies in the prior art, such as varsetumab, durigotuzumab, elgemtumab, istiratumab, lanletuzumab, patritumab, seribantumab, xenoctuzumab, 202-2-1 antibody, and the antibody having a heavy chain set forth in SEQ ID NO: 10 and a light chain set forth in SEQ ID NO: 14 disclosed in Chinese Patent No. 103189392B, and the antibody represented by IMGT / mAb-DB ID: 546.

[0244] In some embodiments, the targeting moiety is an anti-Her3 antibody, and the anti-Her3 antibody or antigen-binding fragment thereof is an antibody or antigen-binding fragment thereof comprising the complementarity determining regions (CDRs) shown below, wherein the CDRs are numbered according to the IMGT numbering system: HCDR1 consisting of the sequence of SEQ ID NO: 56, HCDR2 consisting of the sequence of SEQ ID NO: 57, HCDR3 consisting of the sequence of SEQ ID NO: 58, and / or It is defined by LCDR1 consisting of the sequence of SEQ ID NO:41, LCDR2 consisting of the sequence AAS, and LCDR3 consisting of the sequence of SEQ ID NO:21.

[0245] In some embodiments, the anti-Her3 antibody or antigen-binding fragment thereof as defined by the IMGT numbering system is It comprises a VH comprising the above-described heavy chain HCDR1, HCDR2, and HCDR3, and a VL comprising the above-described light chain LCDR1, LCDR2, and LCDR3.

[0246] In some embodiments, the anti-Her3 antibody or antigen-binding fragment thereof is an antibody or antigen-binding fragment thereof that comprises the complementarity-determining regions (CDRs) shown below, wherein the CDRs are numbered according to the Kabat numbering system: HCDR1 consisting of the sequence of SEQ ID NO: 53, HCDR2 consisting of the sequence of SEQ ID NO: 54, HCDR3 consisting of the sequence of SEQ ID NO: 55, and / or It is defined by LCDR1 consisting of the sequence of SEQ ID NO:59, LCDR2 consisting of the sequence of SEQ ID NO:60, and LCDR3 consisting of the sequence of SEQ ID NO:21.

[0247] In some embodiments, the anti-Her3 antibody or antigen-binding fragment thereof defined by the Kabat numbering system comprises a VH comprising the heavy chain HCDR1, HCDR2, and HCDR3 described above, and a VL comprising the light chain LCDR1, LCDR2, and LCDR3 described above.

[0248] In some embodiments, the anti-Her3 antibody or antigen-binding fragment thereof is (a) Below: (i) the sequence set forth in SEQ ID NO: 49; (ii) a sequence having one or more amino acid substitutions, deletions, or additions, or any combination thereof, compared to the sequence set forth in SEQ ID NO: 49 (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acid substitutions, deletions, or additions, or any combination thereof); or (iii) a heavy chain variable region (VH) comprising an amino acid sequence selected from the group consisting of a sequence having at least 70%, at least 75%, 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%, or at least 99% sequence identity to the sequence set forth in SEQ ID NO: 49; and / or (b) Below: (iv) the sequence set forth in SEQ ID NO: 50; (v) a sequence having one or more amino acid substitutions, deletions, or additions, or any combination thereof, compared to the sequence set forth in SEQ ID NO: 50 (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acid substitutions, deletions, or additions, or any combination thereof); or (vi) a light chain variable region (VL) comprising an amino acid sequence selected from a sequence having at least 70%, at least 75%, 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%, or at least 99% sequence identity to the sequence set forth in SEQ ID NO: 50.

[0249] In some embodiments, the anti-Her3 antibody or antigen-binding fragment thereof is (a) VH set forth in SEQ ID NO: 49 and VL set forth in SEQ ID NO: 50; (b) a heavy chain variable region (VH) and a light chain variable region (VL), each of which independently 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%, or at least 99% sequence identity compared to the VH and VL in group (a); or (c) a heavy chain variable region (VH) and a light chain variable region (VL), each of which independently has one or more amino acid substitutions, deletions, or additions, or any combination thereof (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acid substitutions, deletions, or additions, or any combination thereof), compared to the VH and VL described in group (a). Preferably, the substitutions are conservative substitutions.

[0250] In some embodiments, the heavy chain of the anti-Her3 antibody comprises a human immunoglobulin heavy chain constant region (CH) or a variant thereof, wherein the variant has up to 50 conservative amino acid substitutions (e.g., up to 45, up to 40, up to 35, up to 30, up to 25, up to 20, up to 15, up to 10, or up to 5 conservative amino acid substitutions, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 conservative amino acid substitutions) compared to the wild-type sequence from which it is derived. In some embodiments, the light chain of the anti-B7H3 antibody comprises a human immunoglobulin light chain constant region (CL) or a variant thereof, wherein the variant has up to 50 conservative amino acid substitutions (e.g., up to 45, up to 40, up to 35, up to 30, up to 25, up to 20, up to 15, up to 10, or up to 5 conservative amino acid substitutions, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 conservative amino acid substitutions) compared to the wild-type sequence from which it is derived.

[0251] In some embodiments, the constant region is altered, e.g., mutated, to modify the properties of the anti-Her3 antibody molecule (e.g., to change one or more of the following properties: Fc receptor binding, antibody glycosylation, the number of cysteine ​​residues, effector cell function, or complement function). Functional alterations can be made by substituting at least one amino acid residue in the constant region of the antibody with a different residue, e.g., to alter the affinity of the antibody for an effector ligand (e.g., FcR or complement C1q), thereby altering (e.g., decreasing) the effector function. The Fc region of an antibody mediates several important effector functions, such as ADCC, phagocytosis (ADCP), and CDC.

[0252] In some embodiments, an anti-Her3 antibody or antigen-binding fragment thereof has a heavy chain constant region (CH) selected from, for example, the heavy chain constant regions of IgG1, IgG2, IgG3, IgG4, IgM, IgA1, IgA2, IgD, and IgE, particularly, for example, the heavy chain constant regions of IgG1, IgG2, IgG3, and IgG4, more particularly, the heavy chain constant region of IgG1 (e.g., human IgG1). In some embodiments, an antibody or antigen-binding fragment thereof of the present disclosure has a light chain constant region selected from, for example, a kappa or lambda light chain constant region, preferably a kappa light chain constant region (e.g., a human kappa light chain constant region).

[0253] In some embodiments, the anti-Her3 antibody or antigen-binding fragment thereof has a heavy chain constant region (CH) set forth in SEQ ID NO: 43 and a light chain constant region (CL) set forth in SEQ ID NO: 44.

[0254] In some embodiments, the anti-Her3 antibody or antigen-binding fragment thereof comprises a heavy chain and a light chain, The heavy chain is (i) the sequence set forth in SEQ ID NO: 51; (ii) a sequence having one or more amino acid substitutions, deletions or additions, or any combination thereof, compared to the sequence set forth in (i) (e.g., up to 50, up to 45, up to 40, up to 35, up to 30, up to 25, up to 20, up to 15, up to 10, or up to 5 amino acid substitutions, deletions or additions, or any combination thereof, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acid substitutions, deletions or additions, or any combination thereof); or (iii) a sequence having 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%, or at least 99% sequence identity compared to the sequence set forth in (i); and The light chain is (iv) the sequence set forth in SEQ ID NO: 52; (v) a sequence having one or more amino acid substitutions, deletions or additions, or any combination thereof, compared to the sequence set forth in (iv) (e.g., up to 50, up to 45, up to 40, up to 35, up to 30, up to 25, up to 20, up to 15, up to 10, or up to 5 amino acid substitutions, deletions or additions, or any combination thereof, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acid substitutions, deletions or additions, or any combination thereof); or (vi) A sequence having 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%, or at least 99% sequence identity compared to the sequence set forth in (iv). Preferably, the substitutions described in (ii) or (v) are conservative substitutions.

[0255] In some embodiments, the anti-Her3 antibody or antigen-binding fragment thereof is a chimeric antibody, a humanized antibody, or a fully human antibody.

[0256] In some embodiments, the anti-Her3 antibody or antigen-binding fragment thereof is selected from an scFv, a Fab, a Fab', a (Fab')2, an Fv fragment, a disulfide-linked Fv (dsFv), and a diabody.

[0257] In some embodiments, the anti-Her3 antibody or antigen-binding fragment thereof is an scFv. In certain embodiments, the scFv of the present disclosure comprises: (a) VH set forth in SEQ ID NO: 49 and VL set forth in SEQ ID NO: 50; (b) a heavy chain variable region (VH) and a light chain variable region (VL), each independently having 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%, or at least 99% sequence identity compared to the VH and VL of group (a); or (c) a heavy chain variable region (VH) and a light chain variable region (VL), each of which independently has one or more amino acid substitutions, deletions, or additions, or any combination thereof (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acid substitutions, deletions, or additions, or any combination thereof), compared to the VH and VL of group (a). Preferably, the substitutions are conservative substitutions.

[0258] In some embodiments, the anti-Her3 antibody or antigen-binding fragment thereof is selected from antibody 202-2-1.

[0259] In some embodiments of the present disclosure, the targeting moiety is cetuximab, an anti-EGFR monoclonal antibody whose amino acid sequence is known to those of skill in the art and whose exemplary sequence can refer to the antibody represented by IMGT / mAb-DB ID151.

[0260] In some embodiments, the Ligand Drug Conjugate is selected from: [ka] [ka] [ka] wherein Tb1 is an anti-B7H3 antibody or antigen-binding fragment thereof, e.g., enoblituzumab, mirzotamab, omburtamab, antibody 1D1-01, or antibody 2E3-02, preferably antibody 1D1-01 or antibody 2E3-02, and q is selected from any number between 0.1 and 16.0, preferably any number between 2 and 8. In some embodiments, q is 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10. In some preferred embodiments, q is 2, 4, 6, or 8. In some embodiments, the Ligand Drug Conjugate is selected from: [ka] wherein Tb2 is an anti-Trop-2 antibody or antigen-binding fragment thereof, e.g., datopotamab or sacituzumab, preferably sacituzumab, and q is selected from any number between 0.1 and 16.0, preferably any number between 2 and 8. In some embodiments, q is 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10. In some preferred embodiments, q is more preferably 2, 4, 6, or 8.

[0261] In some embodiments, the Ligand Drug Conjugate is selected from: [ka] wherein Tb3 is an anti-Her2 antibody or antigen-binding fragment thereof, e.g., ambenitamab, coprelotamab, dicitamab, gancotamab, margetuximab, pertuzumab, timigituzumab, zanidatamab, trastuzumab, and pertuzumab, preferably trastuzumab and pertuzumab; and q is selected from any number between 0.1 and 16.0, preferably any number between 2 and 8. In some embodiments, q is 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10. In some preferred embodiments, q is 2, 4, 6, or 8.

[0262] In some embodiments, the Ligand Drug Conjugate is selected from: [ka] Tb4 is an anti-Her3 antibody or antigen-binding fragment thereof, e.g., varsetamab, durigotuzumab, elgemtumab, istiratumab, lanletuzumab, patritumab, seribantumab, xenoctuzumab, antibody 202-2-1, an antibody having a heavy chain set forth in SEQ ID NO: 10 and a light chain set forth in SEQ ID NO: 14 disclosed in Chinese Patent No. 103189392B, an antibody represented by IMGT / mAb-DB ID: 546, and antibody 202-2-1, preferably antibody 202-2-1, wherein q is selected from any number between 0.1 and 16.0, preferably any number between 2 and 8. In some embodiments, q is 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10. In some preferred embodiments, q is 2, 4, 6, or 8.

[0263] In some embodiments, the Ligand Drug Conjugate is selected from: [ka] Tb5 is an anti-EGFR antibody or antigen-binding fragment thereof, e.g., demvitamab, depatucizumab, futuximab, imatuzumab, lapritucimab, rosatucizumab, matuzumab, modutucimab, necitumumab, nimotuzumab, panitumumab, pimurtamab, cercultamab, tomzotucimab, zalutumumab, or cetuximab, preferably cetuximab, and q is selected from any number between 0.1 and 16.0, preferably any number between 2 and 8. In some embodiments, q is 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10. In some preferred embodiments, q is 2, 4, 6, or 8.

[0264] In some embodiments, the Ligand Drug Conjugate is selected from: [ka] In the formula, Tb6 is an antibody that does not have tumor cell endocytosis activity or an antibody that has binding activity to a non-endocytosis antigen (e.g., ALCAM / CD166), for example, an IgG isotype antibody that does not have a corresponding cell surface antigen in humans, an anti-CD166 antibody, preferably an anti-chicken lysozyme human IgG1 isotype antibody; and q is selected from any number between 0.1 and 16.0, preferably any number between 2 and 8. In some embodiments, q is 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10. In some preferred embodiments, q is 2, 4, 6, or 8.

[0265] In some embodiments, the Ligand Drug Conjugate is selected from: [ka] In the formula, Tb7 is an antibody that has weak or no tumor cell endocytosis activity but has tumor cell surface antigen binding activity, and q is selected from any number between 0.1 and 16.0, preferably any number between 2 and 8. In some embodiments, q is 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10. In some preferred embodiments, q is 2, 4, 6, or 8.

[0266] In some embodiments, the antibody has binding activity to the B7H3 antigen but does not have endocytic activity, such as INV721 and I7-01 disclosed in WO2021168379A1. More specifically, the anti-B7H3 antibody has a VH set forth in SEQ ID NO:2 and a VL set forth in SEQ ID NO:1 disclosed in WO2021168379A1.

[0267] In some embodiments, the antibody has GD-2 antigen-binding activity but no endocytic activity, such as INV721 and GD2-5 disclosed in WO2021168379A1. More specifically, the anti-GD-2 antibody has a VH set forth in SEQ ID NO:4 and a VL set forth in SEQ ID NO:3 disclosed in WO2021168379A1.

[0268] In some embodiments, the antibody has activity of binding to the HER3 antigen but does not have endocytic activity, for example, the anti-HER3 antibody 21F06 set forth in SEQ ID NO: 22 disclosed in U.S. Pat. No. 10,808,032 B2.

[0269] In some embodiments, the antibody has binding activity to the CD20 antigen but does not have endocytic activity. While "type II" CD20-specific antibodies have been shown to be difficult to internalize by CD20-positive target cells, other so-called "type I" CD20-specific antibodies have been found to be internalized and degraded to some extent, depending on the activation and inhibition of FcγR expression levels on the target cells they interact with. In some embodiments, the antibody that has binding activity to the antigen CD20 but does not have endocytic activity is a "type II" CD20-specific antibody, such as obinutuzumab.

[0270] In some embodiments, the antibody is an antibody having binding activity to a non-endocytic antigen (e.g., ALCAM / CD166). In some embodiments, the antibody is an antibody comprising a VH set forth in SEQ ID NO: 73 and a VL set forth in SEQ ID NO: 74, a VH set forth in SEQ ID NO: 75 and a VL set forth in SEQ ID NO: 76, a VH set forth in SEQ ID NO: 77 and a VL set forth in SEQ ID NO: 78, or a VH set forth in SEQ ID NO: 79 and a VL set forth in SEQ ID NO: 88, as described in EP 3911682A1.

[0271] In some embodiments, the Ligand Drug Conjugate is selected from: [ka] wherein Tb8 is an antibody having tumor cell endocytosis activity and tumor cell surface antigen binding activity, such as antibody 1D1-01, antibody 2E3-02, sacituzumab, pertuzumab, trastuzumab, or cetuximab antibody, and q is selected from any number between 0.1 and 16.0, preferably any number between 2 and 8. In some embodiments, q is 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10. In some preferred embodiments, q is 2, 4, 6, or 8.

[0272] In addition, it should be noted, and can be understood by those skilled in the art, that L is linked to a sulfhydryl group contained in Tb (e.g., an antibody) after the disulfide bond is cleaved (e.g., reduction of the disulfide bond with the reducing agent TCEP can cleave the disulfide bond to generate the sulfhydryl -SH), i.e., -S- between L and Tb is not an additional foreign sulfur atom. For example, in the formula, -S- is not an additional foreign sulfur atom, but is -S- formed by linking the sulfhydryl group contained in Tb after opening the disulfide bond to L.

[0273] In a second aspect of the disclosure, the disclosure provides a compound of formula II: [ka] or a stereoisomer of the compound, a prodrug thereof, a pharmaceutically acceptable salt thereof, a pharmaceutically acceptable solvate thereof, or a drug linker conjugate thereof, During the ceremony, R1, R2 are each independently selected from H, halogen, —OH, optionally substituted C1-6 alkyl, and optionally substituted C1-6 alkoxy; or R1 and R2, together with the carbon atoms to which they are attached, form a 5- to 7-membered carbocyclic or heterocyclic ring containing one or more O, S, N, carbonyl, sulfoxide or sulfone groups, or any combination thereof; R3 is selected from H, halogen, —OH, —NH2, optionally substituted C1-6 alkyl and optionally substituted C1-6 alkoxy; or R3 and X, together with the carbon atoms to which they are attached, form a 5- to 7-membered carbocyclic or heterocyclic ring containing one or more O, S, N, carbonyl, sulfoxide, or sulfone groups, or any combination thereof; R3 and R2, together with the carbon atoms to which they are attached, form a 5- to 7-membered carbocyclic or heterocyclic ring containing one or more O, S, N, carbonyl, sulfoxide or sulfone groups, or any combination thereof; W is absent or present, and when W is present, W is -O-, -S-, -NHR4-, [ka] and position 1 is attached to X; X is a direct bond, an optionally substituted -O-(CH2) n3 -, -NR4-(CH2) n3 -, -S-(CH2) n3 -, carbonyl-(CH2) n3 -, -SO2-(CH2) n3 -, -(CH2) n1 -, [ka] selected from C3-6 cycloalkyl, C6-10 aryl, 5-10 membered heteroaryl and 4-10 membered heterocyclyl, wherein position 1 is attached to the parent ring and position 2 is attached to W, and the substituents are selected from one or more C1-4 alkyl groups, C3-6 cycloalkyl groups, or multiple C1-4 alkyl groups together with the carbon atoms to which they are both attached form a C3-6 cycloalkyl group; Each M independently represents a direct bond and -CR 5a R 5b - selected from R4, R5, R 5a , R 5b , R6, R7 are each independently selected from H, optionally substituted C1-4 alkyl, optionally substituted C1-4 alkoxy, and optionally substituted C3-6 cycloalkyl; n, n', n1, n2, and n3 are each independently selected from any integer of 0 to 6; R1 and R2 are both H and X is -(CH2) n1 - and n1 is 1, 2, 3, and 4, W is not -OH or -NHR4, and the compound of formula II is [ka] Does not include.

[0274] In some embodiments, R1 and R2 are each independently selected from H, halogen, or C1-4 alkyl.

[0275] In some embodiments, R1 and R2 together with the carbon atoms to which they are attached form a 5-6 membered heterocycle containing 1, 2, or 3 O, S, or N, or any combination thereof.

[0276] In some embodiments, R1 is selected from H and halogen, and R2 is selected from H and C1-4 alkyl.

[0277] In some embodiments, R1 and R2, together with the carbon atom to which they are attached, [ka] where the dotted line indicates where the heterocycle is fused to the benzene ring.

[0278] In some embodiments, R1 is H or F and R2 is H or methyl.

[0279] In some embodiments, R1 and R2, together with the carbon atom to which they are attached, [ka] Form.

[0280] In some embodiments, R1 is F and R2 is methyl, or R1 and R2 together with the carbon atom to which they are attached are: [ka] Form.

[0281] In some embodiments, R1 is F and R2 is methyl.

[0282] In some embodiments, R1 and R2, together with the carbon atom to which they are attached, [ka] Form.

[0283] In some embodiments, R3 is selected from H and C1-4 alkyl.

[0284] In some embodiments, R3 is H.

[0285] In some embodiments, W is absent or present, and when W is present, W is -OH, -SH, -NHR4, [ka] and position 1 is attached to X.

[0286] In some embodiments, W is absent or present, and when W is present, W is -OH, -SH, -NHR4, [ka] and position 1 is attached to X.

[0287] In some embodiments, W is —OH, —NHR 4 , and [ka] and position 1 is attached to X.

[0288] In some embodiments, W is selected from -OH and -NHR4, and position 1 is attached to X.

[0289] In some embodiments, X is optionally substituted -(CH) n1 -, [ka] It is selected from C6-10 aryl, 5- to 10-membered heteroaryl, and 4- to 10-membered heterocyclyl, where position 1 is bonded to the parent ring and position 2 is bonded to W, and the substituents are selected from one or two C1-4 alkyl groups, or two C1-4 alkyl groups together with the carbon atom to which they are both attached form a C3-6 cycloalkyl group.

[0290] In some embodiments, X is optionally substituted [ka] wherein position 1 is attached to the parent ring and position 2 is attached to W, and the substituents are selected from one or two C1-4 alkyl groups (e.g., methyl), or two C1-4 alkyl groups (e.g., methyl) taken together with the carbon atom to which they are both attached form a C3-6 cycloalkyl group (e.g., cyclopropyl).

[0291] In some embodiments, X is [ka] wherein position 1 is attached to the parent ring and position 2 is attached to W.

[0292] In some embodiments, X is [ka] wherein position 1 is attached to the parent ring and position 2 is attached to W.

[0293] In some embodiments, when W is absent, X is [ka] where position 1 is attached to the parent ring and when W is present, X is selected from: [ka] wherein position 1 is attached to the parent ring and position 2 is attached to W.

[0294] In some embodiments, W is —OH, —NHR 4 , and [ka] wherein position 1 is bonded to X and position 2 is bonded to L4 or L3, and X is selected from: [ka] wherein position 1 is attached to the parent ring and position 2 is attached to W. In some embodiments, R4 and R5 are each independently selected from H, C1-4 alkyl, and C3-6 cycloalkyl.

[0295] In some embodiments, each R4 is independently selected from H, C1-4 alkyl, and C3-6 cycloalkyl; and R5 is H.

[0296] In some embodiments, each R4 is independently selected from H, methyl, ethyl, isopropyl, n-propyl, t-butyl, and cyclopropyl; and R5 is H.

[0297] In some embodiments, R 5a and R5b are each independently selected from H and C1-4 alkyl; In some embodiments, R 5a and R 5b are each independently selected from H and methyl; In some embodiments, each R 7 is independently selected from H and C1-4 alkyl; In some embodiments, R 7 is H.

[0298] In some embodiments, n is 1, 2, or 3.

[0299] In some embodiments, n is 1.

[0300] In some embodiments, n1 is 1, 2, 3, or 4.

[0301] In some embodiments, n2 is 1; In some embodiments, n3 is 0.

[0302] In some embodiments, the compound represented by Formula II is the following compound: [ka] [ka] [ka] The value is selected from either: In a third aspect of the present disclosure, the present disclosure provides a drug-linker conjugate of formula III [ka] or a stereoisomer of the drug linker conjugate, a prodrug thereof, a pharmaceutically acceptable salt thereof, or a pharmaceutically acceptable solvate thereof, During the ceremony, R1 and R2 are each independently selected from H, halogen, —OH, optionally substituted C1-6 alkyl, and optionally substituted C1-6 alkoxy; or R1 and R2, together with the carbon atoms to which they are attached, form a 5- to 7-membered carbocyclic or heterocyclic ring containing one or more O, S, N, carbonyl, sulfoxide or sulfone groups, or any combination thereof; R3 is selected from H, halogen, —OH, —NH2, optionally substituted C1-6 alkyl and optionally substituted C1-6 alkoxy; or R3 and X together with the carbon atoms to which they are attached form a 5- to 7-membered carbocyclic or heterocyclic ring containing one or more O, S, N, carbonyl, sulfoxide, or sulfone groups, or any combination thereof; or R3 and R2, together with the carbon atoms to which they are attached, form a 5- to 7-membered carbocyclic or heterocyclic ring containing one or more O, S, N, carbonyl, sulfoxide or sulfone groups, or any combination thereof; W is absent or present, and when W is present, W is -O-, -S-, -NR4-, [ka] wherein position 1 is bonded to X and position 2 is bonded to L4 or L3; X is a direct bond, an optionally substituted -O-(CH2) n3 -, -N(R4)-(CH2) n3 -, -S-(CH2) n3 -, carbonyl-(CH2) n3 , -SO2-(CH2) n3 -, [ka] -(CH2) n1-, C3-6 cycloalkyl, C6-10 aryl, 5-10 membered heteroaryl and 4-10 membered heterocyclyl, wherein position 1 is bonded to the parent ring and position 2 is bonded to W or L4, and the substituents are selected from one or more C1-4 alkyl groups, C3-6 cycloalkyl groups, or multiple C1-4 alkyl groups together with the carbon atoms to which they are both attached form a C3-6 cycloalkyl group; Each M independently represents a direct bond and -CR 5a R 5b - selected from R4, R5, R 5a , R 5b , R6, R7 are each independently selected from H, optionally substituted C1-4 alkyl, optionally substituted C1-4 alkoxy, and optionally substituted C3-6 cycloalkyl; n, n', n1, n2, and n3 are each independently selected from any integer of 0 to 6; L1 is [ka] [ka] is selected from wherein each Z is independently selected from a direct bond, a carbon-carbon triple bond, a carbon-carbon double bond, a C6-10 aryl, a 5-10 membered heteroaryl, an amide, a sulfonamide, an imino, and a CF2; Rx and Ry are each independently selected from H and a C1-4 alkyl; each m is independently selected from 0, 1, 2, 3, 4, 5, and 6; y1, y2, y3, and y4 are each independently selected from any integer from 0 to 20; position 1 is bonded to Lg; position 2 is bonded to L2 or L3; L2 is absent or present, and if present, L2 is [ka] [ka] is selected from In the formula, y1, y2, y3, and y4 are each independently selected from any integer of 0 to 20, the 1-position is bonded to L1, and the 2-position is bonded to L3; L3 is selected from an amino acid residue or a short peptide consisting of 2 to 10 amino acid residues; L4 is absent or present, and if present, L4 is [ka] wherein position 1 is bonded to L3 and position 2 is bonded to W or X; Lg is a leaving group, and Lg is a halogen, a sulfonic acid group, a tertiary amine salt group (MeN + , Et3N + ), a diazonium salt group, selected from -OMs, MeSO2-, and CF3SO3-.

[0303] In some embodiments, R1 and R2 are each independently selected from H, halogen, and C1-4 alkyl.

[0304] In some embodiments, R1 and R2 together with the carbon atoms to which they are attached form a 5-6 membered heterocycle containing 1, 2, or 3 O, S, or N, or any combination thereof.

[0305] In some embodiments, R1 is selected from H and halogen, and R2 is selected from H and C1-4 alkyl.

[0306] In some embodiments, R1 and R2, together with the carbon atom to which they are attached, [ka] where the dotted line indicates where the heterocycle is fused to the benzene ring.

[0307] In some embodiments, R1 is H or F and R2 is H or methyl.

[0308] In some embodiments, R1 is F and R2 is methyl, or R1 and R2 together with the carbon atom to which they are attached are: [ka] Form.

[0309] In some embodiments, R1 is F and R2 is methyl.

[0310] In some embodiments, R1 and R2, together with the carbon atom to which they are attached, [ka] Form.

[0311] In some embodiments, R3 is selected from H, C1-4 alkyl.

[0312] In some embodiments, R3 and X together with the carbon atoms to which they are attached form a 5- to 6-membered carbocyclic ring.

[0313] In some embodiments, R3 is H, or R3 and X together with the carbon atom to which they are attached are [ka] where the dotted lines indicate where the carbocyclic ring is fused to the benzene and pyridine rings.

[0314] [In some embodiments, R3 is H.]

[0315] In some embodiments, R3 and X together with the carbon atom to which they are attached form: [ka] where the dotted lines indicate where the carbocyclic ring is fused to the benzene and pyridine rings.

[0316] In some embodiments, W is absent or present, and when W is present, W is -O-, -S-, -NR4-, [ka] wherein position 1 is bonded to X and position 2 is bonded to L4 or L3.

[0317] In some embodiments, W is absent or present, and when W is present, W is -O-, -S-, -NR4-, [ka] wherein position 1 is bonded to X and position 2 is bonded to L4 or L3.

[0318] In some embodiments, W is —O—, —NR4—, [ka] wherein position 1 is bonded to X and position 2 is bonded to L4 or L3.

[0319] In some embodiments, W is selected from -O-, -NR4-, position 1 is bonded to X, and position 2 is bonded to L4 or L3.

[0320] In some embodiments, X is optionally substituted -(CH) n1 -, [ka] It is selected from C6-10 aryl, 5- to 10-membered heteroaryl, and 4- to 10-membered heterocyclyl, where position 1 is bonded to the parent ring and position 2 is bonded to W or L4, and the substituents are selected from one or two C1-4 alkyl groups, or two C1-4 alkyl groups together with the carbon atom to which they are both attached form a C3-6 cycloalkyl group.

[0321] In some embodiments, X is optionally substituted [ka] where position 1 is attached to the parent ring and position 2 is attached to W or L4. The substituents are selected from one or two C1-4 alkyl groups (e.g., methyl), or two C1-4 alkyl groups together with the carbon atom to which they are both attached form a C3-6 cycloalkyl group (e.g., cyclopropyl).

[0322] In some embodiments, X is [ka] wherein position 1 is attached to the parent ring and position 2 is attached to W or L4.

[0323] In some embodiments, X is [ka] wherein position 1 is attached to the parent ring and position 2 is attached to W.

[0324] In some embodiments, when W is absent, X is [ka] wherein position 1 is attached to the parent ring and position 2 is attached to L4; and when W is present, X is selected from: [ka] wherein position 1 is attached to the parent ring and position 2 is attached to W.

[0325] In some embodiments, W is —O—, —NR4—, [ka] wherein position 1 is bonded to X and position 2 is bonded to L4 or L3, and X is selected from: [ka] wherein position 1 is attached to the parent ring and position 2 is attached to W.

[0326] In some embodiments, R4, R5 are each independently selected from H, C1-4 alkyl, and C3-6 cycloalkyl.

[0327] In some embodiments, each R4 is independently selected from H, C1-4 alkyl, and C3-6 cycloalkyl; and R5 is H.

[0328] In some embodiments, each R4 is independently selected from H, methyl, ethyl, n-propyl, isopropyl, t-butyl, and cyclopropyl; and R5 is H.

[0329] In some embodiments, R 5a , R 5b are each independently selected from H and C1-4 alkyl.

[0330] In some embodiments, R 5a , R 5b are each independently selected from H and methyl.

[0331] In some embodiments, each R 7 is independently selected from H and C1-4 alkyl.

[0332] In some embodiments, R 7 is H.

[0333] In some embodiments, n is selected from 1, 2, or 3.

[0334] In some embodiments, n is 1.

[0335] In some embodiments, n1 is selected from 1, 2, 3, or 4.

[0336] In some embodiments, n2 is 1.

[0337] In some embodiments, n3 is 0.

[0338] In some embodiments, L1 is [ka] wherein each Z is independently selected from a direct bond, a carbon-carbon triple bond, a carbon-carbon double bond, a C6-10 aryl, a 5-10 membered heteroaryl, and an amide (each Z is preferably selected from a direct bond, a carbon-carbon triple bond, and a carbon-carbon double bond); Rx and Ry are each independently selected from H and a C1-4 alkyl; each m is independently selected from 0, 1, 2, 3, 4, 5, and 6; y1 is independently selected from any integer between 1 and 6 (e.g., 4, 5, 6); each y2 is independently selected from any integer between 0 and 15 (e.g., 6-15); each y3 is independently selected from 1, 2, or 3; and each y4 is independently selected from 0 or 1, where position 1 is bonded to Lg and position 2 is bonded to L2 or L3; for example, each Z is independently selected from a direct bond, a carbon-carbon triple bond, or a carbon-carbon double bond; Rx and Ry are each independently selected from H and C1-4 alkyl; each m is independently selected from 0, 1, 2, 3, 4, 5, and 6; y1 is selected from any integer between 1 and 6 (e.g., 4, 5, 6); each y2 is independently selected from any integer between 0 and 15 (e.g., 6 to 15); each y3 is independently selected from 1, 2, or 3; and each y4 is independently selected from 0 or 1; position 1 is bonded to Lg via the S atom, and position 2 is bonded to L2 or L3.

[0339] In some embodiments, L1 is [ka] wherein m is selected from 2, 3, and 4; y1 is selected from any integer from 1 to 6 (e.g., 4, 5, 6); each y2 is independently selected from any integer from 0 to 10 (e.g., 6 to 10); each y3 is independently selected from 1 or 2; and the 1-position is bonded to Lg and the 2-position is bonded to L2 or L3.

[0340] In some embodiments, L1 is [ka] wherein position 1 is attached to Lg and position 2 is attached to L2 or L3.

[0341] In some embodiments, L1 is [ka] wherein position 1 is attached to Lg and position 2 is attached to L2 or L3.

[0342] In some embodiments, L1 is [ka] wherein position 1 is attached to Lg and position 2 is attached to L2 or L3.

[0343] In some embodiments, L1 is [ka] wherein position 1 is attached to Lg and position 2 is attached to L2 or L3.

[0344] In some embodiments, L2 is absent or present, and when L2 is present, L2 is [ka] wherein y1 is selected from any integer from 1 to 6 (e.g., 4, 5, 6), each y2 is independently selected from any integer from 0 to 10 (e.g., 6 to 10), each y3 is independently selected from 1 or 2, and each y4 is independently selected from 0 or 1, and position 1 is bonded to L1 and position 2 is bonded to L3.

[0345] In some embodiments, L2 is absent or present, and when L2 is present, L2 is [ka] wherein position 1 is attached to L1 and position 2 is attached to L3.

[0346] In some embodiments, L2 is absent or present, and when L2 is present, L2 is [ka] wherein position 1 is attached to L1 and position 2 is attached to L3.

[0347] In some embodiments, L2 is absent or present, and when L2 is present, L2 is [ka] wherein position 1 is attached to L1 and position 2 is attached to L3.

[0348] In some embodiments, L2 is absent.

[0349] In some embodiments, L2 is [ka] is selected from.

[0350] In some embodiments, L3 is selected from an amino acid residue or a short peptide consisting of 2 to 10 amino acid residues, wherein the amino acid residues are selected from natural amino acid residues, unnatural amino acid residues, or AA 1 or a stereoisomer thereof.

[0351] In some embodiments, L3 is selected from the group consisting of the amino acid residues Val, D-Val, Cit, Phe, Lys, Lys(Ac), Leu, Gly, Ala, Asn, Asp, Arg, and AA. 1 , or Val, Cit, Phe, Lys, D-Val, Leu, Gly, Ala, Asn, Asp, and AA 1 a short peptide consisting of 2 to 10 amino acid residues selected from In some embodiments, L3 is Val, Cit, Phe, Lys, D-Val, Leu, Gly, Ala, Asn, AA 1 , Val-Cit, Cit-Val, Cit-Ala, Val-Ala, Lys-Val, Val-Lys(Ac), Phe-Lys, Phe-Lys(Ac), Ala-Ala, Val-AA 1 , Ala-AA 1 , Gly-AA 1 , A.A. 1 -Gly, Ala-Ala-Ala, Ala-Ala-Asn, Ala-Ala-Asp, Val-AA 1 -Gly, Ala-AA 1 -Gly, Gly-AA 1 and Lys-Ala-Asn, Gly-Gly-Phe, D-Val-Leu-Lys, Gly-Gly-Arg, Ala-Ala-Asn, Gly-Gly-Phe, Val-Lys-Gly, Val-Lys-Gly-Gly, Val-Lys, and Lys-Ala-Asn.

[0352] In some embodiments, L3 is AA 1 , A.A. 1 -Gly, Val-Cit, Val-AA 1 -Gly, AA 1 -Ala-Asn, and Gly-Gly-Phe-Gly.

[0353] In some embodiments, L3 is AA 1 , Val-AA 1 -Gly.

[0354] In some embodiments, L3 is Val-AA 1 -Gly.

[0355] In some embodiments, L3 is [ka] [ka] wherein X is selected from - is a halide ion, a carboxylate ion, a sulfate ion, a hydrogen sulfate ion, and an OH - wherein position 1 is attached to L1 or L2 and position 2 is attached to L4 or W.

[0356] In some embodiments, L3 is [ka] wherein X is selected from - are halide ions, carboxylate ions, sulfate ions, hydrogen sulfate ions, and OH - wherein position 1 is attached to L1 or L2 and position 2 is attached to L4 or W.

[0357] In some embodiments, L3 is [ka] wherein X is selected from - is a halide ion, a carboxylate ion, a sulfate ion, a hydrogen sulfate ion, and an OH - wherein position 1 is attached to L1 or L2 and position 2 is attached to L4 or W.

[0358] In some embodiments, L3 is [ka] wherein position 1 is attached to L1 or L2 and position 2 is attached to L4 or W.

[0359] In some embodiments, L3 is [ka] wherein position 1 is attached to L1 or L2 and position 2 is attached to L4 or W.

[0360] In some embodiments, AA 1 The structure of the amino acid residue represented by is as follows: [ka] During the ceremony, R a and R b are each independently H, [ka] Selected from R a and R b Is it possible that both of them are H? Or R a and R b together with the carbon atoms to which they are both attached form a 4- to 10-membered heterocyclic ring, and the 4- to 10-membered heterocyclic ring is 0 is optionally replaced by r, r 1 are each independently selected from any integer of 0 to 20, R m1 , R n1 are each independently H, C1-6 alkyl, C3-6 cycloalkyl, or -COOR x1 is selected from R x1 is selected from C1-6 alkyl; Or R m1 and R n1 together with the nitrogen atom to which they are both attached form a 4- to 10-membered heterocyclic ring, and the 4- to 10-membered heterocyclic ring is 0’ is optionally replaced by R z is selected from C1-6 alkyl; R 0 , R 0’ are each independently C1-6 alkyl, C3-6 cycloalkyl, -NR m2 R n2 and 4-10 membered heterocyclyl optionally substituted with C alkyl; R m2 , Rn2 are each independently selected from H and C alkyl; In some embodiments, R a and R b One of them is H and the other is [ka] is selected from.

[0361] In some embodiments, R a and R b One of them is H and the other is [ka] is selected from.

[0362] In some embodiments, R a and R b together with the carbon atom to which they are both attached, form R 0 Forms a 5- to 6-membered heterocyclic ring substituted with

[0363] In some embodiments, R a and R b together with the carbon atom to which they are both attached, form R 0 The piperidine or piperazine ring is formed by the following procedure.

[0364] In some embodiments, R a and R b together with the carbon atom to which they are both attached, form R 0 The compound forms a piperidine ring substituted by

[0365] In some embodiments, R a and R b together with the carbon atom to which they are both attached, [ka] and the carbon atom numbered 1 is Ra and R b is the carbon atom to which both are bonded.

[0366] In some embodiments, R a and R b together with the carbon atom to which they are both attached, [ka] and the carbon atom numbered 1 is R a and R b is the carbon atom to which both are bonded.

[0367] In some embodiments, r and r 1 are each independently selected from 0, 1, 2, 3, 4 and 5.

[0368] In some embodiments, r and r 1 are each independently selected from 0 and 4.

[0369] In some embodiments, r and r 1 One of them is 0 and the other is 4.

[0370] In some embodiments, R m1 and R n1 are each independently selected from H, methyl, ethyl, n-propyl, n-butyl, —COOCH, —COOCHCH, —COOCHCH, —COOCH(CH), —COOC(CH), and —COOCHCHCHCH.

[0371] In some embodiments, R m1 and R n1 are each independently selected from H, C1-6 alkyl, C3-6 cycloalkyl, and t-butoxycarbonyl.

[0372] In some embodiments, R m1 and R n1are each independently selected from H and C1-6 alkyl.

[0373] In some embodiments, R m1 and R n1 are each independently selected from H, methyl, ethyl, and n-propyl.

[0374] In some embodiments, r and r 1 For r = 4, r 1 is 0 and R m1 and R n1 are each independently selected from H and C alkyl (e.g., H, methyl), r is 0, and r 1 If is 4, R m1 and R n1 are each independently selected from C1-6 alkyl (eg, methyl, ethyl, n-propyl), preferably C2-6 alkyl (eg, ethyl, n-propyl).

[0375] In some embodiments, R m1 and R n1 together with the nitrogen atom to which they are both attached, form R 0’ Forms a 5- to 6-membered heterocyclic ring substituted with

[0376] In some embodiments, R m1 and R n1 together with the nitrogen atom to which they are both attached, form R 0’ The piperidine or piperazine ring is formed by the following procedure.

[0377] In some embodiments, R m1 and R n1 together with the nitrogen atom to which they are both attached, [ka] and the nitrogen atom numbered 1 is R m1 and R n1 is the nitrogen atom to which both are bonded.

[0378] In some embodiments, R z is methyl. In some embodiments, R 0 and R 0’ are each independently C1-6 alkyl, -NR m2 R n2 and 5-6 membered heterocyclyl optionally substituted with C1-6 alkyl.

[0379] In some embodiments, R 0 is selected from C1-6 alkyl and 5- to 6-membered heterocyclyl substituted with C1-6 alkyl, said 5- to 6-membered heterocyclyl being selected from piperidinyl and piperazinyl.

[0380] In some embodiments, R 0 is selected from methyl, ethyl, and 5- to 6-membered heterocyclyl substituted with methyl, wherein the 5- to 6-membered heterocyclyl is piperidinyl.

[0381] In some embodiments, R 0 is selected from methyl and 5- to 6-membered heterocyclyl substituted with methyl, and the 5- to 6-membered heterocyclyl is piperidinyl.

[0382] In some embodiments, R 0 is methyl, ethyl and [ka] is selected from.

[0383] In some embodiments, R 0 is methyl and [ka] is selected from.

[0384] In some embodiments, R 0’is C1-6 alkyl and -NR m2 R n2 is selected from.

[0385] In some embodiments, R 0’ is methyl and -NR m2 R n2 is selected from.

[0386] In some embodiments, R m2 and R n2 is methyl.

[0387] In some embodiments, AA 1 The amino acid residue represented by [ka] is selected from.

[0388] In some embodiments, AA 1 The amino acid residue represented by [ka] is selected from.

[0389] In some embodiments, AA 1 The amino acid residue represented by [ka] is selected from.

[0390] In some embodiments, AA 1 The amino acid residue represented by [ka] is selected from.

[0391] In some embodiments, L4 is absent or present, and when L4 is present, L4 is [ka] wherein position 1 is bonded to L3 and position 2 is bonded to W or X.

[0392] In some embodiments, L4 is absent or present, and when L4 is present, L4 is [ka] wherein position 1 is bonded to L3 and position 2 is bonded to W or X.

[0393] In some embodiments, L4 is absent.

[0394] In some embodiments, L4 is [ka] wherein position 1 is bonded to L3 and position 2 is bonded to W or X.

[0395] In some embodiments, L4 is [ka] wherein position 1 is bonded to L3 and position 2 is bonded to W or X.

[0396] In some embodiments, Lg is selected from F, Cl, and MeSO2-.

[0397] In some embodiments, Lg is selected from F and MeSO2-.

[0398] In some embodiments, [ka] The structure of is selected from: [Table 4-1] [Table 4-2] [Table 4-3] Position 1 is attached to Lg and position 2 is attached to W.

[0399] In some embodiments, the structure [ka] is selected from: [ka] [ka] [ka] wherein position 1 is bonded to L4, and if L4 is absent, position 1 is bonded to L3.

[0400] In some embodiments, the drug linker conjugate has the structure of Formula III-(1): [ka] and wherein L1, L2, L3, L4, X, R1, R2, R3, R4, and Lg are as defined above and in any embodiment specifically described herein.

[0401] In some embodiments, the drug linker conjugate has the structure of Formula III-(1A) or III-(1B): [ka] and wherein L2, L3, L4, X, R1, R2, R3, R4, and Lg are as defined above and in any embodiment specifically described herein.

[0402] In some embodiments, the drug linker conjugate has the structure of Formula III-(2): [ka] and wherein L1, L2, L3, L4, X, R1, R2, R3 and Lg are as defined above and in any embodiment specifically described herein.

[0403] In some embodiments, the drug linker conjugate has the structure of Formula III-(2A) or III-(2B): [ka] and wherein L2, L3, L4, X, R1, R2, R3 and Lg are as defined above and in any embodiment specifically described herein.

[0404] In some embodiments, the drug linker conjugate has the structure of Formula III-(3): [ka] and wherein L1, L2, L3, L4, X, R1, R2, R3, R4, R5, n, and Lg are as defined above and in any embodiment specifically described herein.

[0405] In some embodiments, the drug linker conjugate has the structure of Formula III-(3A) or III-(3B): [ka] and wherein Lg, L2, L3, L4, X, R1, R2, R3, R4 and Lg are as defined above and in any embodiment specifically described herein.

[0406] In some embodiments, the drug linker conjugate has the structure of Formula III-A: [ka] and In the formula, Lg, X, R1, R2, R3, R a , R b and q are as defined above and in any embodiment specifically described herein.

[0407] In some embodiments, the drug linker conjugate has the structure of Formula III-B: [ka] and In the formula, Lg, X, R1, R2, R3, R a , R b and q are as defined above and in any embodiment specifically described herein.

[0408] In some embodiments, the drug linker conjugate represented by formula III is selected from the following structures: [Table 5-1] [Table 5-2] [Table 5-3] [Table 5-4] [Table 5-5] [Table 5-6] [Table 5-7] [Table 5-8] [Table 5-9] [Table 5-10] [Table 5-11] [Table 5-12] [Table 5-13] [Table 5-14] [Table 5-15] [Table 5-16] [Table 5-17] [Table 5-18] [Table 5-19] [Table 5-20] [Table 5-21] [Table 5-22]

[0409] In some embodiments, compounds are provided in which the L1-L2-L3 unit in the drug linker-conjugate of Formula III is partially cleaved, thus leaving a drug moiety to which an amino acid residue is attached.

[0410] In some embodiments, the partially released free drug is a compound of formula III-(A): [ka] or a stereoisomer thereof or a mixture of stereoisomers thereof, or a pharmaceutically acceptable salt thereof, wherein L4, X, W, R1, R2 and R3 are as defined above and in any embodiment specifically described herein.

[0411] In some embodiments, the structure: [ka] III-(A) is selected from the following: [ka] [ka] [ka] [ka] [ka] The present disclosure also provides a linker in a ligand drug conjugate comprising the following fragment: [ka] Position 2 binds to the biologically active molecular fragment (position 1 binds to the ligand terminus, e.g., if it is bound to a linking unit (e.g., L2) and then to a ligand or targeting moiety (Tb) via an extender unit (e.g., L1), or if there is no linking unit, position 1 binds directly to the extender unit and then to the ligand); L3, L4 are defined as in any one of the embodiments of the present disclosure.

[0412] In some embodiments, the linker in the Ligand Drug Conjugate has the following structure: [ka] Position 1 binds to a ligand or targeting moiety that binds to a target, and position 2 binds to a biologically active molecular fragment; L1, L2, L3, L4 are defined as in any one of the embodiments of the present disclosure; Preferably, the ligand or targeting moiety that binds to a target and the biologically active molecular fragment are defined for Tb and D, respectively, in any one of the embodiments of the present disclosure.

[0413] In some embodiments, position 1 of the linker is attached to the antibody or antigen-binding fragment thereof and position 2 is attached to the biologically active molecule fragment.

[0414] In some embodiments, position 1 of the linker binds to a cysteine ​​or lysine in the antibody or antigen-binding fragment thereof, and position 2 binds to the biologically active molecule fragment; Preferably, the antibody or antigen-binding fragment thereof is as described in any of the embodiments of the present disclosure.

[0415] In a fourth aspect of the present disclosure, the present disclosure also provides a linker represented by formula III-1: [ka] Lg, L1, L2, L3 and L4 are defined above and in any embodiment specifically described herein, and Lg1 is a leaving group in the reaction with a drug molecule.

[0416] In some embodiments, Lg1 is preferably —OH, [ka] Or a halogen.

[0417] In a fifth aspect of the present disclosure, the present invention also provides a linker represented by formula III-2: [ka] Lg, L1, L2 and L3 are defined above and in any embodiment specifically described herein, and Lg2 is a leaving group in the reaction with L4 or a fragment comprising a drug molecule.

[0418] In some embodiments, Lg2 is preferably -OH, [ka] Or a halogen.

[0419] In a sixth aspect of the present disclosure, the present disclosure provides methods for preparing compounds (drug molecules, bioactive molecules) of formula II, as well as drug-linker-conjugates of formula III.

[0420] In one aspect, the present disclosure provides a method for preparing a compound of formula II (a pharmaceutical molecule, a biologically active molecule), specifically as follows: Compound (VI) can be obtained by the alkylation reaction of compound (IV) with compound (V) catalyzed by an iron compound or other related Minisci reaction. [ka]

[0421] Alternatively, compound (VII) can be synthesized by N-oxidation of compound (IV), and then compound (VII) can be treated with phosphorus oxyhalide to obtain compound (VIII), which is a halide of compound (IV). Compound (VI) can be obtained from compound (VIII) by various reactions such as the Heck reaction, Suzuki reaction, Buchwald reaction, Nishishi reaction, and Stille reaction. [ka]

[0422] A different or more complex molecule may be R in compound (VI). 11 can be synthesized by a variety of chemical transformations, such as oxidation, reduction, substitution and other methods commonly found in textbooks.

[0423] Alternatively, compound (VI) and compound (IV) can be produced from compound (IX) and compound (X) by a ring-closure reaction as shown below. [ka]

[0424] In another aspect, the present disclosure provides methods for preparing fragments represented by formula III-1 and formula III-2. In particular, Compound III-c can be obtained by the reaction of functional group Fg1 in compound III-a with functional group Fg2 in compound III-b, and then compound III-d can be obtained by the reaction of functional group Fg a Compounds III-2 and III-1 represented by the general formula can be obtained by a similar conversion. a , Lg b , Lg1 and Lg2 are, for example, —OH, [ka] or a reactive leaving group such as a halogen. [ka]

[0425] In yet another aspect, the present disclosure provides a method for preparing a drug-linker-conjugate of formula III, in particular: The target product can be obtained by coupling fragment B with fragment C, as shown in the following reaction scheme: The conjugate product is subjected to several common chemical modifications, such as oxidation, deprotection, etc., to obtain the drug-linker conjugate represented by general formula III.

[0426] [ka]

[0427] In a seventh aspect of the present disclosure, the present disclosure provides a method for preparing the aforementioned Ligand Drug Conjugate, comprising: Tb can be converted to a compound of formula III in a suitable solvent and under suitable conditions. [ka] is coupled to a drug linker conjugate represented by During the ceremony, Tb is as defined above and in any embodiment specifically described herein; R1, R2, R3, X, W, L1, L2, L3, L4 and Lg are defined above and in any embodiment specifically described herein.

[0428] In some embodiments, the method comprises reacting Tb with a compound of formula III in a suitable solvent and under suitable conditions. [ka] to form a C-S bond.

[0429] In some embodiments, the ratio of the Tb to the drug linker conjugate in molar amounts is 1:(1-20), e.g., 1:2, 1:3, 1:4, 1:5, 1:6, 1:7, 1:8, 1:9, 1:10, 1:12, 1:14, 1:16, 1:18, 1:(10-20), 1:(12-20), 1:(14-20), 1:(16-20), or 1:(18-20).

[0430] In some embodiments, the coupling reaction is carried out in water and / or an organic solvent.

[0431] In some embodiments, the organic solvent is selected from N,N-dimethylformamide, dimethyl sulfoxide, N-methylpyrrolidone, a nitrile (eg, acetonitrile), an alcohol (eg, methanol, ethanol), or any combination thereof.

[0432] In some embodiments, the process further comprises purifying the coupling product.

[0433] In some embodiments, the coupling product is purified by chromatography.

[0434] In some embodiments, the chromatography comprises one or more of ion exchange chromatography, hydrophobic chromatography, reverse phase chromatography, or affinity chromatography.

[0435] In an eighth aspect of the present disclosure, the present disclosure provides an antibody or antigen-binding fragment thereof that binds to B7H3, wherein the antibody or antigen-binding fragment comprises a complementarity-determining region (CDR) as follows: HCDR1 or a variant thereof, HCDR2 or a variant thereof, and HCDR3 or a variant thereof contained in the heavy chain variable region (VH) set forth in SEQ ID NO: 3 or 23, and / or LCDR1 or a variant thereof, LCDR2 or a variant thereof, and LCDR3 or a variant thereof contained in the light chain variable region (VL) set forth in SEQ ID NO: 13 or 33.

[0436] In certain embodiments, the antibody or antigen-binding fragment thereof comprises HCDR1 or a variant thereof, HCDR2 or a variant thereof, and HCDR3 or a variant thereof contained in the VH set forth in SEQ ID NO: 3, and / or LCDR1 or a variant thereof, LCDR2 or a variant thereof, and LCDR3 or a variant thereof contained in the VL set forth in SEQ ID NO: 13.

[0437] In certain embodiments, the antibody or antigen-binding fragment thereof comprises HCDR1 or a variant thereof, HCDR2 or a variant thereof, and HCDR3 or a variant thereof contained in the VH set forth in SEQ ID NO: 3, and / or LCDR1 or a variant thereof, LCDR2 or a variant thereof, and LCDR3 or a variant thereof contained in the VL set forth in SEQ ID NO: 33.

[0438] In certain embodiments, the antibody or antigen-binding fragment thereof comprises HCDR1 or a variant thereof, HCDR2 or a variant thereof, and HCDR3 or a variant thereof contained in the VH set forth in SEQ ID NO: 23, and / or LCDR1 or a variant thereof, LCDR2 or a variant thereof, and LCDR3 or a variant thereof contained in the VL set forth in SEQ ID NO: 33.

[0439] In certain embodiments, the antibody or antigen-binding fragment thereof comprises HCDR1 or a variant thereof, HCDR2 or a variant thereof, and HCDR3 or a variant thereof contained in the VH set forth in SEQ ID NO: 23, and / or LCDR1 or a variant thereof, LCDR2 or a variant thereof, and LCDR3 or a variant thereof contained in the VL set forth in SEQ ID NO: 13.

[0440] In certain preferred embodiments, the sequence variant is a CDR that has one or more amino acid substitutions, deletions or additions (e.g., 1, 2 or 3 amino acid substitutions, deletions or additions) compared to the CDR from which it is derived.

[0441] In certain preferred embodiments, the substitutions are conservative substitutions.

[0442] Preferably, the CDRs are defined according to the AbM, Chothia, Kabat or IMGT numbering systems.

[0443] In certain embodiments, the VH and / or VL of the antibody or antigen-binding fragment thereof comprises a framework region (FR) derived from a human immunoglobulin.

[0444] In certain embodiments, the antibody or antigen-binding fragment thereof binds to human B7H3 and / or monkey B7H3. In certain embodiments, the antibody or antigen-binding fragment thereof binds to human 2Ig B7H3. In certain embodiments, the antibody or antigen-binding fragment thereof binds to human 4Ig B7H3. In certain embodiments, the antibody or antigen-binding fragment thereof binds to monkey 4Ig B7H3. In certain embodiments, the antibody or antigen-binding fragment thereof binds to human 2Ig B7H3 and human 4Ig B7H3, preferably human 4Ig B7H3. In certain embodiments, the antibody or antigen-binding fragment thereof binds to human 4Ig B7H3 but not human 2Ig B7H3.

[0445] In one aspect, the disclosure provides an antibody or antigen-binding fragment thereof capable of binding to B7H3, wherein the antibody or antigen-binding fragment thereof comprises a heavy chain variable region (VH) and / or a light chain variable region (VL).

[0446] In certain embodiments, an antibody or antigen-binding fragment thereof according to the present disclosure comprises the following heavy chain variable region (VH) and / or light chain variable region (VL), wherein the CDRs are numbered according to the IMGT numbering system: (a) a heavy chain variable region (VH) comprising the following three CDRs: HCDR1 consisting of the sequence of SEQ ID NO: 10 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 10; HCDR2 consisting of the sequence of SEQ ID NO: 11 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 11; and HCDR3 consisting of the sequence of SEQ ID NO: 12 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 12; and / or a light chain variable region (VL) comprising the following three CDRs: LCDR1 consisting of the sequence of SEQ ID NO: 20 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 20; LCDR2 consisting of the sequence of GTF or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to GTF; and LCDR3 consisting of the sequence of SEQ ID NO: 22 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 22; (b) a heavy chain variable region (VH) comprising the following three CDRs: HCDR1 consisting of the sequence of SEQ ID NO: 30 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 30; HCDR2 consisting of the sequence of SEQ ID NO: 31 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 31; and HCDR3 consisting of the sequence of SEQ ID NO: 32 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 32; and / or a light chain variable region (VL) comprising the following three CDRs: LCDR1 consisting of the sequence of SEQ ID NO: 40 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 40; LCDR2 consisting of the sequence of GAS or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to GAS; and LCDR3 consisting of the sequence of SEQ ID NO: 42 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 42; (c) a heavy chain variable region (VH) comprising the following three CDRs: HCDR1 consisting of the sequence of SEQ ID NO: 10 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 10; HCDR2 consisting of the sequence of SEQ ID NO: 11 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 11; and HCDR3 consisting of the sequence of SEQ ID NO: 12 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 12; and / or a light chain variable region (VL) comprising the following three CDRs: LCDR1 consisting of the sequence of SEQ ID NO: 40 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 40; LCDR2 consisting of the sequence of GAS or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to GAS; and LCDR3 consisting of the sequence of SEQ ID NO: 42 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 42; or (d) a heavy chain variable region (VH) comprising the following three CDRs: HCDR1 consisting of the sequence of SEQ ID NO: 30 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 30; HCDR2 consisting of the sequence of SEQ ID NO: 31 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 31; and HCDR3 consisting of the sequence of SEQ ID NO: 32 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 32; and / or a light chain variable region (VL) comprising the following three CDRs: LCDR1 consisting of the sequence of SEQ ID NO: 20 or a sequence having one or more amino acid substitutions, deletions, or additions (e.g., one, two, or three amino acid substitutions, deletions, or additions) compared to SEQ ID NO: 20; LCDR2 consisting of the sequence of GTF or a sequence having one or more amino acid substitutions, deletions, or additions (e.g., one, two, or three amino acid substitutions, deletions, or additions) compared to GTF; and LCDR3 consisting of the sequence of SEQ ID NO: 22 or a sequence having one or more amino acid substitutions, deletions, or additions (e.g., one, two, or three amino acid substitutions, deletions, or additions) compared to SEQ ID NO: 22. is defined according to

[0447] In certain embodiments, an antibody or antigen-binding fragment thereof according to the present disclosure comprises the following heavy chain variable region (VH) and / or light chain variable region (VL), wherein the CDRs are numbered according to the IMGT numbering system: (a) a heavy chain variable region (VH) comprising the following three CDRs: HCDR1 consisting of the sequence of SEQ ID NO: 10, HCDR2 consisting of the sequence of SEQ ID NO: 11, and HCDR3 consisting of the sequence of SEQ ID NO: 12; and / or a light chain variable region (VL) comprising the following three CDRs: LCDR1 consisting of the sequence of SEQ ID NO: 20, LCDR2 consisting of the sequence GTF, and LCDR3 consisting of the sequence of SEQ ID NO: 22; (b) a heavy chain variable region (VH) comprising the following three CDRs: HCDR1 consisting of the sequence of SEQ ID NO: 10, HCDR2 consisting of the sequence of SEQ ID NO: 11, and HCDR3 consisting of the sequence of SEQ ID NO: 12; and / or a light chain variable region (VL) comprising the following three CDRs: LCDR1 consisting of the sequence of SEQ ID NO: 40, LCDR2 consisting of the sequence GAS, and LCDR3 consisting of the sequence of SEQ ID NO: 42; (c) a heavy chain variable region (VH) comprising the following three CDRs: HCDR1 consisting of the sequence of SEQ ID NO: 30, HCDR2 consisting of the sequence of SEQ ID NO: 31, and HCDR3 consisting of the sequence of SEQ ID NO: 32; and / or a light chain variable region (VL) comprising the following three CDRs: LCDR1 consisting of the sequence of SEQ ID NO: 40, LCDR2 consisting of the sequence GAS, and LCDR3 consisting of the sequence of SEQ ID NO: 42; or (d) a heavy chain variable region (VH) comprising the following three CDRs: HCDR1 consisting of the sequence of SEQ ID NO: 30, HCDR2 consisting of the sequence of SEQ ID NO: 31, and HCDR3 consisting of the sequence of SEQ ID NO: 32; and / or a light chain variable region (VL) comprising the following three CDRs: LCDR1 consisting of the sequence of SEQ ID NO: 20, LCDR2 consisting of the sequence GTF, and LCDR3 consisting of the sequence of SEQ ID NO: 22; is defined according to

[0448] In certain embodiments, an antibody or antigen-binding fragment thereof according to the present disclosure comprises the following heavy chain variable region (VH) and / or light chain variable region (VL), wherein the CDRs are numbered according to the Chothia numbering system: (a) a heavy chain variable region (VH) comprising the following three CDRs: HCDR1 consisting of the sequence of SEQ ID NO: 4 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 4; HCDR2 consisting of the sequence of SEQ ID NO: 5 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 5; and HCDR3 consisting of the sequence of SEQ ID NO: 6 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 6; and / or a light chain variable region (VL) comprising the following three CDRs: LCDR1 consisting of the sequence of SEQ ID NO: 14 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 14; LCDR2 consisting of the sequence of SEQ ID NO: 15 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 15; and LCDR3 consisting of the sequence of SEQ ID NO: 16 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 16; (b) a heavy chain variable region (VH) comprising the following three CDRs: HCDR1 consisting of the sequence of SEQ ID NO: 4 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 4; HCDR2 consisting of the sequence of SEQ ID NO: 5 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 5; and HCDR3 consisting of the sequence of SEQ ID NO: 6 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 6; and / or a light chain variable region (VL) comprising the following three CDRs: LCDR1 consisting of the sequence of SEQ ID NO: 34 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 34; LCDR2 consisting of the sequence of SEQ ID NO: 35 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 35; and LCDR3 consisting of the sequence of SEQ ID NO: 36 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 36; (c) a heavy chain variable region (VH) comprising the following three CDRs: HCDR1 consisting of the sequence of SEQ ID NO: 24 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 24; HCDR2 consisting of the sequence of SEQ ID NO: 25 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 25; and HCDR3 consisting of the sequence of SEQ ID NO: 26 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 26; and / or a light chain variable region (VL) comprising the following three CDRs: LCDR1 consisting of the sequence of SEQ ID NO: 34 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 34; LCDR2 consisting of the sequence of SEQ ID NO: 35 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 35; and LCDR3 consisting of the sequence of SEQ ID NO: 36 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 36; or (d) a heavy chain variable region (VH) comprising the following three CDRs: HCDR1 consisting of the sequence of SEQ ID NO: 24 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 24; HCDR2 consisting of the sequence of SEQ ID NO: 25 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 25; and HCDR3 consisting of the sequence of SEQ ID NO: 26 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 26; and / or a light chain variable region (VL) comprising the following three CDRs: LCDR1 consisting of the sequence of SEQ ID NO: 14 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two, or three amino acid substitutions, deletions, or additions) compared to SEQ ID NO: 14; LCDR2 consisting of the sequence of SEQ ID NO: 15 or a sequence having one or more amino acid substitutions, deletions, or additions (e.g., one, two, or three amino acid substitutions, deletions, or additions) compared to SEQ ID NO: 15; and LCDR3 consisting of the sequence of SEQ ID NO: 16 or a sequence having one or more amino acid substitutions, deletions, or additions (e.g., one, two, or three amino acid substitutions, deletions, or additions) compared to SEQ ID NO: 16. is defined according to In certain embodiments, an antibody or antigen-binding fragment thereof according to the present disclosure comprises the following heavy chain variable region (VH) and / or light chain variable region (VL), wherein the CDRs are numbered according to the Chothia numbering system: (a) a heavy chain variable region (VH) comprising the following three CDRs: HCDR1 consisting of the sequence of SEQ ID NO: 4, HCDR2 consisting of the sequence of SEQ ID NO: 5, and HCDR3 consisting of the sequence of SEQ ID NO: 6; and / or a light chain variable region (VL) comprising the following three CDRs: LCDR1 consisting of the sequence of SEQ ID NO: 14, LCDR2 consisting of the sequence of SEQ ID NO: 15, and LCDR3 consisting of the sequence of SEQ ID NO: 16; (b) a heavy chain variable region (VH) comprising the following three CDRs: HCDR1 consisting of the sequence of SEQ ID NO: 24, HCDR2 consisting of the sequence of SEQ ID NO: 25, and HCDR3 consisting of the sequence of SEQ ID NO: 26; and / or a light chain variable region (VL) comprising the following three CDRs: LCDR1 consisting of the sequence of SEQ ID NO: 34, LCDR2 consisting of the sequence of SEQ ID NO: 35, and LCDR3 consisting of the sequence of SEQ ID NO: 36; (c) a heavy chain variable region (VH) comprising the following three CDRs: HCDR1 consisting of the sequence of SEQ ID NO: 4, HCDR2 consisting of the sequence of SEQ ID NO: 5, and HCDR3 consisting of the sequence of SEQ ID NO: 6; and / or a light chain variable region (VL) comprising the following three CDRs: LCDR1 consisting of the sequence of SEQ ID NO: 34, LCDR2 consisting of the sequence of SEQ ID NO: 35, and LCDR3 consisting of the sequence of SEQ ID NO: 36; or (d) a heavy chain variable region (VH) comprising the following three CDRs: HCDR1 consisting of the sequence of SEQ ID NO: 24, HCDR2 consisting of the sequence of SEQ ID NO: 25, and HCDR3 consisting of the sequence of SEQ ID NO: 26; and / or a light chain variable region (VL) comprising the following three CDRs: LCDR1 consisting of the sequence of SEQ ID NO: 14, LCDR2 consisting of the sequence of SEQ ID NO: 15, and LCDR3 consisting of the sequence of SEQ ID NO: 16; is defined according to In certain embodiments, an antibody or antigen-binding fragment thereof according to the present disclosure comprises the following heavy chain variable region (VH) and / or light chain variable region (VL), wherein the CDRs are numbered according to the Kabat numbering system: (a) a heavy chain variable region (VH) comprising the following three CDRs: HCDR1 consisting of the sequence of SEQ ID NO: 7 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 7; HCDR2 consisting of the sequence of SEQ ID NO: 8 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 8; and HCDR3 consisting of the sequence of SEQ ID NO: 9 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 9; and / or a light chain variable region (VL) comprising the following three CDRs: LCDR1 consisting of the sequence of SEQ ID NO: 17 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 17; LCDR2 consisting of the sequence of SEQ ID NO: 18 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 18; and LCDR3 consisting of the sequence of SEQ ID NO: 19 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 19; (b) a heavy chain variable region (VH) comprising the following three CDRs: HCDR1 consisting of the sequence of SEQ ID NO: 27 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 27; HCDR2 consisting of the sequence of SEQ ID NO: 28 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 28; and HCDR3 consisting of the sequence of SEQ ID NO: 29 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 29; and / or a light chain variable region (VL) comprising the following three CDRs: LCDR1 consisting of the sequence of SEQ ID NO: 37 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 37; LCDR2 consisting of the sequence of SEQ ID NO: 38 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 38; and LCDR3 consisting of the sequence of SEQ ID NO: 39 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 39; (c) a heavy chain variable region (VH) comprising the following three CDRs: HCDR1 consisting of the sequence of SEQ ID NO: 7 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 7; HCDR2 consisting of the sequence of SEQ ID NO: 8 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 8; and HCDR3 consisting of the sequence of SEQ ID NO: 9 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 9; and / or a light chain variable region (VL) comprising the following three CDRs: LCDR1 consisting of the sequence of SEQ ID NO: 37 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 37; LCDR2 consisting of the sequence of SEQ ID NO: 38 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 38; and LCDR3 consisting of the sequence of SEQ ID NO: 39 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 39; or (d) a heavy chain variable region (VH) comprising the following three CDRs: HCDR1 consisting of the sequence of SEQ ID NO: 27 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 27; HCDR2 consisting of the sequence of SEQ ID NO: 28 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 28; and HCDR3 consisting of the sequence of SEQ ID NO: 29 or a sequence having one or more amino acid substitutions, deletions or additions (e.g., one, two or three amino acid substitutions, deletions or additions) compared to SEQ ID NO: 29; and / or a light chain variable region (VL) comprising the following three CDRs: LCDR1 consisting of the sequence of SEQ ID NO: 17 or a sequence having one or more amino acid substitutions, deletions, or additions (e.g., one, two, or three amino acid substitutions, deletions, or additions) compared to SEQ ID NO: 17; LCDR2 consisting of the sequence of SEQ ID NO: 18 or a sequence having one or more amino acid substitutions, deletions, or additions (e.g., one, two, or three amino acid substitutions, deletions, or additions) compared to SEQ ID NO: 18; and LCDR3 consisting of the sequence of SEQ ID NO: 19 or a sequence having one or more amino acid substitutions, deletions, or additions (e.g., one, two, or three amino acid substitutions, deletions, or additions) compared to SEQ ID NO: 19. is defined according to

[0449] In certain embodiments, an antibody or antigen-binding fragment thereof according to the present disclosure comprises the following heavy chain variable region (VH) and / or light chain variable region (VL), wherein the CDRs are numbered according to the Kabat numbering system: (a) a heavy chain variable region (VH) comprising the following three CDRs: HCDR1 consisting of the sequence of SEQ ID NO: 7, HCDR2 consisting of the sequence of SEQ ID NO: 8, and HCDR3 consisting of the sequence of SEQ ID NO: 9; and / or a light chain variable region (VL) comprising the following three CDRs: LCDR1 consisting of the sequence of SEQ ID NO: 17, LCDR2 consisting of the sequence of SEQ ID NO: 18, and LCDR3 consisting of the sequence of SEQ ID NO: 19; (b) a heavy chain variable region (VH) comprising the following three CDRs: HCDR1 consisting of the sequence of SEQ ID NO: 27, HCDR2 consisting of the sequence of SEQ ID NO: 28, and HCDR3 consisting of the sequence of SEQ ID NO: 29; and / or a light chain variable region (VL) comprising the following three CDRs: LCDR1 consisting of the sequence of SEQ ID NO: 37, LCDR2 consisting of the sequence of SEQ ID NO: 38, and LCDR3 consisting of the sequence of SEQ ID NO: 39; (c) a heavy chain variable region (VH) comprising the following three CDRs: HCDR1 consisting of the sequence of SEQ ID NO: 7, HCDR2 consisting of the sequence of SEQ ID NO: 8, and HCDR3 consisting of the sequence of SEQ ID NO: 9; and / or a light chain variable region (VL) comprising the following three CDRs: LCDR1 consisting of the sequence of SEQ ID NO: 37, LCDR2 consisting of the sequence of SEQ ID NO: 38, and LCDR3 consisting of the sequence of SEQ ID NO: 39; or (d) a heavy chain variable region (VH) comprising the following three CDRs: HCDR1 consisting of the sequence of SEQ ID NO: 27, HCDR2 consisting of the sequence of SEQ ID NO: 28, and HCDR3 consisting of the sequence of SEQ ID NO: 29; and / or a light chain variable region (VL) comprising the following three CDRs: LCDR1 consisting of the sequence of SEQ ID NO: 17, LCDR2 consisting of the sequence of SEQ ID NO: 18, and LCDR3 consisting of the sequence of SEQ ID NO: 19; is defined according to

[0450] In certain embodiments, an antibody or antigen-binding fragment thereof of the present disclosure comprises a heavy chain variable region (VH) and / or a light chain variable region (VL), wherein at least one CDR of the heavy chain variable region (VH) and / or the light chain variable region (VL) contains a mutation compared to the CDR defined by IMGT, Chothia, or Kabat, supra, wherein the mutation is one or more amino acid substitutions, deletions, or additions, or any combination thereof (1, 2, or 3 amino acid substitutions, deletions, or additions, or any combination thereof).

[0451] Preferably, the substitutions referred to in this disclosure are conservative substitutions.

[0452] In certain embodiments, the VH of an antibody or antigen-binding fragment thereof of the present disclosure comprises a framework region (FR) derived from a heavy chain variable region (VH) of a human immunoglobulin, and / or the VL of an antibody or antigen-binding fragment thereof comprises a framework region (FR) derived from a light chain variable region (VL) of a human immunoglobulin. Thus, in certain embodiments, an antibody or antigen-binding fragment thereof of the present disclosure is humanized. In certain embodiments, an antibody or antigen-binding fragment thereof of the present disclosure is fully humanized.

[0453] In certain embodiments, the VH of an antibody or antigen-binding fragment thereof of the present disclosure comprises a framework region (FR) derived from a heavy chain variable region (VH) of a human immunoglobulin, and / or the VL of an antibody or antigen-binding fragment thereof comprises a framework region (FR) derived from a light chain variable region (VL) of a human immunoglobulin. Thus, in certain embodiments, an antibody or antigen-binding fragment thereof of the present disclosure is humanized. In certain embodiments, an antibody or antigen-binding fragment thereof of the present disclosure is fully humanized.

[0454] In certain embodiments, the antibody or antigen-binding fragment thereof of the present disclosure: (a) a heavy chain framework region of a human immunoglobulin or variant thereof, wherein the variant has up to 20 conservative amino acid substitutions (e.g., up to 20, up to 15, up to 10, or up to 5 conservative amino acid substitutions, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 conservative amino acid substitutions) compared to the amino acid sequence encoded by the germline antibody gene from which it is derived; and / or (b) a light chain framework region of a human immunoglobulin or variant thereof, wherein the variant has up to 20 conservative amino acid substitutions (e.g., up to 20, up to 15, up to 10, or up to 5 conservative amino acid substitutions, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 conservative amino acid substitutions) compared to the amino acid sequence encoded by the germline antibody gene from which it is derived. In certain embodiments, the degree of humanization of an antibody or antigen-binding fragment thereof of the present disclosure is at least 75%, 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%, or at least 99%.

[0455] In certain embodiments, the antibody or antigen-binding fragment thereof of the present disclosure: (a) Below: (i) a sequence set forth in SEQ ID NO: 3 or 23; (ii) a sequence having one or more substitutions, deletions, or additions, or any combination thereof, compared to the sequence set forth in SEQ ID NO: 3 or 23 (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acid substitutions, deletions, or additions, or any combination thereof); or (iii) a heavy chain variable region (VH) comprising an amino acid sequence selected from the group consisting of a sequence having at least 70%, at least 75%, 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%, or at least 99% sequence identity compared to the sequence set forth in SEQ ID NO: 3 or 23; and / or (b) Below: (iv) a sequence set forth in SEQ ID NO: 13 or 33; (v) a sequence having one or more substitutions, deletions, or additions, or any combination thereof, compared to the sequence set forth in SEQ ID NO: 13 or 33 (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acid substitutions, deletions, or additions, or any combination thereof); or (vi) a light chain variable region (VL) comprising an amino acid sequence selected from a sequence having at least 70%, at least 75%, 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%, or at least 99% sequence identity to the sequence set forth in SEQ ID NO: 13 or 33.

[0456] In certain embodiments, an antibody or antigen-binding fragment thereof of the present disclosure comprises a VH set forth in SEQ ID NO:3 and / or a VL set forth in SEQ ID NO:13.

[0457] In certain embodiments, an antibody or antigen-binding fragment thereof of the present disclosure comprises a VH set forth in SEQ ID NO:23 and / or a VL set forth in SEQ ID NO:33.

[0458] In certain embodiments, an antibody or antigen-binding fragment thereof of the present disclosure comprises a VH set forth in SEQ ID NO:3 and / or a VL set forth in SEQ ID NO:33.

[0459] In certain embodiments, an antibody or antigen-binding fragment thereof of the present disclosure comprises a VH set forth in SEQ ID NO:23 and / or a VL set forth in SEQ ID NO:13.

[0460] In certain embodiments, the antibody or antigen-binding fragment thereof of the present disclosure: (a) VH set forth in SEQ ID NO: 3 and VL set forth in SEQ ID NO: 13; (b) VH set forth in SEQ ID NO: 23 and VL set forth in SEQ ID NO: 33; (c) VH set forth in SEQ ID NO: 3 and VL set forth in SEQ ID NO: 33; (d) VH set forth in SEQ ID NO: 23 and VL set forth in SEQ ID NO: 13;

[0461] (e) a heavy chain variable region (VH) and / or a light chain variable region (VL), wherein the heavy chain variable region (VH) and the light chain variable region (VL) each independently have 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%, or at least 99% sequence identity compared to the VH and VL in any one of (a) to (f); or (f) A heavy chain variable region (VH) and / or a light chain variable region (VL) that independently have one or more amino acid substitutions, deletions, or additions, or any combination thereof (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acid substitutions, deletions, or additions, or any combination thereof), compared to the VH and VL in any one of (a) to (d). Preferably, the substitutions are conservative substitutions.

[0462] In certain embodiments, an antibody or antigen-binding fragment thereof of the present disclosure comprises a heavy chain constant region (CH) of a human immunoglobulin or a variant thereof, wherein the variant has up to 50 amino acid conservative substitutions (e.g., up to 45, up to 40, up to 35, up to 30, up to 25, up to 20, up to 15, up to 10, or up to 5 amino acid conservative substitutions, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acid conservative substitutions) compared to the wild-type sequence from which it is derived. In certain embodiments, an antibody or antigen-binding fragment thereof of the present disclosure comprises a light chain constant region (CL) of a human immunoglobulin or a variant thereof, wherein the variant has up to 50 amino acid conservative substitutions (e.g., up to 45, up to 40, up to 35, up to 30, up to 25, up to 20, up to 15, up to 10, or up to 5 amino acid conservative substitutions, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acid conservative substitutions) compared to the wild-type sequence from which it is derived.

[0463] In some embodiments, the constant region is altered, e.g., mutated, to modify the properties of the anti-B7H3 antibody molecule (e.g., to alter one or more of the following properties: Fc receptor binding, antibody glycosylation, the number of cysteine ​​residues, effector cell function, or complement function). Functional alterations can be made by substituting at least one amino acid residue in the constant region of the antibody with a different residue, e.g., to alter the affinity of the antibody for an effector ligand (e.g., FcR or complement C1q), thereby altering (e.g., decreasing) the effector function. The Fc region of an antibody mediates several important effector functions, such as ADCC, phagocytosis (ADCP), and CDC.

[0464] In some embodiments, an antibody or antigen-binding fragment thereof of the present disclosure has a heavy chain constant region (CH) selected from, for example, the heavy chain constant regions of IgG1, IgG2, IgG3, IgG4, IgM, IgA1, IgA2, IgD, and IgE, particularly, for example, the heavy chain constant regions of IgG1, IgG2, IgG3, and IgG4, more particularly, the heavy chain constant region of IgG1 (e.g., human IgG1). In some embodiments, the human IgG1 heavy chain constant region has the sequence set forth in SEQ ID NO: 43. In some embodiments, an antibody or antigen-binding fragment thereof of the present disclosure has a light chain constant region selected from, for example, a kappa or lambda light chain constant region, preferably a kappa light chain constant region (e.g., a human kappa light chain constant region). In some embodiments, the light chain constant region has the sequence set forth in SEQ ID NO: 44.

[0465] In some embodiments, the antibody or antigen-binding fragment thereof has a CH set forth in SEQ ID NO: 43 or a variant thereof, wherein the variant has up to 20 conservative amino acid substitutions (e.g., up to 20, up to 15, up to 10, or up to 5 conservative amino acid substitutions, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 conservative amino acid substitutions) compared to SEQ ID NO: 43, or 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%, or at least 99% sequence identity compared to SEQ ID NO: 43.

[0466] In some embodiments, the antibody or antigen-binding fragment thereof comprises a light chain constant region or variant thereof. In some embodiments, the light chain constant region comprises a kappa light chain constant region. In some embodiments, the light chain constant region comprises a light chain constant region (CL) set forth in SEQ ID NO: 44 or a variant thereof, wherein the variant has up to 20 amino acid conservative substitutions (e.g., up to 20, up to 15, up to 10, or up to 5 amino acid conservative substitutions, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acid conservative substitutions) compared to SEQ ID NO: 44, or 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%, or at least 99% sequence identity compared to SEQ ID NO: 16. In some embodiments, the antibody or antigen-binding fragment thereof comprises a heavy chain constant region (CH) set forth in SEQ ID NO: 43 and a light chain constant region (CL) set forth in SEQ ID NO: 44.

[0467] In some embodiments, the antibody or antigen-binding fragment thereof of the present disclosure: (a) Below: (i) a sequence comprising the VH sequence set forth in SEQ ID NO: 3 and the CH sequence set forth in SEQ ID NO: 43; (ii) a sequence having one or more amino acid substitutions, deletions or additions, or any combination thereof, compared to the sequence set forth in (i) (e.g., up to 50, up to 45, up to 40, up to 35, up to 30, up to 25, up to 20, up to 15, up to 10, or up to 5 amino acid substitutions, deletions or additions, or any combination thereof, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acid substitutions, deletions or additions, or any combination thereof); or (iii) a heavy chain comprising an amino acid sequence selected from the group consisting of a sequence having 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%, or at least 99% sequence identity compared to the sequence set forth in (i); and (b) Below: (iv) a sequence comprising the VL sequence set forth in SEQ ID NO: 13 and the CL sequence set forth in SEQ ID NO: 44; (v) a sequence having one or more amino acid substitutions, deletions or additions, or any combination thereof, compared to the sequence set forth in (iv) (e.g., up to 50, up to 45, up to 40, up to 35, up to 30, up to 25, up to 20, up to 15, up to 10, or up to 5 amino acid substitutions, deletions or additions, or any combination thereof, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acid substitutions, deletions or additions, or any combination thereof); or (vi) a light chain comprising an amino acid sequence selected from a sequence having 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%, or at least 99% sequence identity compared to the sequence set forth in (iv).

[0468] In some embodiments, the substitution described in (ii) or (v) is a conservative substitution.

[0469] In some embodiments, the antibody or antigen-binding fragment thereof of the present disclosure: (a) Below: (i) a sequence comprising the VH sequence set forth in SEQ ID NO: 23 and the CH sequence set forth in SEQ ID NO: 43; (ii) a sequence having one or more amino acid substitutions, deletions or additions, or any combination thereof, compared to the sequence set forth in (i) (e.g., up to 50, up to 45, up to 40, up to 35, up to 30, up to 25, up to 20, up to 15, up to 10, or up to 5 amino acid substitutions, deletions or additions, or any combination thereof, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acid substitutions, deletions or additions, or any combination thereof); or (iii) a heavy chain comprising an amino acid sequence selected from the group consisting of a sequence having 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%, or at least 99% sequence identity compared to the sequence set forth in (i); and (b) Below: (iv) a sequence comprising the VL sequence set forth in SEQ ID NO: 33 and the CL sequence set forth in SEQ ID NO: 44; (v) a sequence having one or more amino acid substitutions, deletions or additions, or any combination thereof, compared to the sequence set forth in (iv) (e.g., up to 50, up to 45, up to 40, up to 35, up to 30, up to 25, up to 20, up to 15, up to 10, or up to 5 amino acid substitutions, deletions or additions, or any combination thereof, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acid substitutions, deletions or additions, or any combination thereof); or (vi) a light chain comprising an amino acid sequence selected from a sequence having 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%, or at least 99% sequence identity compared to the sequence set forth in (iv).

[0470] In some embodiments, the substitution described in (ii) or (v) is a conservative substitution.

[0471] In some embodiments, an antibody or antigen-binding fragment thereof of the present disclosure comprises a heavy chain and a light chain: The heavy chain is (i) the sequence set forth in SEQ ID NO: 45; (ii) a sequence having one or more amino acid substitutions, deletions or additions, or any combination thereof, compared to the sequence set forth in (i) (e.g., up to 50, up to 45, up to 40, up to 35, up to 30, up to 25, up to 20, up to 15, up to 10, or up to 5 amino acid substitutions, deletions or additions, or any combination thereof, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acid substitutions, deletions or additions, or any combination thereof); or (iii) a sequence having 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%, or at least 99% sequence identity compared to the sequence set forth in (i); and The light chain is (iv) the sequence set forth in SEQ ID NO: 46; (v) a sequence having one or more amino acid substitutions, deletions or additions, or any combination thereof, compared to the sequence set forth in (iv) (e.g., up to 50, up to 45, up to 40, up to 35, up to 30, up to 25, up to 20, up to 15, up to 10, or up to 5 amino acid substitutions, deletions or additions, or any combination thereof, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acid substitutions, deletions or additions, or any combination thereof); or (vi) a sequence having 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%, or at least 99% sequence identity compared to the sequence set forth in (iv); Preferably, the substitutions described in (ii) or (v) are conservative substitutions.

[0472] In some embodiments, an antibody or antigen-binding fragment thereof of the present disclosure comprises a heavy chain and a light chain: The heavy chain is (i) the sequence set forth in SEQ ID NO: 47; (ii) a sequence having one or more amino acid substitutions, deletions or additions, or any combination thereof, compared to the sequence set forth in (i) (e.g., up to 50, up to 45, up to 40, up to 35, up to 30, up to 25, up to 20, up to 15, up to 10, or up to 5 amino acid substitutions, deletions or additions, or any combination thereof, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acid substitutions, deletions or additions, or any combination thereof); or (iii) a sequence having 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%, or at least 99% sequence identity compared to the sequence set forth in (i); and The light chain is (iv) the sequence set forth in SEQ ID NO: 48; (v) a sequence having one or more amino acid substitutions, deletions or additions, or any combination thereof, compared to the sequence set forth in (iv) (e.g., up to 50, up to 45, up to 40, up to 35, up to 30, up to 25, up to 20, up to 15, up to 10, or up to 5 amino acid substitutions, deletions or additions, or any combination thereof, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acid substitutions, deletions or additions, or any combination thereof); or (vi) a sequence having 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%, or at least 99% sequence identity compared to the sequence set forth in (iv); Preferably, the substitutions described in (ii) or (v) are conservative substitutions.

[0473] In some embodiments, the antibody or antigen-binding fragment thereof of the present disclosure is a chimeric antibody, a humanized antibody, or a fully human antibody. In some embodiments, the antibody or antigen-binding fragment thereof of the present disclosure is selected from an scFv, a Fab, a Fab', a (Fab')2, an Fv fragment, a disulfide-linked Fv (dsFv), and a diabody.

[0474] In some embodiments, the antibodies of the present disclosure are scFvs. In certain embodiments, the scFvs of the present disclosure are (a) VH set forth in SEQ ID NO: 3 and VL set forth in SEQ ID NO: 13; (b) VH set forth in SEQ ID NO: 23 and VL set forth in SEQ ID NO: 33; (c) VH set forth in SEQ ID NO: 3 and VL set forth in SEQ ID NO: 33; (d) VH set forth in SEQ ID NO: 23 and VL set forth in SEQ ID NO: 13; (e) a heavy chain variable region (VH) and a light chain variable region (VL), wherein the heavy chain variable region (VH) and the light chain variable region (VL) independently have 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%, or at least 99% sequence identity compared to the VH and VL in any one of groups (a) to (d), respectively; or (f) A heavy chain variable region (VH) and a light chain variable region (VL) that independently have one or more amino acid substitutions, deletions, or additions, or any combination thereof (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acid substitutions, deletions, or additions, or any combination thereof), compared to the VH and VL of any one of groups (a) to (d). Preferably, the substitutions are conservative substitutions.

[0475] In some embodiments, the antibodies of the present disclosure are scFvs. In certain embodiments, the scFvs of the present disclosure are (i) a sequence set forth in SEQ ID NO: 1 or 2; (ii) a sequence having one or more amino acid substitutions, deletions or additions, or any combination thereof, compared to the sequence set forth in (i) (e.g., up to 50, up to 45, up to 40, up to 35, up to 30, up to 25, up to 20, up to 15, up to 10, or up to 5 amino acid substitutions, deletions or additions, or any combination thereof, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acid substitutions, deletions or additions, or any combination thereof); or (iii) a sequence having 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%, or at least 99% sequence identity to the sequence set forth in (i); Preferably, the substitutions described in (ii) are conservative substitutions.

[0476] In some embodiments, an antibody of the present disclosure is an scFv.

[0477] In some embodiments, an scFv of the disclosure comprises the sequence set forth in SEQ ID NO: 1 or 2.

[0478] antibody derivative Antibodies or antigen-binding fragments thereof of the present disclosure may be derivatized, e.g., linked to another molecule (e.g., another polypeptide or protein). Generally, derivatization (e.g., labeling) of an antibody or antigen-binding fragment thereof does not adversely affect its binding to B7H3 (particularly human B7H3). Thus, the antibodies or antigen-binding fragments thereof of the present disclosure are also intended to include such derivatized forms. For example, antibodies or antigen-binding fragments thereof of the present disclosure may be linked (by chemical coupling, genetic fusion, non-covalent bonding, etc.) to one or more other molecular groups, such as another antibody (e.g., to form a bispecific antibody), a detection agent, a pharmaceutical agent, and / or a protein or polypeptide that can mediate binding of the antibody or antigen-binding fragment thereof to another molecule (e.g., avidin or a polyhistidine tag).

[0479] One type of derivatized antibody (e.g., bispecific antibodies) is produced by crosslinking two or more antibodies (belonging to the same or different types). Methods for obtaining bispecific antibodies are well known in the art and include, but are not limited to, chemical crosslinking, cell engineering (hybridoma), or genetic engineering.

[0480] Another type of derivatized antibody is a labeled antibody. For example, the antibodies or antigen-binding fragments thereof of the present disclosure can be conjugated to a detectable marker. The detectable marker described in this disclosure can be any substance detectable by fluorescent, spectroscopic, photochemical, biochemical, immunological, electrical, optical, or chemical means. Such markers are well known in the art, and examples include, but are not limited to, enzymes (e.g., horseradish peroxidase, alkaline phosphatase, β-galactosidase, urease, glucose oxidase, etc.), radionuclides (e.g., H, I, S, C, or P), fluorescent dyes (e.g., fluorescein isothiocyanate (FITC), fluorescein, tetramethylrhodamine isothiocyanate (TRITC), phycoerythrin (PE), Texas Red, rhodamine, quantum dots, or cyanine dye derivatives (e.g., Cy7, Alexa750)), acridine esters, magnetic beads (e.g., Dynabeads®), colorimetric markers (e.g., colloidal gold or colored glass or plastic (e.g., polystyrene, polypropylene, latex, etc.) beads), and biotin for binding avidin (e.g., streptavidin) modified with the above markers. Patents that teach the use of markers include, but are not limited to, U.S. Patent Nos. 3,817,837, 3,850,752, 3,939,350, 3,996,345, 4,277,437, 4,275,149, and 4,366,241 (all of which are incorporated herein by reference in their entirety). Detectable markers such as those described above can be detected by methods known in the art. For example, radioactive markers can be detected using photographic film or a scintillation counter, and fluorescent markers can be detected using a photodetector to detect emitted light. Enzymatic markers are generally detected by providing a substrate to the enzyme and detecting the reaction product produced by the action of the enzyme on the substrate, while colorimetric markers are detected by simple visualization of the colored marker.In certain embodiments, such labeling may be suitable for immunological detection (e.g., enzyme-linked immunoassay, radioimmunoassay, fluorescent immunoassay, chemiluminescent immunoassay, etc.) In certain embodiments, such detectable markers may be attached to the antibodies or antigen-binding fragments thereof of the present disclosure by linkers of different lengths to reduce potential steric hindrance.

[0481] In addition, the antibodies or antigen-binding fragments thereof of the present disclosure can be derivatized with chemical groups such as polyethylene glycol (PEG), methyl or ethyl, or glycosyl. These groups can be used to improve the biological properties of the antibody, such as increasing serum half-life.

[0482] The antigen-binding fragments of the present disclosure can be obtained by hydrolysis of intact antibody molecules (see Morimoto et al., J. Biochem. Biophys. Methods 24:107-117 (1992) and Brennan et al., Science 229:81 (1985)). In addition, these antigen-binding fragments can also be produced directly by recombinant host cells (reviewed in Hudson, Curr. Opin. Immunol. 11:548-557 (1999); Little et al., Immunol. Today, 21:364-370 (2000)). For example, Fab' fragments can be obtained directly from host cells. Fab' fragments can be chemically coupled to form F(ab')2 fragments (Carter et al., Bio / Technology, 10:163-167 (1992)). In addition, Fv, Fab, or F(ab')2 fragments can also be isolated directly from recombinant host cell culture. Other techniques for preparing these antigen-binding fragments are well known to those skilled in the art.

[0483] The IgG isotype control antibody of the present disclosure is well known to those skilled in the art and can be purchased or prepared. For example, the sequence of human anti-hen egg lysozyme IgG (anti-HEL, such as human IgG1, abbreviated as hIgG1) is derived from the variable region of the Fab F10.6.6 sequence in "Affinity maturation increases the stability and plasticity of the Fv domain of anti-protein antibodies" (Acierno et al. J Mol Biol. 2007;374(1):130-46) published by Acierno et al. The preparation method is as follows: Amino acid codon optimization and gene synthesis of the heavy and light chain (full sequence or variable region) genes of the human IgG antibody are performed by Nanjing GenScript Biotech Corp. according to the standard techniques introduced in "Molecular Cloning, A Laboratory Manual (3rd Edition)." Using standard molecular cloning techniques, such as PCR, enzyme digestion, DNA gel recovery, ligation transformation, colony PCR, or enzyme digestion identification, the heavy and light chain genes were subcloned into antibody heavy and light chain expression vectors in a mammalian expression system, respectively. The heavy and light chain genes in the recombinant expression vectors were then sequenced. After accurate sequence confirmation by sequencing, a number of endotoxin-free expression plasmids were prepared, and the heavy and light chain expression plasmids were transiently co-transfected into HEK293 cells for recombinant antibody expression. After 7 days of culture, the cell culture was harvested and purified using an rProtein A affinity column (GE). The quality of the collected antibody samples was determined using standard analytical techniques, such as SDS-PAGE and SEC-HPLC.

[0484] In a ninth aspect, the present disclosure provides a multispecific antibody comprising an antibody or antigen-binding fragment thereof according to any item of the eighth aspect of the present disclosure and an additional antibody or fragment thereof or antibody mimetic.

[0485] In certain embodiments, the multispecific antibody is a bispecific antibody or a trispecific antibody or a tetraspecific antibody.

[0486] Thus, in a tenth aspect, the present disclosure provides an isolated nucleic acid molecule comprising a nucleotide sequence encoding an antibody or antigen-binding fragment thereof according to the present disclosure, or the heavy and / or light chain variable region thereof, or one or more CDRs thereof. In certain embodiments, the nucleotide sequence may be substituted according to codon degeneracy, as known in the art. In certain embodiments, the nucleotide sequence is codon-optimized.

[0487] In certain embodiments, an isolated nucleic acid molecule of the present disclosure comprises (i) a first nucleic acid and a second nucleic acid encoding the heavy chain variable region and the light chain variable region, respectively, of an antibody or antigen-binding fragment thereof of the present disclosure, or (ii) a first nucleic acid and a second nucleic acid encoding the heavy chain variable region and the heavy chain constant region, and the light chain variable region and the light chain constant region, respectively, of an antibody or antigen-binding fragment thereof of the present disclosure, or (iii) a first nucleic acid and a second nucleic acid encoding the heavy chain and the light chain, respectively, of an antibody or antigen-binding fragment thereof of the present disclosure. In certain embodiments, the first nucleic acid and the second nucleic acid comprise nucleic acids that are degenerate sequences of or substantially identical to the sequences of the first and second nucleic acids in any of items (i) to (iii) above. In certain embodiments, a degenerate or substantially identical sequence refers to a sequence that has at least about 85%, 90%, 95%, or more sequence identity, or has one or more nucleotide substitutions, or has no more than 3, 6, 15, 30, or 45 nucleotide differences, compared to the nucleic acid molecule described in items (i) to (iii).

[0488] In an eleventh aspect, a vector (e.g., a cloning vector or an expression vector) is provided that comprises an isolated nucleic acid molecule of the present disclosure. In certain embodiments, the vector of the present disclosure is, for example, a plasmid, cosmid, bacteriophage, lentivirus, etc. In certain embodiments, the vector is capable of expressing an antibody or antigen-binding fragment thereof of the present disclosure in vivo in a subject (e.g., a mammal such as a human).

[0489] In a twelfth aspect, a host cell is provided comprising an isolated nucleic acid molecule of the present disclosure or a vector of the present disclosure. The host cell may be a eukaryotic cell (e.g., a mammalian cell, an insect cell, a yeast cell) or a prokaryotic cell (e.g., Escherichia coli). Suitable eukaryotic cells include, but are not limited to, NS0 cells, Vero cells, Hela cells, COS cells, CHO cells, HEK293 cells, BHK cells, and MDCKII cells. Suitable insect cells include, but are not limited to, Sf9 cells. In certain embodiments, the host cell of the present disclosure is a mammalian cell such as a CHO (e.g., CHO-K1, CHO-S, CHO DXB11, CHO DG44).

[0490] In a thirteenth aspect, there is provided a method for preparing an antibody or antigen-binding fragment thereof or a multispecific antibody according to the present disclosure, the method comprising culturing a host cell of the present disclosure under conditions that allow for expression of the antibody or antigen-binding fragment thereof or multispecific antibody, and recovering the antibody or antigen-binding fragment thereof from the cultured host cell culture.

[0491] In a fourteenth aspect, an antibody-drug conjugate is provided, wherein the antibody is an anti-B7H3 antibody as described above, wherein the antibody is linked by a linker to a coupling moiety selected from a detectable marker, a radioisotope, a fluorescent agent, a luminescent agent, a colorant, an enzyme, polyethylene glycol (PEG), a nuclide, a nucleic acid, a small molecule toxin, a polypeptide having binding activity, a protein, a receptor, a ligand, and other active agents that inhibit tumor cell proliferation and promote tumor cell apoptosis or necrosis.

[0492] In certain instances, the antibody-drug conjugate has the following formula: Ab-(LD)n, an anti-B7H3 antibody drug conjugate (B7H3-ADC) of During the ceremony, Ab is the antibody or antigen-binding fragment thereof of the eighth aspect; D is a small molecule toxin drug moiety; L is a bond or linker covalently linking Ab to D; n is an integer of 1 to 16 and represents the number of LDs covalently bound to Ab.

[0493] In some examples, L in the antibody-drug conjugate is selected from an amino acid residue or a short peptide consisting of 2 to 10 amino acid residues, and the amino acid residues can be selected from natural amino acid residues, unnatural amino acid residues, and AA 1 or a stereoisomer thereof, [ka] AA 1 In the amino acid residues represented by R a1 and R b are each independently H, [ka] Selected from or Or R a1 and R b together with the carbon atoms to which they are both attached form a 4-10 membered heterocyclic ring, in which the heteroatoms are selected from 1, 2, 3, or 4 O, N, and S, and the 4-10 membered heterocyclic ring is 0 is optionally replaced by r, r 1 , r 1a and r 1b are each independently selected from any integer of 0 to 20, R m1, R n1 , R m1a , R n1a , R m1b and R n1b are each independently H, C 1~6 Alkyl, C 3~6 Cycloalkyl and -COOR x1 Selected from R x1 is C 1~6 Is it alkyl? Or R m1 and R n1 , R m1a and R n1a , and R m1b and R n1b are taken together with the nitrogen atom to which they are both attached to form a 4-10 membered heterocyclic ring, in which the heteroatoms are selected from 1, 2, 3 or 4 O, N and S, and the 4-10 membered heterocyclic ring is also selected from one or more R 0’ is optionally replaced by R z is C 1~6 alkyl, R 0 , R 0’ are each independently C 1~6 Alkyl, C 3~6 Cycloalkyl, -NR m2 R n2 and C 1~6 and 4-10 membered heterocyclyl optionally substituted with alkyl, wherein in the 4-10 membered heterocycle the heteroatoms are selected from 1, 2, 3 or 4 O, N and S; R m2 , R n2 are each independently H and C 1~6 alkyl.

[0494] In some embodiments, AA 1 In the amino acid residue represented by a1 and R b together with the carbon atoms to which they are both attached, form a 5- to 6-membered heterocyclic ring in which the heteroatom is N and the number of heteroatoms is 1 or 2, and the 5- to 6-membered heterocyclic ring is 0The 5- to 6-membered heterocyclic ring is preferably a piperidine ring or a piperazine ring, and more preferably a piperidine ring, for example, [ka] and the carbon atom numbered 1 is R a1 and R b is the carbon atom to which both are bonded.

[0495] In some embodiments, r, r 1 , r 1a and r 1b are each independently 0, 1, 2, 3, 4, or 5, and preferably r, r 1 , r 1a and r 1b are each independently 0 or 4, more preferably r is 0 and r 1 , r 1a and r 1b is 4, or r is 4 and r 1 , r 1a and r 1b is 0.

[0496] In some embodiments, r, r 1 , r 1a , r 1b is never all 0.

[0497] In some embodiments, R m1 , R n1 , R m1a , R n1a , R m1b and R n1b are each independently H, C 1~6 Alkyl, or -COOR x1 and R x1 is C 1~6 alkyl, preferably each independently H, methyl, ethyl, n-propyl, -COOCH3, -COOCH2CH3, -COOCH2CH2CH3, -COOCH(CH3)2, -COOC(CH3)3 or -COOCH2CH2CH2CH3.

[0498] In some embodiments, R m1 and R n1 , R m1a and R n1a , and R m1b and R n1b together with the nitrogen atom to which they are both attached to form a 5- to 6-membered heterocycle in which the heteroatom is selected from 1 or 2 N atoms, and the 5- to 6-membered heterocycle is 0’ The 5- to 6-membered heterocycle is preferably a piperidine ring or a piperazine ring, more preferably [ka] and the nitrogen atom numbered 1 is R m1 and R n1 is the nitrogen atom to which both are bonded.

[0499] In some embodiments, R z is preferably C 1~6 It is alkyl, more preferably methyl.

[0500] In some embodiments, R 0 and R 0’ are each independently C 1~6 Alkyl, -NR m2 R n2 or C 1~6 A 5- to 6-membered heterocyclyl optionally substituted with alkyl, wherein the heteroatoms in the 5- to 6-membered heterocyclyl are selected from 1 or 2 N atoms.

[0501] In some embodiments, R 0 is preferably C 1~6 Alkyl or C 1~6 The 5- to 6-membered heterocyclyl is piperidinyl or piperazinyl, more preferably methyl or piperidinyl substituted with methyl, for example, methyl or [ka] is.

[0502] In some embodiments, each R 01’ are independently preferably C 1~6 Alkyl, -NR m2 R n2 or C 1~6 and 5- to 6-membered heterocyclyl optionally substituted with alkyl, wherein the heteroatoms are selected from 1 or 2 N atoms, and more preferably, each R 01’ is C 1~6 Alkyl or -NR m2 R n2 and more preferably, each R 01’ are independently methyl or -NR m2 R n2 and R m2 and R n2 are each independently preferably H or C 1~6 It is preferably alkyl, and more preferably methyl.

[0503] In some embodiments, R 0’ is preferably C 1~6 Alkyl or -NR m2 R n2 Yes, R m2 and R n2 are each independently preferably H or C 1~6 It is alkyl (eg, methyl).

[0504] In some embodiments, AA 1 In the amino acid residue of a1 and R b One of them is H and the other is [ka] and preferably R a1 and R b One of them is H and the other is [ka] is selected from.

[0505] In some embodiments, AA 1 The amino acid residue represented by [ka] and more preferably, [ka] and more preferably, [ka] and Most preferably, [ka] is.

[0506] In some embodiments, the antibody-drug conjugate consists of antibody-linker-drug, where the drug is selected from microtubule blocking drugs, DNA damaging drugs, Bcl-xL inhibitors, and the like.

[0507] In some embodiments, the antibody-drug conjugate consists of antibody-linker-drug, and the drug is selected from auristatins (e.g., MMAF, MMAE), maytansine derivatives (e.g., DM1, DM4), tubulysins, cryptomycins, antimitotic inhibitors, pyrrolobenzazepines and indole chlorobenzazepines, ducamycin, camptothecin, calicheamicin, and the like.

[0508] In some embodiments, the antibody-drug conjugate consists of an antibody-linker-drug, where the drug is selected from camptothecin (CPT) and its derivatives.

[0509] In some embodiments, the antibody-drug conjugate consists of an antibody-linker-drug, wherein the drug is selected from a topoisomerase I inhibitor.

[0510] In some embodiments, the antibody-drug conjugate consists of an antibody-linker-drug, wherein the drug is selected from SN-38 and DXd.

[0511] In some embodiments, D in the antibody-drug conjugate has Formula II: [ka] is a drug unit represented by During the ceremony, R1 is H or F R2 is H or methyl; or R1 and R2 together with the carbon atoms to which they are attached [ka] Forming R3 is H, R4 is selected from H, -(C1-C4 alkyl)-OH, -(C1-C4 alkenyl)-OH, -(C1-C4 alkyl)-NH2 or -(C1-C4 alkenyl)-NH2; L is linked by a hydroxyl or amine group in D.

[0512] In some embodiments, L in the antibody-drug conjugate is selected from: [ka] Position 1 binds to Ab, position 2 binds to D, and AA1 is as defined above.

[0513] In addition, it should be noted, and can be understood by those skilled in the art, that L is linked to a sulfhydryl group contained in Ab (e.g., antibody) after the disulfide bond is cleaved (e.g., reduction of the disulfide bond with the reducing agent TCEP can cleave the disulfide bond to generate a sulfhydryl -SH group), i.e., -S- between L and Ab is not an additional foreign sulfur atom. For example, in the formula, -S- is not an additional foreign sulfur atom, but is -S- formed by linking a sulfhydryl group contained in Tb after the disulfide bond is opened to L.

[0514] In some preferred embodiments, the antibody-drug conjugate is selected from: [Table 6-1] [Table 6-2] wherein Ab is anti-B7H3 antibody 1D1-01 or 2E3-02, n is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, and q is preferably 1, 2, 3, 4, 5, 6, 7, 8.

[0515] In a fifteenth aspect of the present disclosure, the present disclosure provides a ligand drug conjugate comprising the aforementioned ligand drug conjugate or the aforementioned antibody-drug conjugate. The ligand drug conjugate has a q value of two or more, and the antibody-drug conjugate has an n value of two or more. The ligand drug conjugate refers to a ligand drug conjugate or antibody-drug conjugate (ADC) comprising a heterogeneous DAR distribution.

[0516] In some embodiments, when a majority of Ligand Drug Conjugates have a particular q or n value, the q or n value is close to the DAR.

[0517] In some embodiments, the drug-antibody ratio (DAR) in the ligand drug conjugate is selected from integers or decimals between 1 and 10.

[0518] In some embodiments, the drug-antibody ratio (DAR) in the ligand drug conjugate is selected from 2, 2.5, 3, 3.5, 4, 4.5, 5, 5.5, 6, 6.5, 7, 7.2, 7.4, 7.5, 7.6, 7.7, 7.8, 7.9, 8.0, 8.1, 8.2, 8.3, 8.4, 8.5, 8.7, 8.9 and 9.

[0519] In some embodiments, the ligand drug conjugate ADCs include ADCs with a DAR distribution of 1 to 8, e.g., 1.5, 2, 4, 6, and 8 (i.e., 1.5, 2, 4, 6, and 8 payload species). It should be noted that degradation products may be generated such that the ligand drug conjugates may also contain DARs of 1, 3, 5, and 7. Additionally, the ADCs in the ligand drug conjugates may also have a DAR greater than 8. The ligand drug conjugates are generated by reduction of interchain disulfides followed by coupling. In some embodiments, the ligand drug conjugates include both of the following: ligand drug conjugates with a DAR of 4 or less (i.e., payload species of 4 or less) and ligand drug conjugates with a DAR of 6 or greater (i.e., payload species of 6 or more).

[0520] In another aspect, the present disclosure provides use of an antibody or antigen-binding fragment thereof of the present disclosure in the manufacture of a kit for detecting the presence or level of B7H3 in a sample. In another aspect, the present disclosure provides a diagnostic or therapeutic kit comprising one or more of the following agents: an antibody or antigen-binding fragment thereof, a nucleic acid, a vector, a host cell, a multispecific antibody, a conjugate, or a pharmaceutical composition of the present disclosure. Optionally, the diagnostic or therapeutic kit further comprises instructions for use.

[0521] In a sixteenth aspect of the present disclosure, the present disclosure provides a pharmaceutical composition comprising a substance A and optionally one or more pharmaceutically acceptable adjuvants, wherein the substance A is the aforementioned ligand drug conjugate, or a stereoisomer of the ligand drug conjugate, a prodrug thereof, a pharmaceutically acceptable salt thereof, or a pharmaceutically acceptable solvate thereof, or the aforementioned antibody or antigen-binding fragment thereof, a nucleic acid molecule, a vector, a host cell, a multispecific antibody, and / or a ligand drug conjugate, a conjugate, or the aforementioned antibody-drug conjugate, or the aforementioned compound, or a stereoisomer of the compound, a prodrug thereof, a pharmaceutically acceptable salt thereof, or a pharmaceutically acceptable solvate thereof. The substance A may be in a therapeutically effective amount.

[0522] In certain embodiments, a pharmaceutical composition of the present disclosure comprises an antibody or antigen-binding fragment thereof of the present disclosure and a pharmaceutically acceptable carrier and / or excipient.

[0523] In certain embodiments, a pharmaceutical composition of the present disclosure comprises a host cell of the present disclosure and a pharmaceutically acceptable carrier and / or excipient, wherein the host cell comprises the isolated nucleic acid molecule or vector described above.

[0524] In certain embodiments, a pharmaceutical composition of the present disclosure comprises a multispecific antibody of the present disclosure and a pharmaceutically acceptable carrier and / or excipient.

[0525] In certain embodiments, a pharmaceutical composition of the present disclosure comprises a conjugate of the present disclosure and a pharmaceutically acceptable carrier and / or excipient.

[0526] In a seventeenth aspect of the present disclosure, the present disclosure provides the aforementioned substance A or the aforementioned pharmaceutical composition in the manufacture of a medicament for the treatment and / or prevention of a disease associated with abnormal cell activity (e.g., a cancer disease), wherein the substance A or the aforementioned pharmaceutical composition may be in a therapeutically effective amount.

[0527] In some examples, the present disclosure provides for the use of an antibody or antigen-binding fragment thereof, nucleic acid, vector, host cell, antibody drug conjugate, or multispecific antibody according to the present disclosure in the manufacture of a medicament, wherein the medicament is used to modulate (inhibit or block) the activity of B7H3.

[0528] In some examples, the present disclosure provides for the use of an antibody or antigen-binding fragment thereof, nucleic acid, vector, host cell, antibody drug conjugate, or multispecific antibody according to the present disclosure in the manufacture of a medicament, wherein the medicament is used for the treatment or prevention of a disease associated with B7H3 activity. In some examples, the present disclosure provides use of an antibody or antigen-binding fragment thereof, nucleic acid, vector, host cell, antibody drug conjugate, or multispecific antibody according to the present disclosure in the manufacture of a medicament, wherein the medicament is used for the treatment or prevention of a tumor associated with B7H3 activity.

[0529] In some embodiments, the cancer disease is selected from esophageal cancer (e.g., esophageal adenocarcinoma and esophageal squamous cell carcinoma), brain tumor, lung cancer (e.g., small cell lung cancer, non-small cell lung cancer, or lung adenocarcinoma), squamous cell carcinoma, bladder cancer, gastric cancer, ovarian cancer, peritoneal cancer, pancreatic cancer, breast cancer, head and neck cancer, cervical cancer, endometrial cancer, colon cancer (e.g., human colon adenocarcinoma), rectal cancer, colorectal cancer, liver cancer, kidney cancer, urothelial cancer, epidermal cancer, non-Hodgkin's lymphoma, central nervous system tumors (e.g., neuroglioma, glioblastoma multiforme, glioma, or sarcoma), prostate cancer, or thyroid cancer.

[0530] In some embodiments, the cancer disease is a cancer disease associated with Trop-2, Her2, B7H3, Her3, EGFR, hi some embodiments, the cancer disease is a cancer disease associated with Trop-2 or Her2.

[0531] In some embodiments, the cancer disease is a Trop-2 associated cancer disease.

[0532] In some embodiments, the cancer disease is a Her3-associated cancer disease.

[0533] In some embodiments, the cancer disease is an EGFR-associated cancer disease.

[0534] In some embodiments, the cancer disease is a B7H3-associated cancer disease.

[0535] In some embodiments, the cancer disease is a solid tumor.

[0536] In some embodiments, the cancer disease is breast cancer or lung cancer (preferably non-small cell lung cancer).

[0537] In a seventeenth aspect of the present disclosure, the present disclosure provides the aforementioned substance A or the aforementioned pharmaceutical composition for use in the treatment and / or prevention of a disease associated with abnormal cellular activity (e.g., a cancer disease).

[0538] In some embodiments, the cancer disease is selected from esophageal cancer (e.g., esophageal adenocarcinoma and esophageal squamous cell carcinoma), brain tumor, lung cancer (e.g., small cell lung cancer, non-small cell lung cancer, or lung adenocarcinoma), squamous cell carcinoma, bladder cancer, gastric cancer, ovarian cancer, peritoneal cancer, pancreatic cancer, breast cancer, head and neck cancer, cervical cancer, endometrial cancer, colon cancer (e.g., human colon adenocarcinoma), rectal cancer, colorectal cancer, liver cancer, kidney cancer, urothelial cancer, epidermal cancer, non-Hodgkin's lymphoma, central nervous system tumors (e.g., glioma, glioblastoma multiforme, glioma, or sarcoma), prostate cancer, or thyroid cancer.

[0539] In some embodiments, the cancer disease is a cancer disease associated with Trop-2, Her2, B7H3, Her3, EGFR, hi some embodiments, the cancer disease is a cancer disease associated with Trop-2 or Her2.

[0540] In some embodiments, the cancer disease is a Trop-2 associated cancer disease.

[0541] In some embodiments, the cancer disease is a Her3-associated cancer disease.

[0542] In some embodiments, the cancer disease is an EGFR-associated cancer disease.

[0543] In some embodiments, the cancer disease is a B7H3-associated cancer disease.

[0544] In some embodiments, the cancer disease is a solid tumor. In some embodiments, the cancer disease is breast cancer or lung cancer (preferably non-small cell lung cancer).

[0545] In an eighteenth aspect of the present disclosure, the present disclosure provides a method for preventing and / or treating a disease associated with abnormal cellular activity (e.g., a cancer disease), comprising administering a prophylactically and / or therapeutically effective amount of the aforementioned substance A or the aforementioned pharmaceutical composition to an individual in need thereof.

[0546] In some embodiments, the cancer disease is selected from esophageal cancer (e.g., esophageal adenocarcinoma and esophageal squamous cell carcinoma), brain tumor, lung cancer (e.g., small cell lung cancer, non-small cell lung cancer, or lung adenocarcinoma), squamous cell carcinoma, bladder cancer, gastric cancer, ovarian cancer, peritoneal cancer, pancreatic cancer, breast cancer, head and neck cancer, cervical cancer, endometrial cancer, colon cancer (e.g., human colon adenocarcinoma), rectal cancer, colorectal cancer, liver cancer, kidney cancer, urothelial cancer, epidermal cancer, non-Hodgkin's lymphoma, central nervous system tumors (e.g., glioma, glioblastoma multiforme, glioma, or sarcoma), prostate cancer, or thyroid cancer.

[0547] In some embodiments, the cancer disease is a cancer disease associated with Trop-2, Her2, B7H3, Her3, or EGFR.

[0548] In some embodiments, the cancer disease is a Trop-2 or Her2 associated cancer disease.

[0549] In some embodiments, the cancer disease is a Trop-2 associated cancer disease.

[0550] In some embodiments, the cancer disease is a Her3-associated cancer disease.

[0551] In some embodiments, the cancer disease is an EGFR-associated cancer disease.

[0552] In some embodiments, the cancer disease is a B7H3-associated cancer disease.

[0553] In some embodiments, the cancer disease is a solid tumor.

[0554] In some embodiments, the cancer disease is breast cancer or lung cancer (preferably non-small cell lung cancer).

[0555] In this disclosure, unless otherwise specified, scientific and technical terms used herein have the meanings commonly understood by those skilled in the art. Furthermore, cell culture, molecular genetics, nucleic acid chemistry, and immunology laboratory procedures used herein are all routine procedures widely used in the corresponding fields. Meanwhile, for a better understanding of the present invention, definitions and explanations of relevant terms are provided below.

[0556] As used herein, the term "pharmaceutically acceptable salt" refers to an organic acid addition salt formed using an organic acid that provides a pharmaceutically acceptable anion, including, but not limited to, formate, acetate, propionate, benzoate, maleate, fumarate, succinate, tartrate, citrate, ascorbate, α-ketoglutarate, α-glycerophosphate, alkylsulfonate, or arylsulfonate, preferably, the alkylsulfonate is methylsulfonate or ethylsulfonate, and the arylsulfonate is benzenesulfonate or p-toluenesulfonate. Suitable inorganic salts may also be formed, including, but not limited to, hydrochloride, hydrobromide, hydroiodide, nitrate, bicarbonate, carbonate, sulfate, or phosphate.

[0557] As used herein, 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 active ingredient, and is known in the art (see, e.g., Remington's Pharmaceutical Sciences. Edited by Gennaro AR, 19th ed. Pennsylvania: Mack Publishing Company, 1995), and includes, but is not limited to, pH adjusters, surfactants, adjuvants, ionic strength enhancers, diluents, agents for maintaining osmotic pressure, agents for delaying absorption, and preservatives.

[0558] Pharmaceutically acceptable salts can be obtained using standard procedures well known in the art, for example, by reacting a sufficient amount of a basic compound with a suitable acid to provide a pharmaceutically acceptable anion.

[0559] In this disclosure, pharmaceutically acceptable adjuvants refer to excipients and additives used in the manufacture of medicines and formulations, and refer to substances contained in pharmaceutical formulations other than active ingredients, whose safety has been reasonably evaluated. In addition to shaping, acting as a carrier, and improving stability, pharmaceutically acceptable adjuvants also have important functions such as solubilization, hydrotropy, sustained release, and controlled release, and are important components that can affect the quality, safety, and efficacy of drugs. According to their source, they can be classified into natural, semi-synthetic, and fully synthetic. According to their functions and uses, they can be classified into solvents, propellants, solubilizers, cosolvents, emulsifiers, colorants, binders, disintegrants, fillers, lubricants, wetting agents, osmotic pressure adjusters, stabilizers, glidants, flavorings, preservatives, suspending agents, coating materials, fragrances, anti-adhesive agents, antioxidants, chelating agents, penetration enhancers, pH adjusters, buffers, plasticizers, surfactants, foaming agents, antifoaming agents, thickeners, clathrates, humectants, absorbents, diluents, flocculants and deflocculants, filter aids, release blockers, etc. According to their administration routes, they can be classified into oral, injection, mucosal, transdermal or topical administration, nasal or oral inhalation administration, and intraocular administration. The same pharmaceutically acceptable adjuvant can be used in pharmaceutical preparations with different administration routes and has different effects and uses.

[0560] The pharmaceutical composition can be prepared into various suitable dosage forms according to the route of administration. For example, tablets, capsules, granules, oral liquids, oral suspensions, oral emulsions, powders, tinctures, syrups, injections, suppositories, ointments, creams, pastes, ophthalmic preparations, pills, implants, aerosols, powder aerosols, sprays, etc., and the pharmaceutical composition or suitable dosage form may contain 0.01 mg to 1000 mg, appropriately 0.1 mg to 800 mg, preferably 0.5 mg to 500 mg, preferably 0.5 mg to 350 mg, particularly preferably 1 mg to 250 mg of the compound of the present disclosure or its pharmaceutically acceptable salt or conjugate.

[0561] The pharmaceutical composition can be administered in the form of injection, including injection solution, sterile powder for injection and concentrated solution for injection.Among the above, useful vehicles and solvents are water, Ringer's solution and physiological saline solution.In addition, sterile non-volatile oils (such as monoglycerides or diglycerides) can also be used as solvents or suspension media.

[0562] The term "treatment," as used herein, generally refers to obtaining a desired pharmacological and / or physiological effect. The effect can be preventative, in terms of complete or partial prevention of a disease or its symptoms, and / or therapeutic, in terms of partial or complete stabilization or cure of the disease and / or side effects due to the disease. "Treatment," as used herein, encompasses any treatment of a disease in a patient, including (a) prevention of a disease or condition in a patient who is susceptible to the disease or condition but has not yet been diagnosed as having the disease or condition, (b) suppression of symptoms of the disease, i.e., prevention of its onset, or (c) alleviation of symptoms of the disease, i.e., regression of the disease or condition.

[0563] In this disclosure, the term "individual" includes a human or a non-human animal. Exemplary human individuals include human individuals (referred to as patients) with a disease (e.g., a disease described herein) or normal individuals. In this disclosure, the term "non-human animal" includes all vertebrates, such as non-mammals (e.g., birds, amphibians, reptiles), and mammals, such as non-human primates, farm animals and / or domesticated animals (e.g., sheep, dogs, cats, cows, pigs, etc.).

[0564] In the present disclosure, the term "effective dose" refers to that amount of a compound which, upon administration, will relieve to some extent one or more of the symptoms of the disorder being treated.

[0565] In the present disclosure, the term "ligand drug conjugate" refers to a substance in which a biologically active molecule (drug molecule) is linked to a targeting moiety. In some embodiments of the present disclosure, the biologically active molecule is linked to the targeting moiety by a linker. The linker can be cleaved under certain circumstances (e.g., in the presence of hydrolases and / or low pH in tumors) or under certain actions (e.g., the action of lysosomal proteases) so that the biologically active molecule is separated from the targeting moiety. In some embodiments of the present disclosure, the linker comprises a cleavable or non-cleavable unit, such as a peptide or disulfide bond. In some embodiments of the present disclosure, the biologically active molecule is directly linked to the targeting moiety by a covalent bond that can be cleaved under certain circumstances or actions, thereby separating the biologically active molecule from the targeting moiety. In some embodiments of the present disclosure, the ligand drug conjugate comprises a targeting moiety, a linker, and a compound fragment of Formula II of the present disclosure.

[0566] In the present disclosure, the terms "biologically active substance," "biologically active molecule," or "drug molecule" refer to a substance that inhibits or prevents the function of a cell and / or causes cell death or destruction. In some embodiments of the present disclosure, the biologically active substance, biologically active molecule, or drug molecule in the conjugate is a molecule with anti-tumor biological activity. For example, a radioisotope, e.g., At 211 , I 131 , I 125 , Y 90 ,Re 186 ,Re 188 , Sm 153 , Bi 212 , P 32 , Pb 212and Lu, metal complexes, such as metal platinum complexes and metal gold complexes, oxaliplatin, glycopeptide antibiotics, such as bleomycin and pingyangmycin, DNA topoisomerase inhibitors, such as topoisomerase I inhibitors, camptothecin, hydroxylcamptothecin, 9-aminocamptothecin, SN-38, irinotecan, topotecan, belotecan, rubitecan, topoisomerase II inhibitors, actinomycin D, adriamycin, doxorubicin, duocarmycin, daunorubicin, mitoxantrone, podophyllotoxin, etoposide, drugs that inhibit DNA synthesis, such as methotrexate, 5-fluorouracil, cytarabine, gemcitabine, mercaptopurine, pentostatin, fludarabine, cladribine, nelarabine, drugs that act on structural proteins, such as tubulin inhibitors, vinca alkaloids, These include tumor signaling pathway inhibitors such as vincristine, vinblastine, paclitaxel, docetaxel, and cabazitaxel, for example, serine / threonine kinase inhibitors, tyrosine kinase inhibitors, aspartate kinase inhibitors, or histidine kinase inhibitors, proteasome inhibitors, histone deacetylase inhibitors, tumor angiogenesis inhibitors, cyclin inhibitors, maytansine derivatives, calicheamicin derivatives, auristatin derivatives, pyrrolobenzodiazepine (PBD) derivatives, melphalan, mitomycin C, chlorambucil, or other active substances that inhibit tumor cell proliferation and promote tumor cell apoptosis and necrosis, enzymes such as nucleolysin and fragments thereof, antibiotics, toxins such as small molecule toxins or enzymatically active toxins derived from bacteria, fungi, plants, or animals (including fragments and / or variants thereof), growth inhibitors, and drug modules. The term "toxin" refers to a substance that can have a deleterious effect on cell growth or proliferation.

[0567] In this disclosure, the term "small molecule" refers to a biologically active small molecule drug. The term small molecule toxin has cell-damaging activity, i.e., a state that causes pathological changes in cells in several forms, and cell damage is not limited to direct damage but includes any kind of damage to the structure and function of cells, such as DNA breakage, base dimer formation, chromosome breakage, damage to the cell division machinery, and reduction of various enzyme activities.

[0568] In this disclosure, the term "linker" refers to a fragment that connects a bioactive molecule (drug molecule) to a targeting moiety.

[0569] In this disclosure, the term "targeting moiety" refers to a moiety in a conjugate that can specifically bind to a target (or a portion of a target) on a cell surface. Through the interaction of the targeting moiety with the target, the conjugate can be delivered to a specific cell population.

[0570] In the present disclosure, when the targeting moiety in the conjugate is an antibody, the conjugate may be referred to as a "drug-antibody conjugate."

[0571] In the present disclosure, antibodies or antigen-binding fragments thereof include derivatized antibodies or antigen-binding fragments thereof, such as antibodies or antigen-binding fragments thereof bearing thiol groups, where the derivatization allows the antibody to have a group or ability to react with a drug-linker conjugate. The thiol -SH can be derivatized by cleaving a disulfide bond (e.g., by reduction with the reducing agent TCEP).

[0572] As used herein, the terms "cancer" and "tumor" are used interchangeably.

[0573] As used herein, the term "gene" includes not only DNA, but also its mRNA, its cDNA, and its cRNA.

[0574] As used herein, the term "polynucleotide" is used interchangeably with nucleic acid and includes DNA, RNA, probes, oligonucleotides, and primers.

[0575] As used herein, the term "Tb" is an abbreviation for "target binding" and includes an antibody or any molecule that binds to a target, and the term "Ab" is an abbreviation for "antibody" and is used interchangeably with "antibody."

[0576] As used herein, the terms "polypeptide" and "protein" are used interchangeably.

[0577] As used herein, the term "cell" also includes cells within an individual animal and cells in culture.

[0578] The B7H3 involved in the anti-B7H3 antibodies described in this disclosure may be any B7H3 conventional in the art, such as soluble B7H3, membrane-form B7H3, and also referred to as B7H3 variant 1 and / or B7H3 variant 2.

[0579] KD refers to the dissociation constant, which is obtained from the ratio of Kd (the dissociation rate of a specific binding molecule-target protein interaction) to Ka (the association rate of a specific binding molecule-target protein interaction) (or Kd / Ka, expressed as molar concentration (M)). KD values ​​can be determined using methods well established in the art. A preferred method for determining the KD of a binding molecule is by surface plasmon resonance, for example, using a biosensor system such as a Biacore™ (GE Healthcare Life Sciences) system.

[0580] The term "B7H3 variant" refers to a polypeptide that has a similar or identical function to a B7H3 polypeptide, a B7H3 fragment, an anti-B7H3 antibody, or an antibody fragment thereof, but does not necessarily contain the amino acid sequence of the similar or identical B7H3 polypeptide, a B7H3 fragment, an anti-B7H3 antibody, or a fragment thereof, or has the structure of the similar or identical B7H3 polypeptide, a B7H3 fragment, an anti-B7H3 antibody, or a fragment thereof.

[0581] The term "Her3 variant" refers to a polypeptide that has a similar or identical function to a HER3 polypeptide, a HER3 fragment, an anti-HER3 antibody or an antibody fragment thereof, but does not necessarily contain the amino acid sequence of the similar or identical HER3 polypeptide, HER3 fragment, anti-HER3 antibody or fragment thereof, or the structure of the similar or identical HER3 polypeptide, HER3 fragment, anti-Her3 antibody or fragment thereof.

[0582] As used herein, the percent homology between two amino acid sequences is equal to the percent identity between the two sequences. The percent sequence identity between two sequences is a function of the number of identical positions shared by the sequences, taking into account the number of gaps that need to be introduced for optimal alignment of the two sequences and the length of each gap (i.e., % homology = number of identical positions / total number of positions × 100). Sequence comparison and determination of the percent identity between sequences can be performed by methods generally known in the art, and such sequence comparison and determination of the percent identity can be achieved by a mathematical algorithm. For example, the algorithm in Meyers and Miller, 1988, Comput. Appl. Biosci. 4:11-17 (incorporated into the ALIGN program (version 2.0)) can be used to determine the percent identity between amino acid sequences and / or nucleotide sequences. In addition, the GAP program (using its default parameters) in the GCG package available online from Accelrys can be used to determine the percent identity between amino acid sequences or nucleotide sequences. In one embodiment, the two sequences are of equal length.

[0583] The term "epitope" refers to a portion of a B7H3 polypeptide or protein that has antigenic or immunogenic activity in an animal, preferably a mammal. The epitope of an anti-B7H3 antibody or antigen-binding fragment thereof of the present disclosure can be determined by existing techniques, such as synthetic peptide methods, immunoinformatics prediction, polypeptide activity determination, epitope peptide scanning, phage display technology, X-ray diffraction and nuclear magnetic resonance analysis, homologous antibody modeling, and protein docking prediction. As used herein, the phrase "antibodies that bind to the same epitope" refers to different antibodies that bind to a common epitope. A first antibody and a second antibody can be determined to bind to the same epitope if the second antibody binds to a portion of the peptide or tertiary structure to which the first antibody binds.

[0584] In this disclosure, the term "antibody" is used in its broadest sense to include intact monoclonal antibodies, polyclonal antibodies, and multispecific antibodies (e.g., bispecific antibodies) formed from at least two intact antibodies, so long as they possess all the necessary biological activity. In this disclosure, "antibody" and "immunoglobulin" are used interchangeably. As used herein, "antibody molecule" or "antibody" refers to immunoglobulin molecules and immunologically active portions of immunoglobulin molecules, i.e., molecules that contain an antigen-binding site that enables them to immunospecifically bind to an antigen. Thus, the term antibody broadly encompasses not only whole antibody molecules, but also fragments of such antibodies and variants (including derivatives) of such antibodies and antibody fragments. When "antibody molecule" or "antibody" is used in the same context as an antigen-binding fragment, "antibody molecule" or "antibody" refers to an intact antibody molecule or a full-length antibody. The term antibody molecule referred to herein includes, but is not limited to, for example, single-chain Fv (scFv), Fab fragment, Fab' fragment, F(ab'), disulfide-linked Fv (sdFv), Fv, and intact or full-length antibodies. The term "single-chain Fv" or "scFv" refers to a polypeptide comprising an antibody VL domain linked to an antibody VH domain. For example, an antibody that immunospecifically binds to B7H3 may cross-react with other antigens, preferably not cross-react with other antigens. Antibodies that immunospecifically bind to B7H3 can be identified, for example, by immunoassays or other methods known to those of skill in the art. An "intact" or "full-length" antibody refers to a protein containing two heavy (H) chains and two light (L) chains linked together by disulfide bonds, which protein includes: (1) for the heavy chain, a heavy chain variable region (abbreviated herein as "VH") containing three domains CH1, CH2, CH3 and a constant region; and (2) for the light chain, a light chain variable region (abbreviated herein as "VL") and a light chain constant region containing domain CL.Antibodies of the present disclosure include, but are not limited to, monoclonal antibodies, multispecific antibodies, human or chimeric antibodies, single-chain antibodies, Fab fragments, F(ab') fragments, anti-idiotypic (anti-Id) antibodies (including, for example, the anti-Id antibodies of the present disclosure), and epitope-binding fragments of any of the foregoing antibodies. Immunoglobulin molecules of the present disclosure can be of any type (e.g., IgG, IgE, IgM, IgD, IgA, and IgY), any class (e.g., IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2), or any subclass of immunoglobulin. Preferably, antibodies of the present disclosure comprise or consist of a VH domain, a VH CDR (herein referred to as HCDR), a VL domain, or a VL CDR (often referred to herein as LCDR) having any of the amino acid sequences set forth in the Table of Sequences and Specific Information thereof, or fragments or variants thereof.

[0585] In this disclosure, the term "monoclonal antibody" refers to an antibody from a population of substantially homogeneous antibodies, i.e., each antibody in the population is identical except for possible minor natural mutations. Monoclonal antibodies have high specificity for one determinant (epitope) on an antigen, whereas polyclonal antibodies contain different antibodies directed against different determinants (epitopes). In addition to specificity, an advantage of monoclonal antibodies is that they may be synthesized without contamination by other antibodies. As used herein, the modifier "monoclonal" means that the antibody is characterized as being derived from a substantially homogeneous antibody population and should not be construed as requiring a particular method of preparation.

[0586] In some embodiments of the present disclosure, monoclonal antibodies specifically include chimeric antibodies, i.e., a portion of the heavy and / or light chain is identical to or homologous to one type, class, or subclass of antibody, and the remainder is identical to or homologous to another type, class, or subclass of antibody, so long as they possess the desired biological activity (see, e.g., U.S. Pat. No. 4,816,567 and Morrison et al., 1984, PNAS, 81:6851-6855). Chimeric antibodies useful in the present disclosure include primatized antibodies comprising variable region antigen-binding sequences derived from a non-human primate (e.g., an ancient monkey, chimpanzee, etc.) and human constant region sequences.

[0587] The term "antigen-binding fragment" refers to a portion of an antibody, preferably the antigen-binding or variable region. Examples of antibody fragments include Fab, Fab', F(ab')2, Fd, Fv, dAb, and complementarity-determining region fragments, diabodies, linear antibodies, and single-chain antibody molecules. As used herein, the term "antigen-binding fragment" refers to a partial fragment of an antibody that retains antigen-binding activity. This fragment retains complete or partial functionality of the antibody, and includes, but is not limited to, single-chain Fv (scFv), Fab, Fab', F(ab')2, disulfide-linked Fv (sdFv), Fv, and dis-scFv. This term also includes Fab', a monovalent fragment of the antibody variable region obtained by treating F(ab')2 under reducing conditions. However, this term is not limited to these molecules, as long as the fragment retains binding affinity for the antigen. Furthermore, these functional fragments include not only fragments obtained by treating a full-length antibody protein molecule with an appropriate enzyme, but also proteins produced in a suitable host cell using genetically engineered antibody genes.

[0588] As used herein, the term "Fab" refers to a monovalent fragment of the variable region of an antibody obtained by treating F(ab')2 under reducing conditions as described above. However, Fab' in the present disclosure also includes Fab' produced using genetically engineered antibody genes.

[0589] As used herein, the term "scFv" refers to a single polypeptide chain comprising a VL and a VH domain, where the VL and VH are linked by a linker or directly (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, Eds. Roseburg and Moore, Springer-Verlag, New York, pp. 269-315 (1994)). Such scFv molecules can have the general structure: NH2-VL-linker-VH-COOH or NH2-VH-linker-VL-COOH. Suitable linkers in the prior art consist of a repeated GGGGS amino acid sequence or a variant thereof. For example, a linker having the amino acid sequence (GGGGS)4 can be used, and variants thereof can also be used (Holliger et al. (1993), Proc. Natl. Acad. Sci. USA 90:6444-6448). Other linkers that can be used in the present disclosure were described in 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 can also exist between the VH and VL of an scFv. As used herein, the term "di-scFv" refers to an antibody fragment formed by linking two scFvs.

[0590] As used herein, an antibody or antigen-binding fragment thereof may contain mutations, amino acid substitutions, deletions, or additions, yet still have the activity of binding to an antigen.

[0591] The term "bispecific antibody," also known as "bifunctional antibody conjugate," refers to a conjugate formed by a first antibody (fragment) and a second antibody (fragment) via coupling arms, wherein the conjugate retains the activity of each antibody and is therefore bifunctional and bispecific.

[0592] The term "multispecific antibody" includes, for example, triabodies, which are antibodies with three different antigen-binding specificities, and tetrabodies, which are antibodies with four different antigen-binding specificities.

[0593] The term "intact antibody" or "full length antibody" refers to an antibody comprising an antigen-binding variable region and a light chain constant region (CL), a heavy chain constant region (CH1, CH2, and CH3). The constant region may be a native sequence (e.g., a human native constant region sequence) or a variant thereof. An intact antibody preferably has one or more effector functions.

[0594] The term "probody" refers to a modified antibody, including an antibody or antibody fragment, that is capable of specifically binding to its target and that can be coupled to a masking group, the masking group meaning that the cleavage constant for the binding ability of the antibody or antibody fragment with the masking group to its target is at least 100-fold or 1000-fold or 10,000-fold greater than that of the antibody or antibody fragment without the masking group.

[0595] In this disclosure, "humanized" forms of non-human (e.g., murine) antibodies refer to chimeric antibodies containing minimal non-human immunoglobulin sequence. Most humanized antibodies are human recipient immunoglobulins with hypervariable region residues replaced by non-human (e.g., mouse, rat, rabbit, or non-human primate) hypervariable region residues (donor antibody) having the desired specificity, affinity, and function. In some embodiments, framework region (FR) residues of the human immunoglobulin are also replaced with non-human residues. Furthermore, humanized antibodies may also comprise residues that are not found in the recipient antibody or the donor antibody. These modifications are intended to further optimize antibody performance. Humanized antibodies generally contain at least one, and usually two, variable regions, with all or nearly all of the hypervariable loops corresponding to those of a non-human immunoglobulin and the FRs representing complete or nearly complete human immunoglobulin protein sequences. Humanized antibodies may also comprise at least a portion of an immunoglobulin constant region (Fc, typically a human immunoglobulin Fc). For further details, see, e.g., Jones et al., 1986, Nature, 321:522-525; Riechmann et al., 1988, Nature, 332:323-329; and Presta, 1992, Curr Op Struct Bwl 2:593-596.

[0596] Intact antibodies can be assigned to different "classes" based on the amino acid sequence of their heavy chain constant region. The five main classes are IgA, IgD, IgE, IgG, and IgM, and some of these can be further divided into different "subclasses" (isotypes) (e.g., IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2). The heavy chain constant regions of the different classes of antibodies are called α, β, ε, γ, and μ, respectively. The subunit structures and three-dimensional configurations of different classes of immunoglobulins are well known in the art.

[0597] Herein, the CDRs contained in the antibody or antigen-binding fragment thereof of the present disclosure can be identified according to various numbering systems known in the art. In certain embodiments, the CDRs contained in the antibody or antigen-binding fragment thereof of the present disclosure are preferably determined according to the Kabat, Chothia, AbM or IMGT numbering system. According to the IMGT numbering system, the corresponding numbers of CDRs and FRs are as follows: [Table 7] As used herein, the term "framework region" or "FR residues" refers to amino acid residues in an antibody variable region other than the CDR residues as defined above.

[0598] As used herein, the term "germline antibody gene" refers to a gene encoding an immunoglobulin expressed by a non-lymphocyte that has not undergone the maturation process that results in gene rearrangement and maturation of the particular immunoglobulin expressed. One advantage provided by various embodiments of the present disclosure resides in the recognition that the amino acid sequences encoded by germline antibody genes retain more characteristic and important amino acid sequence structures of an individual of an animal species than those encoded by mature antibody genes, and are therefore less likely to be recognized as foreign by that species when applied therapeutically to that species.

[0599] Conservative amino acid substitutions are preferred herein for amino acid substitutions. Conservative amino acid substitutions refer to substitutions that occur within a set of amino acids that are related in their side chains. Preferred amino acid sets are as follows: the acidic set (aspartic acid and glutamic acid), the basic set (lysine, arginine, and histidine), the nonpolar set (alanine, valine, leucine, isoleucine, proline, phenylalanine, methionine, and tryptophan), and the uncharged polar family (glycine, asparagine, glutamine, cysteine, serine, threonine, and tyrosine). More preferred amino acid sets are as follows: the aliphatic hydroxyl set (serine and threonine), the amide-containing set (asparagine and glutamine), the aliphatic set (alanine, valine, leucine, and isoleucine), and the aromatic set (phenylalanine, tryptophan, and tyrosine). Such amino acid substitutions are preferably made within a range that does not impair the properties of the substance having the original amino acid sequence.

[0600] In addition, it is known that the lysine residue at the carboxyl terminus of the heavy chain of an antibody produced in cultured mammalian cells is deleted (Journal of Chromatography A, 705:129-134 (1995)), and it is also known that two amino acid residues (glycine and lysine) are deleted at the carboxyl terminus of the heavy chain of an antibody produced in cultured mammalian cells, and a new proline residue at the carboxyl terminus is amidated (Analytical Biochemistry, 360:75-83 (2007)). However, such deletions and modifications of the heavy chain sequence do not affect the antigen-binding affinity and effector functions (complement activation, antibody-dependent cellular cytotoxicity, etc.) of the antibody.

[0601] The 20 conventional amino acids referred to herein are written 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)), incorporated herein by reference. As used herein, the terms "polypeptide" and "protein" have the same meaning and are used interchangeably. Also, in this disclosure, amino acids are generally represented by one-letter and three-letter abbreviations well known in the art. For example, alanine can be represented by A or Ala, arginine can be represented by R or Arg, glycine can be represented by G or Gly, and glutamine can be represented by Q or Gln.

[0602] As used herein, the term "prophylaxis" refers to a method performed to prevent or delay the onset of a disease or disorder or condition (e.g., tumors and infections) in a subject. As used herein, the term "treatment" refers to a method performed to obtain beneficial or desired clinical results. For purposes of this disclosure, beneficial or desired clinical results include, but are not limited to, alleviation of symptoms, reduction in the extent of disease, stabilization of the disease state (i.e., not worsening), delay or slowing of disease progression, improvement or mitigation of the disease state, and alleviation of symptoms (whether partial or complete), whether detectable or undetectable. In addition, "treatment" can also mean prolonging survival as compared to expected survival (if not receiving treatment).

[0603] As used herein, the term "subject" refers to a mammal, e.g., a primate mammal, such as a non-human primate mammal or a human. In certain embodiments, the subject (e.g., a human) has or is at risk of having a tumor or infectious disease.

[0604] As used herein, the term "effective amount" refers to an amount sufficient to achieve or at least partially achieve a desired effect. For example, an effective amount for preventing a disease (e.g., tumors and infections) is an amount sufficient to prevent, stop, or delay the onset of the disease (e.g., tumors and infections). An effective amount for treating a disease is an amount sufficient to cure or at least partially stop the disease and its complications in a patient already suffering from the disease and its complications. Determining such effective amounts is entirely within the capabilities of one skilled in the art. For example, an amount effective for therapeutic use will depend on the severity of the disease being treated, the general condition of the patient's own immune system, the patient's general condition, such as age, weight, and sex, the method by which the drug is administered, and other treatments administered simultaneously.

[0605] As used herein, the term "effector functions" refers to the biological activities attributable to the Fc region of an antibody (either a native sequence Fc region or amino acid sequence variant Fc region), which vary depending on the antibody isotype.

[0606] The term "pharmaceutically acceptable" means that the molecule itself, molecular fragment, or composition does not produce an adverse, allergic, or other untoward reaction when appropriately administered to an animal or human. Specific examples of some substances that may be pharmaceutically acceptable carriers or components thereof include sugars (e.g., lactose), starch, cellulose and its derivatives, vegetable oils, gelatin, polyols (e.g., propylene glycol), alginic acid, etc.

[0607] The in vivo therapeutic effects of antibodies and antibody-drug conjugates on cancer can be determined using experimental animals, for example, by administering the antibody to nude mice implanted with a B7H3-expressing tumor cell line and measuring any changes in the cancer cells.

[0608] In the present disclosure, most of the amino acid substitutions in antibodies are, but are not limited to, L-amino acids. In some embodiments, one or more D-amino acids may be included in the antibody peptide chain. Peptides containing D-amino acids are more stable and less susceptible to degradation in the oral cavity, intestine, or plasma than peptides containing only L-amino acids.

[0609] Monoclonal antibodies used in the present disclosure can be produced by several methods. For example, the monoclonal antibodies used in the present disclosure can be obtained by the hybridoma method using several species (including cells from mouse, hamster, rat, and human) (see, e.g., Kohler et al., 1975, Nature, 256:495), or by recombinant DNA technology (see, e.g., U.S. Pat. No. 4,816,567), or can be isolated from phage antibody libraries (see, e.g., Clackson et al., 1991, Nature, 352:624-628, and Marks et al., 1991, Journal of Molecular Biology, 222:581-597).

[0610] In this specification, the expressions "independently selected from each ..." and "independently selected from each ..." used throughout this specification, unless otherwise clearly stated, can be interchangeable and should be understood broadly, meaning that specific options expressed for the same or different symbols in different groups do not affect each other, and may also mean that specific options expressed for the same or different symbols in the same group do not affect each other.

[0611] As used herein, the term "direct bond" refers to a bond of Formula II [ka] When X is a direct bond, the structural formula is: [ka] Other direct bonds can be understood with reference to the above.

[0612] As used herein, the term "absent" refers to a compound of Formula II [ka] When W is absent, the structural formula is [ka] is.

[0613] Formula II [ka] In the compound represented by the formula: R1 and R2 together with the carbon atom to which they are both attached represent: [ka] where the "dotted line" indicates that the heterocycle is fused to a benzene ring, e.g. [ka] It shows that the

[0614] As used herein, the compound of formula III [ka] In the drug-linker conjugate represented by [ka] In this case, the symbols 1 and 2 indicate the binding positions of L4 and other groups. Specifically, when the 1-position is bound to L3 and the 2-position is bound to a drug molecule, i.e., [ka] The remaining marks 1 and 2 can be understood by referring to the above.

[0615] As used herein, Formula II [ka] In a compound represented by the formula: R3 and X together with the carbon atom to which they are both attached [ka] wherein the "dotted line" indicates that the heterocycle is fused to the benzene ring, [ka] It shows that the

[0616] As used herein, X may be used in the form of, for example, "X is optionally substituted [ka] and the substituents are selected from 1 or 2 C1-4 alkyl groups (e.g., methyl), X can be, for example, [ka] Other similar definitions of X can be understood with reference to the foregoing.

[0617] As used herein, X may be used in the form of, for example, "X is optionally substituted [ka] and the substituents are selected from two C alkyl groups (e.g., methyl) which, together with the carbon atom to which they are both attached, form a C cycloalkyl (e.g., cyclopropyl), then X can be, for example, [ka] Other similar definitions of X can be understood with reference to the foregoing.

[0618] AA 1 In the structure of amino acid residues represented by [ka] If r is 0, then AA 1 It will be understood by those skilled in the art that the structure of an amino acid residue represented by [ka]

[0619] AA 1 In the structure of amino acid residues represented by [ka] R a and R b are taken together with the carbon atoms to which they are both attached to form a 4- to 10-membered heterocyclic ring, and the 4- to 10-membered heterocyclic ring is bound to one or more R 0 When the 4- to 10-membered heterocycle is optionally substituted with one or more R 0 The term "optionally substituted with" refers to a 4- to 10-membered heterocycle that is unsubstituted but contains one or more R 0 and a plurality of R 0 In each R 0 It means that the definitions of may be the same or different. Other similar definitions can be understood by referring to the above content.

[0620] As used herein, for example, when L3 is selected from Lys, Val-Cit, Ala-Ala-Asn, Ala-Ala-Asp, Gly-Gly-Phe-Gly, Val-Lys-Gly, Val-Ala, and Lys-Ala-Asn, "the distal amino group of lysine (Lys) is optionally substituted with one, two, or three substituents selected from tert-butoxycarbonyl, C1-6 alkyl (preferably methyl), and O" means that the distal amino group of Lys in each of the L3 options is optionally substituted with one, two, or three substituents selected from tert-butoxycarbonyl, C1-6 alkyl (preferably methyl), and O. The "distal amino group of Lys" refers to a lysine residue. [ka] The phrase "the distal amino group of lysine (Lys) is optionally substituted with one, two or three substituents selected from tert-butoxycarbonyl, C1-6 alkyl (preferably methyl), and O" refers to the naked amino group -NH2 in the formula (I). The phrase "the distal amino group of lysine (Lys) is optionally substituted with one, two or three substituents selected from tert-butoxycarbonyl, C1-6 alkyl (preferably methyl), and O" means that the distal amino group of lysine (Lys) may be unsubstituted, or may be substituted with one, two or three substituents selected from tert-butoxycarbonyl, C1-6 alkyl (preferably methyl), and O. For example, it may be substituted with one tert-butoxycarbonyl group, i.e., it may be [ka] or may be substituted with two methyls, i.e., it is [ka] or may be substituted by two methyls and one O, i.e. it is [ka] "The distal amino group of lysine (Lys) is substituted with O" means that the distal amino group of lysine (Lys) is substituted with oxo, i.e., [ka] and if the distal amino group is further substituted with two methyl groups, the distal amino group becomes [ka] This means that

[0621] At various points in this specification, substituents of compounds in this disclosure are disclosed according to group types or ranges. Specifically, the present invention includes each individual subcombination of each element of these group types and ranges. For example, the term "C1-6 alkyl" specifically refers to methyl, ethyl, C3 alkyl, C4 alkyl, C5 alkyl, and C6 alkyl, each of which is independently disclosed.

[0622] As used herein, the term "C1-6 alkyl" refers to a straight or branched chain alkyl group containing 1 to 6 carbon atoms, such as "C1-3 alkyl" or "C1-4 alkyl", methyl, ethyl, etc., and specific examples include, but are not limited to, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, pentyl, and hexyl.

[0623] As used herein, "C 1~4 The term "alkyl" refers to a straight or branched chain alkyl group containing from 1 to 4 carbon atoms, including, for example, "C1-3 alkyl," methyl, ethyl, etc. Specific examples include, but are not limited to, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl.

[0624] As used herein, "C 2~6 The term "alkenyl" refers to a straight, branched, or cyclic alkenyl group containing at least one double bond and 2 to 6 carbon atoms, e.g., "C 2~4Examples include, but are not limited to, vinyl, 1-propenyl, 2-propenyl, 1-butenyl, 2-butenyl, 1,3-butadienyl, 1-pentenyl, 2-pentenyl, 3-pentenyl, 1,3-pentadienyl, 1,4-pentadienyl, 1-hexenyl, 2-hexenyl, 3-hexenyl, 1,4-hexadienyl, cyclopentenyl, 1,3-cyclopentadienyl, cyclohexenyl, 1,4-cyclohexadienyl, and the like.

[0625] As used herein, "C 2~6 The term "alkynyl" refers to a straight or branched chain alkynyl group containing at least one triple bond and 2 to 6 carbon atoms, e.g., "C 2~4 Examples include, but are not limited to, ethynyl, propynyl, 2-butynyl, 2-pentynyl, 3-pentynyl, 4-methyl-2-pentynyl, 2-hexynyl, 3-hexynyl, 5-methyl-2-hexynyl, and the like.

[0626] As used herein, the term "halogen" includes fluorine, chlorine, bromine, and iodine.

[0627] As used herein, "3- to 6-membered cycloalkyl" or "C 3~6 The term "cycloalkyl" refers to saturated cyclic alkyl containing from 3 to 6 carbon atoms, including cyclopropanyl (ie, cyclopropyl), cyclobutanyl (ie, cyclobutyl), cyclopentanyl (ie, cyclopentyl), and cyclohexyl.

[0628] As used herein, "3- to 7-membered carbocycloalkyl" or "C 3~7 The term "cycloalkyl" refers to saturated cyclic alkyls containing 3 to 7 carbon atoms, including cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, and cycloheptyl.

[0629] As used herein, "C 1~6The term "alkoxy" refers to an alkyl group, as defined above, attached to the parent molecular moiety through an oxygen atom. Illustrative examples include, but are not limited to, methoxy, ethoxy, propoxy, isopropoxy, n-propoxy, isopropoxy, n-butoxy, isobutoxy, tert-butoxy, pentoxy, hexyloxy, and the like.

[0630] As used herein, "C 1~4 The term "alkoxy" refers to an alkyl group, as defined above, attached to the parent molecular moiety through an oxygen atom. Illustrative examples include, but are not limited to, methoxy, ethoxy, propoxy, isopropoxy, n-propoxy, isopropoxy, n-butoxy, isobutoxy, tert-butoxy, and the like.

[0631] As used herein, the term "4- to 10-membered heterocyclic group" refers to a cyclic group containing 4 to 10 ring atoms, at least one of which is a heteroatom such as a nitrogen, oxygen, or sulfur atom. The term "4- to 6-membered heterocyclic group" refers to a cyclic group containing 4 to 6 ring atoms (at least one of which is a heteroatom such as a nitrogen, oxygen, or sulfur atom). Optionally, a ring atom (e.g., a carbon, nitrogen, or sulfur atom) in the ring structure may be replaced by oxygen. Examples of the "4- to 8-membered heterocyclic group" include "4- to 8-membered nitrogen-containing heterocyclic group", "4- to 8-membered oxygen-containing heterocyclic group", "4- to 7-membered heterocyclic group", "4- to 7-membered oxygen-containing heterocyclic group", "4- to 7-membered heterocyclic group", "4- to 6-membered heterocyclic group", "5- to 7-membered heterocyclic group", "5- to 6-membered heterocyclic group", and "5- to 6-membered nitrogen-containing heterocyclic group", including, but not limited to, oxocyclobutanyl, pyrrolidinyl, tetrahydrofuryl, piperdinyl, piperazinyl, tetrahydropyranyl, homopiperazinyl, and the like.

[0632] As used herein, the term "4- to 10-membered heterocycle" refers to a ring containing 4 to 10 ring atoms (at least one ring atom is a heteroatom such as a nitrogen, oxygen, or sulfur atom). The term "5- to 6-membered heterocycle" refers to a ring containing 5 to 6 ring atoms (at least one ring atom is a heteroatom such as a nitrogen, oxygen, or sulfur atom), including, but not limited to, pyrrolidine, tetrahydrofuran, piperidine, piperazine, tetrahydropyran, and the like.

[0633] As used herein, the term "aryl" refers to a monocyclic or polycyclic hydrocarbon group having aromaticity, such as a 6- to 10-membered aryl or a 5- to 8-membered aryl. Specific examples include, but are not limited to, phenyl, naphthyl, anthracenyl, phenanthryl, and the like. A "6- to 10-membered aryl" refers to an aryl having 6 to 10 ring atoms. A "C6-10 aryl" refers to an aryl group containing 6 to 10 carbon atoms.

[0634] As used herein, the term "heteroaryl" refers to a cyclic group having aromaticity, in which at least one ring atom is a heteroatom such as a nitrogen, oxygen, or sulfur atom. Optionally, a ring atom (e.g., a carbon, nitrogen, or sulfur atom) in the ring structure may be replaced with oxygen. Specific examples include 5- to 10-membered heteroaryls, 5- to 6-membered heteroaryls, 5- to 10-membered nitrogen-containing heteroaryls, 6- to 10-membered oxygen-containing heteroaryls, 6- to 8-membered nitrogen-containing heteroaryls, and 5- to 8-membered oxygen-containing heteroaryls, such as furyl, thienyl, pyrrolyl, thiazolyl, isothiazolyl, thiadiazolyl, oxazolyl, isoxazolyl, oxadiazolyl, imidazolyl, pyrazolyl, 1,2,3-triazolyl, 1,2,4-triazolyl, 1,2,3-oxadiazolyl, 1,2,4-oxadiazolyl, 1,2,5-oxadiazolyl, and 1,3,4- These include, but are not limited to, oxadiazolyl, pyridyl, 2-pyridonyl, 4-pyridonyl, pyrimidinyl, 1,4-dioxinyl, 2H-1,2-oxazinyl, 4H-1,2-oxazinyl, 6H-1,2-oxazinyl, 4H-1,3-oxazinyl, 6H-1,3-oxazinyl, 4H-1,4-oxazinyl, pyridazinyl, pyrazinyl, 1,2,3-triazinyl, 1,3,5-triazinyl, 1,2,4,5-tetrazinyl, azacycloheptatrienyl, 1,3-diazacycloheptatrienyl, azacyclooctatetraenyl, and the like.

[0635] A bond in a structural formula represented herein by a wavy line "~" is intended to indicate that the structure represents either a cis or trans isomer, or a mixture of cis and trans isomers in any ratio.

[0636] The term "drug-antibody ratio" or "DAR" refers to the amount of drug, e.g., a small molecule toxin, conjugated to the antibody of an ADC. The DAR of an ADC can range from 1 to 16, although it depends on the number of binding sites in the antibody, and thus higher loadings (e.g., 20) are possible. The term DAR can be used to refer to the amount of drug loaded onto a single antibody, or to refer to the average or mean DAR of a group of ADCs.

[0637] Beneficial effects of the present invention: Through extensive research, the present invention has provided highly potent and highly cell-permeable bioactive drugs (bioactive molecules or payloads): linkers with high systemic circulation stability and high chemical stability in vivo; linker-driven ADC enrichment in the tumor microenvironment by adjusting the linker's physicochemical properties, such as basicity and lipophilicity; unique linker cleavage properties in vivo; and coupling modes between the targeting moiety and the linker. The above discoveries, combined with numerous in vitro and in vivo screenings and tests, have provided a novel class of antibody-drug conjugates that offer the following several surprising technical advantages: The conjugate products obtained according to the above method have better solubility and excellent chemical stability, and avoid the retro-Michael reaction of conventional maleimide-based ADCs. Therefore, the ADC products of the present disclosure can have a high drug-to-antibody ratio (DAR). In some embodiments, the DAR value of the conjugates can reach 6-8. High coupling efficiency: In some embodiments, coupling efficiency can reach or exceed 90%. Through numerous experimental studies, a class of highly plasma-stable linkers has been discovered that can be cleaved both intracellularly and extracellularly in the tumor microenvironment. The ADC products of the present disclosure can achieve good antitumor effects in tumors with low or no antigen expression. By adjusting the physical and chemical properties of the linker and the overall ADC molecule, the ADC product obtained according to the above method increases its exposure in the relatively acidic tumor environment, and therefore the ADC product of the present disclosure has better tumor tissue targeting. The ability of the ADC to concentrate in the tumor microenvironment increases the payload concentration ratio between the tumor and the blood. This reduces ADC mechanism-related toxicity (i.e., toxicity of the ADC after binding to cell surface antigens and endocytosis in non-tumor tissues, or referred to as "on-target toxicity"), thereby achieving a higher therapeutic index. The conjugates obtained according to the above methods have high stability in the circulation in vivo, low cleavage of the drug molecule in non-target tissues, and therefore attenuate the "off-target" toxicity caused by cleavage of the toxin in non-target tissues. The biologically active molecules of the conjugates of the present invention have higher anti-tumor cell activity and therefore have excellent bystander effects, and the ADCs can more effectively kill tumor cells with high antigen expression as well as tumor cells with low or no antigen expression in tumor tissues. By utilizing the extracellular cleavage ability of the linker of the present disclosure in the tumor microenvironment, the toxin-linker of the present disclosure can form an antibody-drug conjugate with an antibody that does not have endocytic ability, and such an antibody-drug conjugate has high in vivo anti-tumor activity. By utilizing both the extracellular cleavage ability and tumor microenvironment concentration ability, the toxin-linker of the present disclosure can form an antibody-drug conjugate with an antibody that does not have endocytosis ability or is incapable of cell surface antigen binding, and such an antibody-drug conjugate has high in vivo anti-tumor activity, The antibodies provided in the present disclosure are highly humanized or fully human antibodies so that they can be safely administered to human subjects without eliciting an immunogenic response. These molecules have high binding affinity to human and non-human B7H3, particularly antibody molecules that are cross-reactive with B7H3 of non-human primates (e.g., cynomolgus monkeys). In conclusion, the linkers, antibodies and ADCs of the present disclosure have great clinical value. [Brief explanation of the drawings]

[0638] Text description of the illustration image022.gif. [Figure 1A] Binding detection of anti-B7H3 antibody 1D1-01 to human B7H3-4 IgG-His protein by ELISA.

[0639] [Figure 1B]Binding detection of anti-B7H3 antibody 2E3-02 to human B7H3-4 IgG-His protein by ELISA.

[0640] [Figure 2A] 10: Detection of binding of anti-B7H3 antibody 1D1-01 to monkey B7H3-4 IgG-His protein by ELISA.

[0641] [Figure 2B] Binding of anti-B7H3 antibody 2E3-02 to monkey B7H3-4 IgG-His protein detected by ELISA.

[0642] [Figure 3A] Binding detection of anti-B7H3 antibody 1D1-01 to MCF-7 tumor cells by flow cytometry.

[0643] [Figure 3B] Binding detection of anti-B7H3 antibody 2E3-02 to MCF-7 tumor cells by flow cytometry.

[0644] [Figure 4A] Binding detection of anti-B7H3 antibody 1D1-01 to A549 tumor cells by flow cytometry.

[0645] [Figure 4B] Binding detection of anti-B7H3 antibody 2E3-02 to A549 tumor cells by flow cytometry.

[0646] [Figure 5] Binding of anti-B7H3 antibody 2E3-02 to PC-3 tumor cells detected by flow cytometry.

[0647] [Figure 6A] This is the detection of the specific binding of anti-B7H3 antibody 1D1-01 to B7H3 protein.

[0648] [Figure 6B] This is the detection of the specific binding of anti-B7H3 antibody 2E3-02 to B7H3 protein.

[0649] [Figure 7A] This is the detection of endocytosis of tumor cells A549 toward the candidate antibody 1D1-01.

[0650] [Figure 7B] This is the detection of endocytosis of tumor cells A549 toward the candidate antibody 2E3-02.

[0651] [Figure 7C] This is the detection of endocytosis of tumor cells A549 toward candidate antibody 202-2-1.

[0652] [Figure 8] SEC profiles before and after conjugation.

[0653] [Figure 9] Assay I for anti-B7H3-ADCs inhibiting tumor growth in the NCI-HT29 xenograft model.

[0654] [Figure 10] Assay II for anti-B7H3-ADCs inhibiting tumor growth in the NCI-HT29 xenograft model.

[0655] [Figure 11] Assay for anti-B7H3-ADCs inhibiting tumor growth in the NCI-H358 xenograft model.

[0656] [Figure 12] Assay for anti-B7H3-ADC inhibiting tumor growth in an esophageal squamous cell carcinoma PDX model.

[0657] [Figure 13]Assay for anti-B7H3-ADC inhibiting tumor growth in a prostate PDX model.

[0658] [Figure 14] Assay for anti-Her3-ADCs inhibiting tumor growth in the NCI-H358 xenograft model.

[0659] [Figure 15] Assay I for anti-Her3-ADC inhibiting tumor growth in the SW480 xenograft model.

[0660] [Figure 16] Assay II for anti-Her3-ADCs inhibiting tumor growth in the SW480 xenograft model.

[0661] [Figure 17] Distribution ratio of A1.9 in plasma and tumor in the HT29 model.

[0662] [Figure 18] Distribution ratio of ADC in plasma and tumor in the HT29 model.

[0663] [Figure 19A] 1 shows the detection results of incubation of antibody drug conjugates in tumor homogenates.

[0664] [Figure 19B] 1 shows the incubation results of antibody drug conjugates in muscle tissue homogenate. [Example]

[0665] The present disclosure will be further described below by describing specific examples, but these are not intended to limit the present disclosure. Based on the teachings of the present disclosure, those skilled in the art can make various modifications or improvements without departing from the basic spirit and scope of the present disclosure. The reagents or equipment used without manufacturer's instructions are conventional products that can be commercially available.

[0666] The abbreviations in this disclosure have the following meanings: [Table 8] Sequence and its specific information: [Table 9-1] [Table 9-2] [Table 9-3] [Table 9-4] The present disclosure will now be described with reference to the following examples which are intended to illustrate, but not limit, the present disclosure.

[0667] Unless otherwise specified, the molecular biology experimental methods and immunodetection methods used in the present disclosure are essentially those described in J. Sambrook et al., Molecular Cloning: Laboratory Handbook, 2nd Edition, Cold Spring Harbor Laboratory Press, 1989, and F.M.Ausubel et al., Short Protocols in Molecular Biology, 3rd Edition, John Wiley & Sons, Inc., 1995. Those skilled in the art will understand that these examples are provided to illustrate the present disclosure by way of illustration and are not intended to limit the scope of protection claimed in the present disclosure.

[0668] Preparation procedure The structures of the compounds described in the following examples were determined by nuclear magnetic resonance ( 1 The hydroxyl groups were determined by 1 H NMR or mass spectrometry (MS).

[0669] Nuclear magnetic resonance ( 1 The instrument for detecting H NMR was a Bruker 400 MHz nuclear magnetic resonance spectrometer, the test solvent was deuterated methanol (CD3OD), deuterated chloroform (CDCl3) or hexadeuterated dimethyl sulfoxide (DMSO-d6), and the internal standard was tetramethylsilane (TMS).

[0670] The abbreviations for nuclear magnetic resonance (NMR) spectra used in the examples are shown below.

[0671] s: singlet, d: doublet, t: triplet, q: quartet, dd: double doublet, qd: quartet doublet, ddd: double double doublet, ddt: double double triplet, dddd: double double double doublet, m: multiplet, br: broad, J: coupling constant, Hz: hertz, DMSO-d6: deuterated dimethyl sulfoxide. δ values ​​are expressed in ppm.

[0672] An Agilent (ESI) mass spectrometer (model Agilent 6120B) was used as the instrument for mass spectrometry (MS) testing.

[0673] I. Synthesis of biologically active molecules and intermediates used in the synthetic procedures for "drug-linker compounds." A. Synthesis of Bioactive Molecules Example A1.1: Synthesis of (S)-4-ethyl-8-fluoro-4-hydroxy-9-methyl-11-(1H-pyrazol-4-yl)-1,12-dihydro-14H-pyrano[3',4':6,7]indolizino[1,2-b]quinoline-3,14(4H)-dione (A1.1) [ka] Step 1: 4-Bromo-1-(tetrahydro-2H-pyran-2-yl)-1H-pyrazole (4.6 g) was dissolved in anhydrous tetrahydrofuran (50 ml), and the solution was cooled to -78 °C in dry ice-acetone under nitrogen protection. Then, n-butyllithium (12 ml, 2 M) was added dropwise. The reaction solution was stirred at -78 °C for 20 minutes, and then methyl 2-amino-4-fluoro-5-methylbenzoate (1.83 g) was added. After the addition, the reaction mixture was allowed to warm to room temperature and stirred continuously for 5 hours. The reaction mixture was quenched by the addition of methanol (3 ml), followed by the addition of ethyl acetate (200 ml). The solution was washed with water (100 ml × 3), and the organic phase was dried. The organic solvent was removed, and the residue was separated by silica gel column chromatography to obtain the target product (2-amino-4-fluoro-5-methylphenyl)(1-(tetrahydro-2H-pyran-2-yl)-1H-pyrazol-4-yl)methanone. ESI-MS (m / z): 304 [M+H] + .

[0674] Step 2: (S)-4-Ethyl-4-hydroxy-7,8-dihydro-1H-pyrano[3,4-f]indolizine-3,6,10(4H)-trione (2.63 g), (2-amino-4-fluoro-5-methylphenyl)(1-(tetrahydro-2H-pyran-2-yl)-1H-pyrazol-4-yl)methanone (3.03 g), and p-toluenesulfonic acid (1.74 g) were dissolved in dichloromethane (50 ml), and the solvent was removed. The mixture was heated to 120°C under nitrogen protection and reacted for 4 hours. The mixture was dissolved in ethyl acetate (300 ml), and the organic phase was washed with water (100 ml x 2), dried, and the organic solvent was removed. The residue was separated by silica gel column chromatography to obtain the target product (S)-4-ethyl-8-fluoro-4-hydroxy-9-methyl-11-(1H-pyrazol-4-yl)-1,12-dihydro-14H-pyrano[3',4':6,7]indolizino[1,2-b]quinoline-3,14(4H)-dione (A1.1). ESI-MS (m / z): 447 [M+H] + .

[0675] Example A1.2: Synthesis of (S)-7-ethyl-7-hydroxy-14-(1H-pyrazol-4-yl)-10,13-dihydro-11H-[1,3]dioxolo[4,5-g]pyrano[3',4':6,7]indolizino[1,2-b]quinoline-8,11(7H)-dione (A1.2) [ka] Using the same method and reaction conditions as in Example A1.1, methyl 2-amino-4-fluoro-5-methylbenzoate was replaced with methyl 6-aminobenzo[d][1,3]dioxolane-5-carboxylate to give the target product (S)-7-ethyl-7-hydroxy-14-(1H-pyrazol-4-yl)-10,13-dihydro-11H-[1,3]dioxolo[4,5-g]pyrano[3',4':6,7]indolizino[1,2-b]quinoline-8,11(7H)-dione (A1.2). ESI-MS (m / z): 459 [M+H] + .

[0676] Example A1.3: Synthesis of (S)-14-(3-aminophenyl)-7-ethyl-7-hydroxy-10,13-dihydro-11H-[1,3]dioxolo[4,5-g]pyrano[3',4':6,7]indolizino[1,2-b]quinoline-8,11(7H)-dione (A1.3) [ka] Compound (A1.3-A) (1.4 g), compound (A1.3-B) (2.2 g), Pd2(DBA)3 (300 mg), tricyclohexylphosphane (300 mg), and potassium acetate (1.1 g) were added sequentially to a mixture of dioxane (30 ml) and water (5 ml), and the mixture was heated to 100 °C and reacted for 12 hours with stirring under nitrogen protection. After cooling, ethyl acetate (200 ml) was added, and the resulting solution was washed with water (100 ml). After drying, the target product (S)-14-(3-aminophenyl)-7-ethyl-7-hydroxy-10,13-dihydro-11H-[1,3]dioxolo[4,5-g]pyrano[3',4':6,7]indolizino[1,2-b]quinoline-8,11(7H)-dione (A1.3) was isolated by silica gel column chromatography. ESI-MS (m / z): 484 [M+H] + .

[0677] Example A1.4: Synthesis of (S)-14-(4-aminophenyl)-7-ethyl-7-hydroxy-10,13-dihydro-11H-[1,3]dioxolo[4,5-g]pyrano[3',4':6,7]indolizino[1,2-b]quinoline-8,11(7H)-dione (A1.4) [ka] Compound (A1.4-A) (1.4 g), compound (A1.4-B) (2.2 g), Pd2(DBA)3 (300 mg), tricyclohexylphosphane (300 mg), and potassium acetate (1.1 g) were added sequentially to a mixture of dioxane (30 ml) and water (5 ml), and the mixture was heated to 100 °C and reacted for 12 hours with stirring under nitrogen protection. After cooling, ethyl acetate (200 ml) was added, and the resulting solution was washed with water (100 ml). After drying, the target product (S)-14-(4-aminophenyl)-7-ethyl-7-hydroxy-10,13-dihydro-11H-[1,3]dioxolo[4,5-g]pyrano[3',4':6,7]indolizino[1,2-b]quinoline-8,11(7H)-dione (A1.4) was obtained by separation using silica gel column chromatography.

[0678] ESI-MS (m / z): 484 [M+H]. + , 1 H NMR(400MHz,DMSO)δ7.56(s,1H),7.34(d,J=8.0Hz,2H),7.27(s,1H),7.14(s,1H),6.89(d,J= 7.8Hz,2H),6.26(s,2H),5.40(s,2H),5.05(s,2H),1.96-1.78(m,2H),0.88(t,J=7.2Hz,3H).

[0679] Example A1.5: Synthesis of (S)-14-(3-aminopropyl)-7-ethyl-7-hydroxy-10,13-dihydro-11H-[1,3]dioxolo[4,5-g]pyrano[3',4':6,7]indolizino[1,2-b]quinoline-8,11(7H)-dione (A1.5) [ka] Step 1: Synthesis of (S)-14-(3-chloropropyl)-7-ethyl-7-hydroxy-10,13-dihydro-11H-[1,3]dioxolo[4,5-g]pyrano[3',4':6,7]indolizino[1,2-b]quinoline-8,11(7H)-dione [ka] In an ice bath, ferrous sulfate heptahydrate (570 mg of ferrous sulfate heptahydrate was dissolved in 1 mL of water) and 4-dimethoxychlorobutane (3.89 g) were added to a 75% sulfuric acid solution (5 mL) of compound (S) 7-ethyl-7-hydroxy-10,13-dihydro-11H-[1,3]dioxolo[4,5-g]pyrano[3',4':6,7]indeno[1,2-b]quinoline-8,11(7H)-dione (A1.5-A, 500 mg). The reaction solution was stirred for 3 minutes, and then hydrogen peroxide (29%, 2.5 mL) was added dropwise. The reaction solution was stirred at 0°C for 5 minutes, warmed to room temperature, and reacted for 3 hours with stirring. The reaction solution was diluted with water (50 mL) and extracted with ethyl acetate (80 mL x 2). The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated to give the crude product, which was further purified by C18 column (acetonitrile / 0.05% formic acid aqueous solution: 5% to 60%) to give the target compound (yellow solid, 400 mg, yield: 67%).

[0680] LCMS(ESI)[M+H] + :468.9, 1 H NMR(400MHz,DMSO-d6)δ7.65(s,1H),7.51(s,1H),7.24(s,1H),6.50(s,1H),6.30(s,2H),5.42(s, 2H),5.26(s,2H),3.81(d,J=5.9Hz,2H),3.22(s,2H),1.98(d,J=6.7Hz,4H),0.88(t,J=7.2Hz,3H).

[0681] Step 2: Synthesis of (S)-14-(3-azidopropyl)-7-ethyl-7-hydroxy-10,13-dihydro-11H-[1,3]dioxolo[4,5-g]pyrano[3',4':6,7]indolizino[1,2-b]quinoline-8,11(7H)-dione [ka] Sodium azide (284 mg) was added to a solution of the compound (S)-14-(3-chloropropyl)-7-ethyl-7-hydroxy-10,13-dihydro-11H-[1,3]dioxolo[4,5-g]pyrano[3',4':6,7]indolizino[1,2-b]quinoline-8,11(7H)-dione (200 mg) in N,N-dimethylformamide (3 ml), and the reaction solution was stirred at 100° C. for 1 hour. Water (30 ml) was added to the reaction solution, and the resulting solution was extracted with ethyl acetate (60 ml × 2). The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated to give the target compound (S)-14-(3-azidopropyl)-7-ethyl-7-hydroxy-10,13-dihydro-11H-[1,3]dioxolo[4,5-g]pyrano[3',4':6,7]indolizino[1,2-b]quinoline-8,11(7H)-dione (yellow solid, 180 mg, yield: 88%).

[0682] LCMS(ESI)[M+H] + :476, 1 H NMR(400MHz,DMSO-d6)δ7.65(s,1H),7.51(s,1H),7.24(s,1H),6.48(s,1H),6.29(s,2H),5.42(s, 2H),5.25(s,2H),3.53-3.49(m,2H),3.16-3.12(m,2H),1.93-1.80(m,4H),0.88(t,J=7.2Hz,3H).

[0683] Step 3: Synthesis of (S)-14-(3-aminopropyl)-7-ethyl-7-hydroxy-10,13-dihydro-11H-[1,3]dioxolo[4,5-g]pyrano[3',4':6,7]indolizino[1,2-b]quinoline-8,11(7H)-dione [ka] To a solution of the compound (S)-14-(3-azidopropyl)-7-ethyl-7-hydroxy-10,13-dihydro-11H-[1,3]dioxolo[4,5-g]pyrano[3',4':6,7]indolizino[1,2-b]quinoline-8,11(7H)-dione (130 mg, 0.274 mmol) in a mixture of tetrahydrofuran (3 mL) and water (1 mL), triphenylphosphine (108 mg, 0.411 mmol) was added, and the mixture was reacted at 55°C for 16 hours. LCMS indicated the reaction was complete. Water (5 mL) was added to the reaction solution, and the resulting solution was acidified with 2N hydrochloric acid (3 mL) and extracted with ethyl acetate (10 mL). The aqueous phase was purified by high-performance liquid chromatography (HPLC) (acetonitrile / 0.05% formic acid in water) to give the target compound (S)-14-(3-aminopropyl)-7-ethyl-7-hydroxy-10,13-dihydro-11H-[1,3]dioxolo[4,5-g]pyrano[3',4':6,7]indolizino[1,2-b]quinoline-8,11(7H)-dione (A1.5, yellow solid, 15 mg, yield: 12%).

[0684] LCMS(ESI)[M+H]+:449.9, 1 H NMR(400MHz,DMSO-d6)δ7.72(s,3H),7.53(s,1H),7.25(s,1H),6.50(s,1H),6.30(s,2H),5.43(s,2H) ,5.24(s,2H),3.15(d,J=6.4Hz,2H),3.03(t,J=6.9Hz,2H),1.96-1.81(m,4H),0.88(t,J=7.3Hz,3H).

[0685] Example A1.6: Synthesis of (S)-N-ethyl-N-(2-(7-ethyl-7-hydroxy-8,11-dioxo-7,8,11,13-tetrahydro-10H-[1,3]dioxolo[4,5-g]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-14-yl)ethyl)-2-hydroxyacetamide (A1.6) [ka] Step 1: Preparation of (S)-2-(ethyl(2-(7-ethyl-7-hydroxy-8,11-dioxo-7,8,11,13-tetrahydro-10H-[1,3]dioxolo[4,5-g]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-14-yl)ethyl)amino)-2-oxoethyl acetate (A1.6-B): [ka] The compound (S)-7-ethyl-14-(2-(ethylamine)ethyl)-7-hydroxy-10,13-dihydro-11H-[1,3]dioxolo[4,5-g]pyrano[3',4':6,7]indolizino[1,2-b]quinoline-8,11(7H)-dione (A1.6-A, 50 mg), acetyloxyacetyl chloride (73 mg), and triethylamine (50 mg) were added sequentially to dichloromethane (5 ml), and the mixture was stirred at room temperature for 1 hour. The reaction solution was rotary evaporated to remove the solvent, yielding 75 mg of crude oil.

[0686] LCMS(ESI)[M+H] + :564.2.

[0687] Step 2: Synthesis of (S)-N-ethyl-N-(2-(7-ethyl-7-hydroxy-8,11-dioxo-7,8,11,13-tetrahydro-10H-[1,3]dioxolo[4,5-g]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-14-yl)ethyl)-2-hydroxyacetamide (A1.6) [ka] The crude compound (S)-2-(ethyl(2-(7-ethyl-7-hydroxy-8,11-dioxo-7,8,11,13-tetrahydro-10H-[1,3]dioxolo[4,5-g]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-14-yl)ethyl)amino)-2-oxoethyl acetate (75 mg, 0.13 mmol) was dissolved in a mixture of concentrated hydrochloric acid and absolute ethanol (volume ratio: 1 / 2, 3 ml), and the reaction solution was refluxed at 85°C for 1 hour. LCMS showed that the reaction was complete. The reaction solution was concentrated, and the crude product was purified by preparative chromatography (0.01% aqueous trifluoroacetic acid, acetonitrile) to give the target compound (S)-N-ethyl-N-(2-(7-ethyl-7-hydroxy-8,11-dioxo-7,8,11,13-tetrahydro-10H-[1,3]dioxolo[4,5-g]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-14-yl)ethyl)-2-hydroxyacetamide (A1.6) (15 mg, 26% yield) as a white solid.

[0688] LCMS(ESI)[M+H] + :522.0, 1 H NMR(400MHz,DMSO)δ7.92(s,1H),7.53(m,1H),7.25(s,1H),6.50(s,1H),6.31(s,2H),5.43(s,2H),5.34(m,2H),4. 65(m,1H),4.07(m,2H),3.52(m,2H),3.41(m,2H),2.00(m,2H),1.88(m,2H),1.16-1.04(m,3H),0.89-0.83(m,3H).

[0689] Example A1.7: Synthesis of (S)-N-methyl-N-(2-(7-ethyl-7-hydroxy-8,11-dioxo-7,8,11,13-tetrahydro-10H-[1,3]dioxolo[4,5-g]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-14-yl)ethyl)-2-hydroxyacetamide (A1.7) [ka] Step 1: Preparation of compound benzylmethyl (3-(6-nitrobenzo[d][1,3]dioxol-5-yl)-3-oxopropyl)carbamate [ka] The compound 1-(6-nitrobenzo[d][1,3]dioxin-5-yl)ethan-1-one (A1.7-A, 500 mg), methylamine hydrochloride (1.6 g), and paraformaldehyde (714 mg) were dissolved in ethanol (8 ml), and the mixture was reacted in a sealed vessel at 100°C for 16 hours. The reaction solution was cooled to room temperature and concentrated under reduced pressure. The resulting residue was dissolved in dichloromethane (80 ml), and the organic phase was extracted with water (50 ml x 3). The pH of the resulting aqueous phase was adjusted to 9 with sodium bicarbonate, and then benzyl chloroformate (513 mg, 3.0 mmol) was added. The reaction was carried out at room temperature for 16 hours. The reaction solution was extracted with ethyl acetate (30 ml × 3), and the organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated to obtain a crude product. This was separated and purified by C18 column chromatography (acetonitrile / water containing 0.05% formic acid = 5-95%) to obtain the target compound (A1.7-B, 180 mg, yield 23%).

[0690] LCMS(ESI)[M+H] + :387.1.

[0691] 1 H NMR (400MHz, CDCl3) δ7.56(s,1H),7.35(m,5H),7.32(m,1H),6.17(s,2H),5.13(s,2H),3.71(m,2H),3.03(m,3H),2.99-2.86(m,2H).

[0692] Step 2: Preparation of compound benzyl (3-(6-aminobenzo[d][1,3]dioxin-5-yl)-3-oxopropyl)(methyl)carbamate (A1.7-C). [ka] Compound A1.7-B methylbenzyl (3-(6-nitrobenzo[d][1,3]dioxol-5-yl)-3-oxopropyl)carbamate (180 mg, 0.47 mmol) was dissolved in a mixture of saturated aqueous ammonium chloride (8 ml) and ethanol (8 ml), and then iron powder (130 mg) was added to the reaction solution. The reaction mixture was stirred at 80°C for 2 hours. The reaction solution was filtered, and the filtrate was concentrated under reduced pressure to a solid. The crude product was purified by TLC (petroleum ether:ethyl acetate = 2 / 1) to give target compound A1.7-C (76 mg).

[0693] LCMS(ESI)[M+H] + :357.0.

[0694] Step 3: Preparation of benzyl (S)-(2-(7-ethyl-7-hydroxy-8,11-dioxo-7,8,11,13-tetrahydro-10H-[1,3]dioxolo[4,5-g]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-14-yl)ethyl)(methyl)carbamate (A1.7-D). [ka] Compound A1.7-C benzyl (3-(6-aminobenzo[d][1,3]dioxin-5-yl)-3-oxopropyl)(methyl)carbamate (38 mg), compound (S)-4-ethyl-4-hydroxy-7,8-dihydro-1H-pyrano[3,4-f]indolizine-3,6,10(4H)-trione (29 mg), and p-toluenesulfonic acid (23 mg) were dissolved in dichloromethane (5 ml) at room temperature. After the solution became clear and homogeneous, it was concentrated under reduced pressure and evacuated with an oil pump. The reaction was carried out under vacuum at 120 °C for 2 hours. After the reaction mixture was cooled to room temperature, water (30 ml) was added, and the resulting mixture was extracted with dichloromethane (30 ml × 3). The combined organic phase was successively dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to give target compound A1.7-D (50 mg).

[0695] LCMS(ESI)[M+H] + =584.0.

[0696] Step 4: Preparation of (S) 7-ethyl-7-hydroxy-14-(2-(methylamino)ethyl)-10,13-dihydro-11H-[1,3]dioxolo[4,5-g]pyrano[3',4':6,7]indolizino[1,2-b]quinoline-8,11(7H)-dione (A1.7-E) [ka] Compound A1.7-D benzyl (S)-(2-(7-ethyl-7-hydroxy-8,11-dioxo-7,8,11,13-tetrahydro-10H-[1,3]dioxolo[4,5-g]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-14-yl)ethyl)(methyl)carbamate (50 mg) was dissolved in dichloromethane (5 ml) at room temperature, and trimethyliodosilane (51 mg, 0.26 mmol) was added thereto at 0 °C. The mixture was then reacted for 3 hours. The reaction solution was concentrated to remove the solvent, and the residue was purified by preparative HPLC (acetonitrile / water containing 0.05% formic acid) to give compound A1.7-E (15 mg) as a brown solid.

[0697] LCMS(ESI)[M+H] + :450.0.

[0698] Step 5: Preparation of compound (S)-2-((2-(7-ethyl-7-hydroxy-8,11-dioxo-7,8,11,13-tetrahydro-10H-[1,3]dioxolo[4,5-g]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-14-yl)ethyl)(methyl)amino)-2-oxoethyl acetate (A1.7-F) [ka] Compound A1.7-E (S) 7-Ethyl-7-hydroxy-14-(2-(methylamino)ethyl)-10,13-dihydro-11H-[1,3]dioxolo[4,5-g]pyrano[3',4':6,7]indolizino[1,2-b]quinoline-8,11(7H)-dione (15 mg) and triethylamine (15 mg) were dissolved in dichloromethane (5 ml), and acetyloxyacetyl chloride (20 mg) was added at 0° C. The mixture was reacted for 1 hour. The reaction solution was concentrated to remove the solvent, and the crude product was used directly in the next step without further purification.

[0699] LCMS(ESI) [M5+H]. + =550.1.

[0700] Step 6: Preparation of compound (S)-N-(2-(7-ethyl-7-hydroxy-8,11-dioxo-7,8,11,13-tetrahydro-10H-[1,3]dioxolo[4,5-g]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-14-yl)ethyl)-2-hydroxy-N-methylacetamide (A1.7) [ka] Compound A1.7-F(S)-2-((2-(7-ethyl-7-hydroxy-8,11-dioxo-7,8,11,13-tetrahydro-10H-[1,3]dioxolo[4,5-g]pyrano[3',4':6,7]indolizino[1,2-b]quinolin-14-yl)ethyl)(methyl)amino)-2-oxoethyl acetate (15 mg) was dissolved in ethanol (5 ml), concentrated hydrochloric acid (1.5 ml) was added, and the mixture was reacted at 80 ° C. for 2 hours. The reaction solution was concentrated to remove the solvent, and the residue was purified by preparative HPLC (acetonitrile / water containing 0.05% formic acid) to obtain compound A1.7 (1.6 mg) as a white solid.

[0701] LCMS(ESI)[M+H] + :508.2.

[0702] Example A1.8: Synthesis of (S)—N-(2-(7-ethyl-7-hydroxy-8,11-dioxo-7,8,11,13-tetrahydro-10H-[1,3]dioxolo[4,5-g]pyrano[3′,4′:6,7]indolizino[1,2-b]quinolin-14-yl)ethyl)-2-hydroxy-N-isopropylacetamide (A1.8) [ka] Step 1: Preparation of isopropylbenzyl (3-(6-nitrobenzo[d][1,3]dioxin-5-yl)-3-oxopropyl)carbamate (A1.8-B) [ka] To a solution of compound (A1.8-A) 3-(isopropylamine)-1-(6-nitrobenzo[d][1,3]dioxin-5-yl)propan-1-one (900 mg) in dichloromethane (10 ml), triethylamine (1.8 g) and benzyloxycarbonyl chloride (734 mg, 4.3 mmol) were added sequentially, and the mixture was allowed to react at room temperature for 2 hours. The reaction solution was diluted with water (50 ml) and extracted with dichloromethane (50 ml x 3). The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated to give the crude product, which was purified by column chromatography (petroleum ether:ethyl acetate = 3 / 1) to give the target compound (A1.8-B) (600 mg).

[0703] LCMS(ESI)[M+H] + :414.9, 1 H NMR(400MHz,DMSO-d6)δ7.72(s,1H),7.34(br s,6H),6.31(s,2H),5.08(s,2H),4.14-4.10(m,1H),3.48(d,J=7.9Hz,2H),3.03(s,2H),1.13-1.11(m,6H).

[0704] Step 2: Preparation of compound benzyl (3-(6-aminobenzo[d][1,3]dioxin-5-yl)-3-oxypropyl)(isopropyl)carbamate (A1.8-C) [ka] Compound (A1.8-B) isopropylbenzyl (3-(6-nitrobenzo[d][1,3]dioxin-5-yl)-3-oxopropyl)carbamate (200 mg, 0.48 mmol) was dissolved in a mixture of saturated ammonium chloride (3 ml) and ethanol (3 ml), and then iron powder (135 mg) was added to the reaction solution. The reaction mixture was stirred at room temperature for 2 hours. The reaction solution was filtered, and the filtrate was concentrated under reduced pressure to obtain a solid. The crude product was purified by column chromatography (petroleum ether:ethyl acetate = 3 / 1) to obtain the target compound (A1.8-C) (65 mg).

[0705] LCMS(ESI)[M+H] + :395.2.

[0706] Step 3: Preparation of (S) 7-ethyl-7-hydroxy-14-(2-(isopropylamine)ethyl)-10,13-dihydro-11H-[1,3]dioxolo[4,5-g]pyrano[3',4':6,7]indolizino[1,2-b]quinoline-8,11(7H)-dione (A1.8-D) [ka] At room temperature, compound (A1.8-C) benzyl (3-(6-aminobenzo[d][1,3]dioxin-5-yl)-3-oxopropyl)(isopropyl)carbamate (65 mg), compound (S)-4-ethyl-4-hydroxy-7,8-dihydro-1H-pyrano[3,4-f]indolizine-3,6,10(4H)-trione (45 mg), an...

Claims

1. Ligand Drug Conjugates of Formula I 【Chemistry 1】 or a pharma- ceutically acceptable salt thereof, During the ceremony, Tb is an antibody or antigen-binding fragment thereof, or a targeting moiety that binds to a target; q is the drug-to-ligand coupling ratio, where q is selected from any number between 0.1 and 16.0; R １ , R ２ each independently represents H, halogen, OH, C １－４ alkyl, or R １ and R ２ together with the carbon atoms to which they are attached form a 5-6 membered heterocyclic ring containing 1, 2 or 3 O, S, or N, or any combination thereof; R ３ But, H, C １－４ alkyl, or R ３ and X together with the carbon atom to which they are attached form a 5- to 6-membered carbocyclic ring; W is -O-, -S-, -NR ４ - and 【Chemistry 2】 Position 1 is bonded to X and position 2 is selected from L ４ Or L ３ Binds to R ４ , R ５ Each independently represents H, C １－４ Alkyl and C ３－６ cycloalkyl; n is selected from 1, 2 and 3; X, 【Chemistry 3】 wherein position 1 is attached to the parent ring and position 2 is attached to W; L １ but, 【Chemistry 4】 and wherein position 1 is bonded to Tb via an S atom and position 2 is selected from L ２ Or L ３ Binds to L ２ is not present or is present, and L ２ If there is L ２ but, 【Chemistry 5】 and the first position is selected from L １ and the 2-position is L ３ Binds to L ３ But Val-AA １ -Gly, AA １ The amino acid residue represented by 【Chemistry 6】 is selected from L ４ is not present or is present, and L ４ If there is, L4 is 【Chemistry 7】 And the first place is L ３ and position 2 is bonded to D; A ligand-drug conjugate or a pharma- ceutically acceptable salt thereof, comprising:

2. 2. The ligand drug conjugate of claim 1, or a pharma- ceutically acceptable salt thereof, wherein q is selected from any number from 2 to 8.

3. L １ but, 【Chemistry 8】 and wherein position 1 is bonded to Tb via an S atom and position 2 is selected from L ２ Or L ３ 2. The ligand drug conjugate of claim 1, or a pharma- ceutically acceptable salt thereof, wherein the ligand drug conjugate is bound to

4. L ２ The ligand drug conjugate or pharma- ceutically acceptable salt thereof of claim 1, wherein:

5. A.A. １ The amino acid residue represented by 【Chemistry 9】 The ligand drug conjugate or a pharma- ceutically acceptable salt thereof according to claim 1, characterized in that it is selected from:

6. A.A. １ The amino acid residue represented by 【Chemistry 10】 The ligand drug conjugate or its pharma- ceutically acceptable salt according to claim 5, characterized in that it is selected from the following:

7. L ３ but, 【Chemistry 11】 and the first position is selected from L １ Or L ２ and the 2-position is L ４ or D. The ligand drug conjugate or a pharma- ceutically acceptable salt thereof according to any one of claims 1 to 6, characterized in that:

8. L ３ but, 【Chemistry 12】 And the first place is L １ Or L ２ and the 2-position is L ４ or D. The ligand drug conjugate or a pharma- ceutically acceptable salt thereof according to claim 7, wherein:

9. L4, 【Chemistry 13】 and the first position is selected from L ３ and position 2 is bonded to D, or a pharma- ceutically acceptable salt thereof, according to any one of claims 1 to 8.

10. L ４ The ligand drug conjugate or pharma- ceutically acceptable salt thereof according to any one of claims 1 to 9, characterized in that:

11.

14. But the following structural fragment: 【Table 1】 The ligand drug conjugate or a pharma- ceutically acceptable salt thereof according to claim 7, characterized in that it is selected from

12.

15. has the following structure: 【Table 2-1】 【Table 2-2】 and wherein position 1 is bonded to Tb and position 2 is bonded to D, or a pharma- ceutically acceptable salt thereof.

13.

16. has the following structure: 【Table 3】 13. The ligand drug conjugate of claim 12, or a pharma- ceutically acceptable salt thereof, wherein: the ligand drug conjugate is selected from the group consisting of:

14. R １ is selected from H and halogen; R ２ But H and C １－４ alkyl, or R １ and R ２ together with the carbon atoms to which they are attached, 【Chemistry 17】 where the dotted line indicates where the heterocycle is fused to the benzene ring, R ３ is H or R ３ and X together with the carbon atoms to which they are attached 【Chemistry 18】 where the dotted lines indicate where the heterocycle is fused to the benzene and pyridine rings. The ligand drug conjugate of claim 13, or a pharma- ceutically acceptable salt thereof.

15. R １ is H or F, R ２ is H or methyl, or R １ and R ２ together with the carbon atoms to which they are attached, 【Chemistry 19】 15. The ligand drug conjugate of claim 14, or a pharma- ceutically acceptable salt thereof, wherein the dotted line indicates where the heterocycle is fused to the benzene ring.

16. R ３ The ligand drug conjugate of claim 14, or a pharma- ceutically acceptable salt thereof, wherein:

17. W is -NR ４ 15. The ligand drug conjugate of claim 14, wherein said ligand is selected from - and -O-, or a pharma- ceutically acceptable salt thereof.

18. R ４ Each independently represents H, C １－４ Alkyl and C ３－６ cycloalkyl; R ５ A ligand drug conjugate according to any one of claims 1 to 17, characterized in that: is H and n is 1; or a pharma- ceutically acceptable salt thereof.

19. X, 【Chemistry 20】 and wherein position 1 is attached to the parent ring and position 2 is attached to W, or a pharma- ceutically acceptable salt thereof.

20. X, 【Chemistry 21】 20. The ligand drug conjugate of claim 19, wherein:

21. D. 【Chemistry 22】 wherein the 1-position is L ４ Binds to W is -O-, -S- and 【Chemistry 23】 Position 1 is bonded to X and position 2 is selected from L ４ Binds to X, 【Chemistry 24】 wherein position 1 is attached to the parent ring and position 2 is attached to W; R １ , R ２ , R ３ , R ４ , R ５ and n is as defined in any one of claims 1 to 16. The ligand drug conjugate according to any one of claims 1 to 16, or a pharma- ceutically acceptable salt thereof.

22.

25. has the following structure: 【Chemistry 26-1】 【Chemistry 26-2】 【Chemistry 26-3】 and the first position is selected from L ４ Binds to L ４ If there is no ３ 22. The ligand drug conjugate or a pharma- ceutically acceptable salt thereof according to any one of claims 1 to 21, characterized in that it is bound to

23.

27. has the following structure: 【Chemistry 28】 and the first position is selected from L ４ Binds to L ４ If there is no ３ 23. The ligand drug conjugate of claim 22, or a pharma- ceutically acceptable salt thereof, wherein the ligand drug conjugate is bound to

24. Compounds represented by formula II 【Chemical Formula 29】 or a pharma- ceutically acceptable salt thereof, wherein the compound represented by formula II is 【Chemistry 30】 A ligand drug conjugate or a pharma- ceutically acceptable salt thereof, characterized in that it is selected from any one of the following:

25. 24. The ligand drug conjugate of any one of claims 1 to 23, or a pharma- ceutically acceptable salt thereof, wherein said Tb comprises a Fab, Fab', F(ab')2, Fd, Fv, dAb, a complementarity determining region fragment, a non-human antibody, a humanized antibody, a chimeric antibody, a fully human antibody, a probody, a monoclonal antibody, a bispecific antibody, or a multispecific antibody.

26. 24. The ligand drug conjugate or a pharma- ceutically acceptable salt thereof of any one of claims 1 to 23, wherein said Tb is selected from a full length antibody.

27. 27. The ligand drug conjugate or a pharma- ceutically acceptable salt thereof according to any one of claims 1 to 26, wherein said Tb is a monoclonal antibody.

28. 28. The ligand drug conjugate of any one of claims 1 to 27, or a pharma- ceutically acceptable salt thereof, wherein Tb is an anti-B7H3 antibody or an antigen-binding fragment thereof, an anti-Trop-2 antibody or an antigen-binding fragment thereof, an anti-Her2 antibody or an antigen-binding fragment thereof, an anti-EGFR antibody or an antigen-binding fragment thereof, an anti-Her3 antibody or an antigen-binding fragment thereof, or an IgG1 isotype antibody anti-chicken lysozyme antibody.

29. Tb is one of the following: (i) Tb is an anti-Her2 antibody or an antigen-binding fragment thereof, and the anti-Her2 antibody is ambenitamab, coprelotamab, dicitamab, gancotamab, margetuximab, pertuzumab, timigituzumab, zanidatamab, trastuzumab or pertuzumab; (ii) Tb is an anti-Trop-2 antibody or an antigen-binding fragment thereof, and the anti-Trop-2 antibody is datopotamab or sacituzumab; (iii) Tb is an anti-B7H3 antibody or an antigen-binding fragment thereof, and the anti-B7H3 antibody is an enoblitzumab, mirzotamab, or omburtamab antibody; (iv) Tb is an anti-EGFR antibody or an antigen-binding fragment thereof, and the anti-EGFR antibody is demvitamab, depatucizumab, futuximab, imatuzumab, lapritucimab, rosatucizumab, matuzumab, modotucimab, necitumumab, nimotuzumab, panitumumab, pimultamab, cercultamab, tomzotucimab, zalutumumab, or cetuximab; 30. The ligand drug conjugate of claim 28, or a pharma- ceutically acceptable salt thereof, which is any one of the embodiments of

30. wherein the Tb is an anti-B7H3 antibody or an antigen-binding fragment thereof, the anti-B7H3 antibody comprising the complementarity determining regions (CDRs) shown below, the CDRs being numbered according to the Kabat numbering system: (a) an HCDR1 consisting of the sequence of SEQ ID NO: 7, an HCDR2 consisting of the sequence of SEQ ID NO: 8, an HCDR3 consisting of the sequence of SEQ ID NO: 9, and / or LCDR1 consisting of the sequence of SEQ ID NO: 17, LCDR2 consisting of the sequence of SEQ ID NO: 18, LCDR3 consisting of the sequence of SEQ ID NO: 19, (b) an HCDR1 consisting of the sequence of SEQ ID NO: 27, an HCDR2 consisting of the sequence of SEQ ID NO: 28, an HCDR3 consisting of the sequence of SEQ ID NO: 29, and / or LCDR1 consisting of the sequence of SEQ ID NO: 37, LCDR2 consisting of the sequence of SEQ ID NO: 38, LCDR3 consisting of the sequence of SEQ ID NO: 39, (c) an HCDR1 consisting of the sequence of SEQ ID NO: 7, an HCDR2 consisting of the sequence of SEQ ID NO: 8, an HCDR3 consisting of the sequence of SEQ ID NO: 9, and / or LCDR1 consisting of the sequence of SEQ ID NO: 37, LCDR2 consisting of the sequence of SEQ ID NO: 38, LCDR3 consisting of the sequence of SEQ ID NO: 39, or (d) an HCDR1 consisting of the sequence of SEQ ID NO: 27, an HCDR2 consisting of the sequence of SEQ ID NO: 28, an HCDR3 consisting of the sequence of SEQ ID NO: 29, and / or LCDR1 consisting of the sequence of SEQ ID NO: 17, LCDR2 consisting of the sequence of SEQ ID NO: 18, and LCDR3 consisting of the sequence of SEQ ID NO: 19; 2. The ligand drug conjugate of claim 1, wherein said ligand drug conjugate is defined by:

31. wherein the Tb is an anti-B7H3 antibody or an antigen-binding fragment thereof, the anti-B7H3 antibody comprising the complementarity determining regions (CDRs) shown below, the CDRs being numbered according to the Kabat numbering system: (a) an HCDR1 consisting of the sequence of SEQ ID NO: 7, an HCDR2 consisting of the sequence of SEQ ID NO: 8, and an HCDR3 consisting of the sequence of SEQ ID NO: 9; and LCDR1 consisting of the sequence of SEQ ID NO: 17, LCDR2 consisting of the sequence of SEQ ID NO: 18, LCDR3 consisting of the sequence of SEQ ID NO: 19, (b) an HCDR1 consisting of the sequence of SEQ ID NO: 27, an HCDR2 consisting of the sequence of SEQ ID NO: 28, and an HCDR3 consisting of the sequence of SEQ ID NO: 29; and LCDR1 consisting of the sequence of SEQ ID NO: 37, LCDR2 consisting of the sequence of SEQ ID NO: 38, LCDR3 consisting of the sequence of SEQ ID NO: 39, (c) an HCDR1 consisting of the sequence of SEQ ID NO: 7, an HCDR2 consisting of the sequence of SEQ ID NO: 8, and an HCDR3 consisting of the sequence of SEQ ID NO: 9; and LCDR1 consisting of the sequence of SEQ ID NO: 37, LCDR2 consisting of the sequence of SEQ ID NO: 38, LCDR3 consisting of the sequence of SEQ ID NO: 39, or (d) an HCDR1 consisting of the sequence of SEQ ID NO: 27, an HCDR2 consisting of the sequence of SEQ ID NO: 28, and an HCDR3 consisting of the sequence of SEQ ID NO: 29; and LCDR1 consisting of the sequence of SEQ ID NO: 17, LCDR2 consisting of the sequence of SEQ ID NO: 18, and LCDR3 consisting of the sequence of SEQ ID NO: 19; 2. The ligand drug conjugate of claim 1, wherein said ligand drug conjugate is defined by:

32. 31. The ligand drug conjugate of claim 30, or a pharma- ceutically acceptable salt thereof, wherein the anti-B7H3 antibody comprises a VH comprising the group of HCDRs (a)-(d) and a VL comprising the group of LCDRs (a)-(d).

33. The anti-B7H3 antibody, VH set forth in SEQ ID NO: 3 and / or VL set forth in SEQ ID NO: 13, or VH set forth in SEQ ID NO: 23 and / or VL set forth in SEQ ID NO: 33; 31. The ligand drug conjugate of claim 30, comprising:

34. The anti-B7H3 antibody, VH set forth in SEQ ID NO: 3 and VL set forth in SEQ ID NO: 13; or VH set forth in SEQ ID NO: 23 and VL set forth in SEQ ID NO: 33; 31. The ligand drug conjugate of claim 30, comprising:

35. The anti-B7H3 antibody, (a) a heavy chain constant region CH of a human immunoglobulin or a variant thereof, and / or (b) a human immunoglobulin light chain constant region CL or a variant thereof; 35. The ligand drug conjugate of any one of claims 30 to 34, or a pharma- ceutically acceptable salt thereof, further comprising:

36. 36. The ligand drug conjugate of claim 35, or a pharma- ceutically acceptable salt thereof, wherein the anti-B7H3 antibody comprises CH set forth in SEQ ID NO: 43 or a variant thereof, and / or CL set forth in SEQ ID NO: 44 or a variant thereof.

37. The anti-B7H3 antibody, A heavy chain as set forth in SEQ ID NO: 45, and / or a light chain as set forth in SEQ ID NO: 46, or A heavy chain according to SEQ ID NO: 47, and / or a light chain according to SEQ ID NO: 48, 34. The ligand drug conjugate of claim 33, comprising:

38. 24. The ligand drug conjugate or a pharma- ceutically acceptable salt thereof according to any one of claims 1 to 23, wherein Tb is an anti-chicken lysozyme human IgG1 isotype antibody.

39. The ligand drug conjugate has the following structure: Structure represented by formula I-1: 【Chemistry 31】 In the formula, Tb, L １ , L ２ , L ３ , L ４ , X, R １ , R ２ , R ３ , R ４ and q is as defined in claim 22, or Structure represented by formula I-2: 【Chemistry 32】 In the formula, Tb, L １ , L ２ , L ３ , L ４ , X, R １ , R ２ , R ３ and q is as defined in claim 22, or Structure represented by formula I-3: 【Chemical 33】 In the formula, Tb, L １ , L ２ , L ３ , L ４ , X, R １ , R ２ , R ３ , R ４ , R ５ , n and q are as defined in claim 22; or A structure represented by formula I-1A or I-1B: 【Chemical 34】 In the formula, Tb, L ２ , L ３ , L ４ , X, R １ , R ２ , R ３ , R ４ and q is as defined in any one of claims 1 to 22, or Structure represented by formula I-2A or formula I-2B: 【Chemistry 35】 In the formula, Tb, L ２ , L ３ , L ４ , X, R １ , R ２ , R ３ and q is as defined in any one of claims 1 to 22, or Structure represented by formula I-3A or formula I-3B: 【Chemical 36】 In the formula, Tb, L ２ , L ３ , L ４ , X, R １ , R ２ , R ３ , R ４ and q is as defined in any one of claims 1 to 22, or A structure represented by formula IA or IB: 【Chemical 37】 In the formula, Tb, X, R １ , R ２ , R ３ and q are as defined in claim 22, 【Chemical 38】 is as defined in claim 22, L ３ A.A. in １ and Structure represented by formula I-B: 【Chemical 39】 In the formula, Tb, X, R １ , R ２ , R ３ and q are as defined in claim 22, 【Chemistry 40】 is as defined in claim 22, L ３ A.A. in １ That is, 2. The ligand drug conjugate of claim 1, wherein the ligand drug conjugate is represented by:

40. The ligand drug conjugate is selected from: 【Table 4-1】 【Table 4-2】 【Table 4-3】 【Table 4-4】 【Table 4-5】 【Table 4-6】 【Table 4-7】 【Table 4-8】 【Table 4-9】 wherein Tb and q are as defined in any one of claims 1 to 29. A ligand-drug conjugate or a pharma- ceutically acceptable salt thereof, comprising:

41. The ligand drug conjugate is selected from: 【Chemistry 41-1】 【Chemistry 41-2】 In the formula, Tb １ is an anti-B7H3 antibody, and q is selected from any number between 0.1 and 16.0; The ligand drug conjugate of claim 1 or a pharma- ceutically acceptable salt thereof.

42. The Tb １ 42. The ligand drug conjugate of claim 41, or a pharma- ceutically acceptable salt thereof, characterized in that: is an enoblituzumab, mirzotamab, or omburtamab antibody.

43. The ligand drug conjugate is selected from: 【Chemistry 42】 In the formula, Tb ２ is an anti-Trop-2 antibody, and q is selected from any number between 0.1 and 16.0; The ligand drug conjugate of claim 1 or a pharma- ceutically acceptable salt thereof.

44. The Tb ２ 44. The ligand drug conjugate of claim 43, or a pharma- ceutically acceptable salt thereof, wherein is datopotamab or sacituzumab.

45. The ligand drug conjugate is selected from: 【Chemistry 43】 In the formula, Tb ３ is an anti-Her2 antibody, and q is selected from any number between 0.1 and 16.0; The ligand drug conjugate of claim 1 or a pharma- ceutically acceptable salt thereof.

46. The Tb ３ 46. ​​The ligand drug conjugate of claim 45, or a pharma- ceutically acceptable salt thereof, wherein is ambenitamab, coprelotamab, dicitamab, gancotamab, margetuximab, pertuzumab, timigituzumab, zanidatamab, or trastuzumab.

47. The ligand drug conjugate is selected from: 【Chemistry 44】 In the formula, Tb ５ is an anti-EGFR antibody, and q is selected from any number between 0.1 and 16.0; The ligand drug conjugate of claim 1 or a pharma- ceutically acceptable salt thereof.

48. The Tb ５ 48. The ligand drug conjugate of claim 47, or a pharma- ceutically acceptable salt thereof, wherein is demvitamab, depatucizumab, futuximab, imatuzumab, laplitucinamab, rosatucizumab, matuzumab, modotucimab, necitumumab, nimotuzumab, panitumumab, pimultamab, cercultamab, tomzotucimab, zalutumumab, or cetuximab.

49. The ligand drug conjugate is selected from: 【Chemistry 45】 In the formula, Tb ６ is an antibody or an antigen-binding fragment thereof that has no tumor cell endocytosis activity and no tumor cell surface antigen-binding activity, or an antibody or an antigen-binding fragment thereof that has binding activity to a non-endocytosis antigen, and q is selected from any numerical value of 0.1 to 16.

0. The ligand drug conjugate of claim 1 or a pharma- ceutically acceptable salt thereof.

50. The ligand drug conjugate is selected from: 【Chemistry 46】 In the formula, Tb ７ is an antibody or an antigen-binding fragment thereof that has weak tumor cell endocytosis activity or has no tumor cell endocytosis activity but has tumor cell surface antigen binding activity, and q is selected from any numerical value of 0.1 to 16.

0. The ligand drug conjugate of claim 1 or a pharma- ceutically acceptable salt thereof.

51. The Tb ７ is an antibody or antigen-binding fragment thereof having binding activity to B7H3, GD-2, Trop-2, EGFR, CD19, CD30, GPNMB, CD20, CD79b and BCMA antigens but not endocytosis activity.

52. The ligand drug conjugate is selected from: 【Chemistry 47】 In the formula, Tb ８ is an antibody or an antigen-binding fragment thereof having tumor cell endocytosis activity and tumor cell surface antigen binding activity, and q is selected from any numerical value between 0.1 and 16.

0. The ligand drug conjugate of claim 1 or a pharma- ceutically acceptable salt thereof.

53. The Tb ８ 53. The ligand drug conjugate of claim 52, or a pharma- ceutically acceptable salt thereof, wherein is a sacituzumab, pertuzumab, trastuzumab or cetuximab antibody.

54. A linker in a ligand drug conjugate, the structure of which is as follows: 【Chemistry 48】 Position 1 binds to a ligand or targeting moiety that binds to a target, and position 2 binds to a biologically active molecular fragment; L １ , L ２ , L ３ , L ４ is as defined in any one of claims 1 to 13, Linker.

55. Drug-Linker Conjugate of Formula III 【Chemistry 49】 or a pharma- ceutically acceptable salt thereof, During the ceremony, R １ , R ２ , R ３ , X, W, L １ , L ２ , L ３ and L ４ is as defined in any one of claims 1 to 40, and L １ is bonded to Lg; Lg is a leaving group; A drug linker conjugate of Formula III:

56. Lg is a halogen, a sulfone group, a tertiary amine salt group, a diazonium salt group, -OMs, MeSO ２ - and CF ３ SO ３ or a pharma- ceutically acceptable salt thereof.

57. The drug linker conjugate is selected from: 【Table 5-1】 【Table 5-2】 【Table 5-3】 【Table 5-4】 【Table 5-5】 【Table 5-6】 【Table 5-7】 【Table 5-8】 【Table 5-9】 【Table 5-10】 A drug linker conjugate or a pharma- ceutically acceptable salt thereof, comprising:

58. In a suitable solvent under suitable conditions, Tb and a compound of formula III 【Chemistry 50】 carrying out a coupling reaction of a drug linker conjugate of During the ceremony, Tb is as defined in any one of claims 1 to 53, L １ , L ２ , L ３ , L ４ , R １ , R ２ , R ３ , X, W, Lg are as defined in any one of claims 55 to 57; The coupling reaction is carried out in water and / or an organic solvent. A process for preparing a ligand drug conjugate according to any one of claims 1 to 53.

59. the molar ratio of Tb to the drug linker conjugate is 1:(1-20); The organic solvent is selected from N,N-dimethylformamide, dimethylsulfoxide, N-methylpyrrolidone, acetonitrile, methanol, ethanol, or any combination thereof; 59. The method of claim 58.

60. the Ligand Drug Conjugate has a q value of 2 or more, and the Drug Antibody Ratio (DAR) for the Ligand Drug Conjugate is selected from an integer or decimal number between 1 and 10; Preferably, the Drug to Antibody Ratio (DAR) in said Ligand Drug Conjugate is selected from 1.5 to 2.5, 3.5 to 4.5, 5.5 to 6.5 and 7.5 to 8.5; Preferably, the Drug to Antibody Ratio (DAR) in said Ligand Drug Conjugate is selected from about 2.0, 4.0, 6.0 and 8.0; Preferably, the Drug Antibody Ratio (DAR) in said Ligand Drug Conjugate is selected from: 2, 2.5, 3, 3.5, 4, 4.5, 5, 5.5, 6, 6.5, 7, 7.2, 7.4, 7.5, 7.6, 7.7, 7.8, 7.9, 8.0, 8.1, 8.2, 8.3, 8.4, 8.5, 8.7, 8.9 and 9; A ligand drug conjugate comprising the ligand drug conjugate of any one of claims 1 to 53.

61. A pharmaceutical composition comprising a substance A and optionally one or more pharma- ceutically acceptable adjuvants, said substance A being a ligand drug conjugate according to any one of claims 1 to 53, or a pharma- ceutically acceptable salt thereof, or a ligand drug conjugate according to claim 60.

62. Use of a substance A or a pharmaceutical composition according to claim 61 for preparing a medicament for the treatment and / or prevention of a disease associated with abnormal cellular activity, wherein said substance A is a ligand drug conjugate according to any one of claims 1 to 53, or a pharma- ceutically acceptable salt thereof, or a ligand drug conjugate according to claim 60, and said disease associated with abnormal cellular activity may be a cancer disease; Preferably, the cancer disease is selected from esophageal cancer, brain tumor, lung cancer, squamous cell carcinoma, bladder cancer, stomach cancer, ovarian cancer, peritoneal cancer, pancreatic cancer, breast cancer, head and neck cancer, cervical cancer, endometrial cancer, colon cancer, rectal cancer, colorectal cancer, liver cancer, kidney cancer, urothelial cancer, epidermal cancer, non-Hodgkin's lymphoma, central nervous system tumor, prostate cancer or thyroid cancer, the esophageal cancer is, for example, esophageal adenocarcinoma or esophageal squamous cell carcinoma, the lung cancer is, for example, small cell lung cancer, non-small cell lung cancer or lung adenocarcinoma, the central nervous system tumor is, for example, glioma, glioblastoma multiforme, glioma or sarcoma, the colon cancer is, for example, human colon adenocarcinoma, Preferably, the cancer disease is selected from colon cancer, colorectal cancer, colon adenocarcinoma, lung cancer, breast cancer, prostate cancer, esophageal squamous cell carcinoma; More preferably, the cancer disease is a B7H3-associated cancer disease; More preferably, the cancer disease is a Her3-associated cancer disease; More preferably, the cancer disease is an EGFR-associated cancer disease; More preferably, the cancer disease is a cancer disease associated with Trop-2 or Her2; Most preferably, said cancer disease is breast cancer or said lung cancer is non-small cell lung cancer.

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